How to improve photography through the microscope

Howto lmprovePhotography
Through
theMicroscope
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Howto get the mostout of this booklet
Thisbooklells divdedintofourman sections.
ltemsmarkedwitha a I st the minmuminformationthat newcomers
to the art of photomicrography
needto know,whie llemsleft unmarked
are addressed
to userswhohavealreadytakenphotoswiththe mlcroscope
butare notsatisfied
withthe results.By relerringto sections1 to 3 for picturetakingand section4 for checking,
youwill findthis bfochurea reliablehelpthe nexttimeyouwantto takea photomicrograph.
FrompEparationto observation
lvlicroscopes
suitabletor photomicrography
. . . . . . . . . . . .8
Equipment
suitabletor photomicrography.
. .. . . . . . . . . . .9
Varioustypesot photomicrographic
equipment
a n dp e r f o r m a n c e
...........10
aEquipmentneededfor photomicrography
.. . . . . . . . . . . . 11
Suitablelocalionsfor settingup thiemicroscope
.......12
aObjectivesand photoeyepiecessuitablefor
photomicroOraphy
..........13
aDifferencesin the peripheralimagesdependingon
o b i e c t i vdee s i o n
.. . ... . .14
aDiffeiencesin re-solution
dependingon lne lype
of objective
. . . . . . . . . . . . . . . . 15
asome hintsaboutthe objective.
. . . .. . . . . . . . . . . . . 16- 18
Selection
of a condenser.
.. . . . . . . . .. .. . . . . . . . . . . . . . 19
a o b s e r v a t iporno c e d u r. e. .s. . . . . . . . . . . . . . . . . . . . . m - 2 1
a H a n d l ionf g
s p e c i m e. .n. s. . . . . . . . . . . . . .. . . . . . . . . .. 2 2
Useof the fieldirisdiaphragm
. . . . . . . . . . . . . . . . . . . . . .23
lJseof the apertureiris diaphragmand its effects . .24 - 25
aBasic focusingmethods
. . . . . . . . . . . . . . . .. . . . . . . .6 - 27
How to spot dirt and specksof dust in the optical
systemsof the microscopeand the Ohotomicrographa
i cl t a c h m e n t
.........28
aHow to cleanthe microscopeframe . . . . . . . . . . . . . . . . . 29
a H o wt o c l e a n
t h eo p l i c aslv s t e m . . . . . . . . . . . . . . . . . 3 0 3
-l
a H o w t o c l e a na n o i i . i m m e r s ioobnj e c t i v e..... . . . . . . . . . 3 2
aHow to cleanspecimens. . . . . . . . . . . . . . . . . . . . . . . . . . .33
Sectionl
FrompEpaaationto
obseryation
Sectlon2
Whenlakingpictur€s
in photomicrography
SectionS
Howto obtain
good pdnls
In the caseof poorphotos
How to obtain good prints
How to obtaingood color prints . . . . . . . . . . . . . . . . .60- 61
p nts . . . . . . . . . . . . . . . . . . . . .62
aPreparingblack-and.white
N l a r kosn t h ep h o t o
...........63
H o wl o a v o i dm a r k sd u r i n g
d e v e l o p m e. n
. .t. . . . . . . 6 4 - 6 5
$ection4
Trcuble€hooting
When taking picturesin photomicrography
tr:oJclion ol eouiDmenisuilable lor
GE-Jal on andphoiomicrcgraphy,
basic
lti.€dge,
observaiion procedures,
cE rc methods,etc. are explainedin
.esylsundeGlandmanner.
t"*'**
l*,
l-r"=
lt<r,
and exprafanon
or
p'oceduresneededfof pholG
drfferences
in filmsandpurchase
can ruin lhe best
+ct processing
' This seciiondealswilh film de
nl, howlo ordercolorPrlnls,
elc.
Basicinlomalion in photomicrography
Operatinginstructionsfor modelsP[,4-1oAD
and
P[,4IoADS(with35mmcameraback) . . . . . . . . . . . 36- 37
M a g n i f i c a toi fopnh o t o g r a pehqi cu i p m e.n. .i . . . . . . . . . . 3 8
Differencesin resolutionaccordingto the combinaiions
o f o b i e c t i va en sd e y e p i e .c.e. .s. . . . . . . . . . . . . . . . . . 3 8
Settingot photographicmagnification(effective
m a g n i f i c a .t .i .o.n. .) .
..............39
F r a m i on fgt h es p e c i m .e. n. . . . . . . . . .- . . . . . . . . . . . . 3 9
C h e c k i nogf t h ef i n i s h epdh o t o m i c r o g r a p h s . . . . . . . . 4 0 - 4 1
Pholomicrography
techniques
E x p o s uar d
e j u s t m el en ct h n i q u e. .s. . . . . . . . . . . . . . 4 2 - 4 3
..........44-45
U s eo f t h eA El o c k
C o m p e n s a t ifnogr a f i l m ' sr e c i p r o c if tayi l u r e . . . . . . . . . . . 4 6
color photomicrography
...........41
O C o l of ri l m
O T v D eosf f i l t e r s
.............48
6 Differences
in colorrendition
depending
on the
of tilm and lightsource. . . . . . . . . . . . . . . 49
combination
E Differences
in colorreproduclion
depending
on differencesin color temperature. . . . . . . . . . . . . . . . 50
on lhe lype
a Diflerences
in colorrendition
depending
........5'1
o ff i l m. . . . . . . . . . . . .
........52
a P u r c h aos fec o l ofri l m. . . . . . . . . .
(cc) filterswhentaking
Useof color-compensating
.....-.........53
c o l opr h o l o s
.............54
a T e sp
l hotography
phologEphy
Black-and.white
55
a Black'and-while
tilrn .
56
aComparisonof ditterentfilm brands . . . . . . . . . . . . . . . . .56
57
4 F i | t e .r. s. . . . . . . . . . . . . . . .
........58
P h o t o g r a pwh iyt hP o l a r o if di l m . . .
Trouble-shooting
ranging linishedphotomicrcgnphs
youcan
Cngto theircharacteristics
:-€ causesfor iailureand take core action. In many instances,the
shoollngseclion can also serve
Problemsin tinishedphotosand thok co.reclion
. . . . . . . . . . . . . . . . . . . . . . . .68-71
Poorcolorreproduction
. . . . . . . . . . . . . . . . . . . . .72-73
Blurredimage
i s i n l o c u sb u tn o ts h a r p . . . . . . . . . . . . . . 7 4 ' 7 9
T h ei m a g e
imaqeappeared
Oojeclsolherrha" the specimen
...............80-81
o n t h ei i l m .
. . . . . . . . . . . 82- 83
lJnevenbrightness
Sectionl
From pr€paralionto obseryation
lvlicroscopes
suitable
for photomicrography
.. ... .. .. .. .8
Equipment
suitable
fo. photomicrography..
... .. .. .. .. .9
Varioustypesof photomicrographic
equipment
a n do e r f o r m a n c e
...........10
aEquipmentneededfor photomicrography
. . . . . . . . . . . . . 1'l
Suitablelocations
for settingup the microscope
. . . . . . .12
aobjectivesandphotoeyep;eces
suitablefor
photomlcrography
..........13
in the peripheral
lmagesdepending
aDiff€rences
on
o b j e c t idvees i g n
............14
in resolution
depending
on the type
aDifferences
of obiective
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
a s o m eh i n t sa b o utth eo b i e c t i v e. . . . . . . . . . . . . . . . 1 6 - 1 8
S e l e c t ioof n
a c o n d e n .s.e. .r . . . . . . . . . . - . . . . . . . . . . . . 1 9
a o b s e r v a t iporno c e d u r.e. .s. . . . . . . . . . . . . . . . . . . . . n - 2 1
s p e c i m e. .n. s. . . . . . . . . . . . . .. . . . . . . . . . . 2
a H a n d l ionfg
Useof the fiefdirisdiaphragm
. . . . . . . . . . . . . . . . . . . . . .23
useof the aperture
irisdiaphragm
andits eftects..24-25
a Baslcfocusing
methods
. . . . . . . . . . . . . . . . .... .. ..26-27
Howto spotdirt andspecksof dustin the optical
systemsof the rnicroscope
andthe photomicrc
g r a p h iact t a c h m e n t
,,,......m
frame. . . . . . . . . . . . . . . . .29
aHow to cleanthe mlcroscope
aHow to cleanthe opticalsyslem. . . . . . . . . . . . . . . . . 30- 31
objective
. . . . . . . . . . . . . .32
aHow to cleanan oifimmersion
. . . . . . . . . . . . . . . . . . . . . . . . . . .33
aHow to cleanspecimens
o
Microscopes
suitablefor photomicrography
1. Thephotolub€acceptsthe pholomicrc
graphiccameraattachmenl.
2.
-Ihe
image can be focused
throughthe binocul lube.
matching
3. Thedesiredlighlinlensily
tre specimenconditionsc€n be set
by the odical palh seleclor,
D
4. Objectivesshouldhavehigh resolution
andgoodllatness(iat imageall the way
to lhe peripheryol lhe Msualfielo.
5. Theslageis rolalable.
6. Thecondenseris equippedwilh an
aperlurei s diaphragmand cemeiing
devicelor aligning10the oplicala.xis.
z$""[if;ffffi;'lXll""*'ipd-r'h
------
8. Sufiicientlighl intensityis assured.
Lightinlensilyis adjuslable10matchthe
observalionand pholographing
condilions. lJnilormilluminationfrom low
10highmagnific€tion.
9. The microscopeslandis resislant
'The phoio shoE m.del BHS
again$ enemal vibralions.
10.Provision
lor insenion
of filters
o
Equipment
suitablefor photomicrography
1. Thec€rnea attachmenlaccepls35rnm
or large{omatfilms.The35mmcamsra
back is equippedwith automaticfllm
2. Theoplicalpalhcan be changedlo
condilio.rs.
matchphotographic
3. The measuringarca c€n be changed
to matchthe specimen,(lntegraled
metefing
3oor-sfDt meiering1%)
4. The cameraattachmentis fmly
clampedon the microscope,
colortemperalure.
5. Devicefor measuring
6. Widerangeforsettinglso/AsAsensilivity
for reciprociylaw failure
7. Coanpensation
is carriedout automaljcallyfor long
8. E\po€ureadjuslment,matching
sp€cimenconditions,is possible.
9. Manualexposureis possible.
suchas 16mm
10.Specialapplicalions.
cine and35mmlime lap6epholography,
can be performed.
(A specialcorfrcl unil is reqrifedin
11. An exposurelock mechanism{AElock)
is built'in.
I''rne phoroshm
mod€ PM.1oAos.
o
rii
Various
typesof photomicrographic
equipment
andperformance
Typesol pholomicrographic
oquipment
' Concernlng
theioclsingscreenior
olympusoM systemcameras:
TTre
screencanbechanged
to sul dflerenl
(OM.]
uses
2.
4) Forpholo
OM
Olv-3.
OM
The',ocus
ngscreenmLrsl
berelse screenNo 1 12
placedior pholomicrography' ' mcrography
Sh|]lterb !r mayoccufli magnificaton
s
increased
Whenusng40Xand100X
obleclives.
adlustlhe lightinlenstyso llrat
Shutlerb urmayresullwhen40X
lheshltterspeedis 1-2 sec.In thiscase,
and1mXobjeclvesareused'
sometypesof coor fim mayrequnecoor
Pi|nOADS/
OM s€desSLRcame6
E se ol
locuslng
Photographic
quallty
Easeol
op€ralion
Useol large
forfial lilrn
.Manualwinding
oi
35mmlim
.Use of cablefe ease
.Manual windng of filrn
.Use of cable release
.Use of VarmagniFnder lof
rocusng
meter
Useot exposure
(EMM,7)
.tlse ol buill-nexposure
meier
,'i
f -'
E+
r
needed
for photomicrography
Equipment
: st checkthe charl to seeif lhe equipment
-:.uned ior pholomicrography
hasbeencom
:'.iely assembld.fihe lypeslistedbeloware
-€ mostbasicattachmenlsneededfor taking
roto6 ol stainedspecimens
inlransmitt€d
lighl.
. quip49!! !99!ie! !!!!9pgEP!
(m
r :ocusiigmagnifier
I :ocusingTelescope
! 35mmcameraback
. iLrlomalicaeosure body
.
t
|
,
:onneciingcord
lrgeformal cameraback
:dapter for largeJormalcamera
tulomatic exposlrecontrollnil
-rff
df^
P"
U-
B _slruclion
manlals
t :crm lor recordingdata
rr leaningutensilsi
blower,cleaningiquid,
?1SCeaning
lissle
oOQ
ooa
G
6")
t n
U'
Suitable
locations
for
settingupthemicroscope
Ressonsfor unsuitablelocalionsand corectivemeasures
Unsultabl€room
Cons€qugnces
.Localedtoocloseto mechanical
appliarices
or
machinerythatcancauseexlernalMbralions
Blurredimageas a rcsullof vibralions
.P aces in whichthe vibralon ol persons
walkingpasl can be transmitled
Ga
T|Ealm6nl
.Remove lhe microscopeJromthe soufce
of vibralions
.Use a slurdylableas suppod
.Use a vibralion
proollable
Brlghllightlromthewndowprevents
neara wall
correci .Sei up lhe microscop€
.Positionlhe microscope
locusrng
in sucha way
lhatihe overhead
lightiallsin slightty
in
froni oi the microscope.
.Place whereroomlighl entersthe eyepiece
ljghtor flaresare rcproduced
on the .Cover ihe eyepieceswith caps
.Shui oul siraylighlgetlingntotheeyepiece
photo
or thefocusing
lelescope
by changing
lhe
oplicaPalhseleclor
.Use of lhe microscope
neara windo/
.sel lhe microscope
.A dusryand dlrly room
up in anotherroom
B ackspolsare reproduced
on lhe specimen.Cover
.Placenea.a winclow
the wholemicroscopewith a dusf
wheredustcanenier image
fromlhe oulside
Exampleol a suitable room lor photomlcrography
L
suitable
andphotoeyepieces
Objectives
for photomicrography
Obieciives
:.' pholomicrography
cbjechigh'resolulion
la€s wilh flahessall the way10the periphery
r - visualfieldare requircd.Oi the LB(ong:a"e ) objectives,seies S PlanApochromal,
a >an Achromai,and D PlanAchromal,
._c oi lhe short-barrel
objeclives,Plan
,rccchromaland PlanAchromaltypesare
Cor€ct comblnalions ol obioctiYeand ey€pl€c€
with FK/P
Combination
Combination
with NFK
tlpto eyepleces
-e pholoeyepieceis opticallycoanpensated
E .ermil the objectivelo deliverils full
r€.'cmance on the film plane.ll musl be
:..'eclly malchedwilh the objectives.
s PlanApochromat
eeri€s
Shod.bargl Achromal s€ties
Differences
in the peripheral
images
depending
on objective
design
Accordng lo lhe lype 01objectve the per
pheryoi both lhe observedimaqeand the
pholographed
rnagernayappearoul ol locus
This efiecl s causedby lhe pe.formance
characlerslicsoj Achromallype oblecives
Us ng Pan Achromaloblectves however.
wi res! t . a sharpand Tlatmageexlendng
a lhe way 10the perpheryof the Jeld
iL
y!
rhe rierd ,rarnessor a D Pan Achromai objeclive eiiend nq a th€ way lo the pefprrery s superof to
rhe one orovidedbv a D ach/omal 0b _.civa
Difference
in resolution
depending
on the typeof objective
:- .: -: on n lhe centerof the lieldoi borh
_- :cserved
andthepholoqraphed
images
: _. : according
to
lhe
type
of
objectve.
_ - -:c
cassobjeclive
series,S Pan Apo:-:1s. as wel the S PanAch.omal
- :: crovidesuperorresolulion.
Sill'9oT.--tl?
: ; i- Apochromat
40x.NFK33XLD
ligss-ffi
IU*H
Gives srrarp ,esorutor .r lh€ whoe mase. allowino obseflation oi mlnul€ dela s
Somehintsaboutthe objective
It ls importanl lo choos€ en obi€ctiye
sultable ior your sp€cltlc purpo66
For all objectives,properusedependson lhe
specilicpurpose,sometypesrequiringsome
adjuslments.ln orderto tulv rJtiiizethe
objective,you shouldknowthe meaningoi
the variousnumbersand letiersengraved
on lhe objeclive
barrel.
1. Obi€ctlvewlth con€ction collsr
(N.A.) Method ot adiustnent
In objeclives
witha numeicalapenure
above0.6- €xcludingoifimmersionl}pes(1)Setthe scalelo lhe 0.17mmposition,
lhe thicknessoi the coverglassslrongly
ailecls ihe imagequalily.Coverglasslhick(2)Rotate
ihe coreclioncollar2 or 3 gradua.
nessis theoretically
designaled
al 0.l7mm,
llon marks(0.02{.o3mm)in the direction
althoughin actualpraciicethis may vary by
oi 2, and refocus.lf lhe imageis sharper
+0.3rnm-By oplically
compensaling
ior this
thanunder(1),rotatethe collaranolher
thickness
devation,
the correction
collar
2 or 3 graduation
maks in the same
assuresthe best image.
direclionand tocusagain.
(3)lt you are not satisfiedwilh the image
quality,try rotatingthecollarin the opposite
direction
tor 1-2gradualion
marks,r+
focusing
andcomparing
the image.
(4)Gradually
reducethe amountoi rolalionot
the correclioncollar,andtry to iind the op
iimumcondilionby repealinqsteps(2H3).
r-Ihe abo,/eadjustmenlproceduresmusl
be repeatedeveryiime lhe specimen
is changed.
objFljw typ€ and hagntfication
Num€icarapenu,e(N.4.)
2. Obieclhr€wlth lds dlaphragm
Someobiectivesabo\€,lOXareequippedwith
prevenls
an irisdiaphragm.
Thisdiaphragm
direcl lightfrornenteing the objectivein
clarKieldor lransmittedlightlluorescence
observalion.By watchingbothconlrastand
resolutionof the image,lhe apenurecan
be adjusledto the optimumposilion.
la
-r
at
d
aa
raa
aJ
al
I
I
q
-a
D Pranf{X, S PlanADo100X,Nc s Pt.n ADo100x
-r
s malk indicaresrhat no cov€r
: \o-cov€r (NC) objeclive
::cmens wilhoui coverglasssuch as
:-: objectvesbearinqthe mark NC
: :over glassattectsthe imasec arity,
.!h ch is padicuarlyobvous in lhe case
:.tves with a largenumerca aperlure.
':s!1, when observlngspecmens nol
: :C by a cover glassat hqh maqnl
: specia NC oblecliveis used.
. ewingufcoveredspecmensthfough
: .bjecllves, on y poor images w th
'ares and insuficieniresoluloncan
'i,,'
'-,
) \]ruSi
3 ;+:'\)/ \
)'. d"
'
.,.-.
Li
I'
'l
)'r /;l- (
sp@imenw ihouicoverolass NcsPanApoloox
-ous LB s€riesno cov€fobjeclvesi
: rn 40X,NC D P.n FL 60x NC S Pran
: : ! a ^ d N CS P a n r m x d r y
SDecimen
wlhoul cove,s ass S PlanApo r00X
4. Nocovor lmx dry objeclive
High.maqn
l calionI00Xobleclivesare uslaly
oj lhe oll mmersiontype bul ior the nocover
l00X objeclve a dry type is avalabe Using
this objectivelogelherwth other non.
imme6on objeclves,e.g. nGcovef40X
and 60X.obviateslhe needlo pul mmersion
o on llre specmen slide The l00X dry type
lens can aso be usedfor pholomc fography.
bLrlior achevng oplimumpiclurequaily,
the lse oi an oil'immersonobjeclve s
Obletlve lypeand m.gnific.tlon
Numdi€r aperluc (N.A-)
Mehanial llb€ l€nglh
ofthenlcD6cop€
5. Oil.immersionobjectlv€
Useof the oil-innercionobiective
Oiljmmersion
objeclives
havea numerical
apenureabovel-0 and use manulacturerspeciiiedoi belweenthe lront lensol the
(Oilmmersiof
objeclive
andlhe specimen.
objeciivescarry lhe mark "oii .) In orderio
makefull lse ol the resolvingpowerof the
objeclive,il is preterableto useoil also
betweenlhe condenseffront lensandthe
How to apply the oil
(l)Focuson lhe specimen,
wilhthe 10X,
or 40Xobiective
andbringlhe desired
specimendetailin ihe lietd of view
'
Tnlckns
ot lie cdor Olas
(2)Rolalelhe revolvingnosepieceso lhal lhe
oilimmersionobjectiveis pointingtowards
you,andapplyoii io theirontlensol
hg
'Use only oll specilied by the
manutacluret
li youus€old Cargilloil or cedaroll,lhe
obieclivecannotdisplayils lull potentiat,
sincetheir difiaclion coetficienlsdiffer irom
lhe nominalvalue.
li theoil is llnted.it affecls
lhe quallyof colorrcndilion
in colorphotos.
Theretore,il is advisableto useonlymanufac.
lurerspecifiedoil.In parlicularfor fluorescence
examinalion,
useonlylhe iluorescenceiree
oil provided
wilhthelluorescence
microscopes.
clicksinlothe lighipalh.Makesurelhai
lhe objectivelronl lensis tully immersed
ntotheoil on lhe speclmen
slide.
(4)lftheirnageis notin vieq slowyrolalelhe
linetocusing
knoblill it comesintofocus.
I Makesurethattheobjectivefronllensdoes
notgel 1oocloselo the specimen,
since
lhe imagewilldeteriorale
il air bubbtes
gel
inlolhe oil.ll a hazeseemsto coverparls
of lhe image,evenlhoughit is in focus,
swng the nosepiece1 or 2 timesfrornlhe
cljckslopin orderto removebe air bubbles.
(3)Applyoi lo the specimensurfaceand
rotatelhe nosepiece
untillheoii.immersiorl
objective,ils tip likewisecovercdwilh oil,
lf the lmagestill doesnol improve,remove
the eyepieceas illuslraledabove,check
ior air bubbles
by viewing
the backtens
of lhe objective,wip€oii the oil and
reapplyoil.
- - 1..j .i
t.,,,. t. cte.n an a I.
I
l}t
all
-rl
rb
al
-
ObjeclivBtyp. .nd magnltrcatlon
NlmsdcalaperllB {N.AJ
Mehanicallub. l.ngth
ol lh. mlcrosope
Thickn6s ol th€ cd.r glss
(2)Rotale
5. Oiljmm€r$on objeclhr€
the revovingnosepiece
so thallhe
Useof the oil-imnercionobiective
objecliveis pointingtowalds
oil-immercion
you,and appy o I lo lhe lronl lensof
Oijmmerslon
objeclives
havea numerical
lhe objeclive.
aperlure
above1.0andusemanuiaclurer,
speciliedoil belweenthe iront lensoi the
(Ollimmersion
objective
andthe specimen.
objectivescarry the mark 'o l ,) In orderto
makelull lse oi th€ resolvingpowerol the
objeciive,I is preferableio useoil also
belween
lhe condenser
lrontlensandlhe
ob.,
How to apply the oll
(l)Focuson the sp€cimen,
wilhthe 1OX,
or 40Xoblectiveandbing the desired
specimen
delailin the fieldol view
'
'Us€ only oil sp€cifiedby th6
manulaciuler
ll youuseold Cargilloil or cedaroil,the
objeclivecannoldisplayils fu I polential,
sincelheir diffraciioncoeflicienlsditfer lrcm
thenominal
value.lf lheoil s tinled,ii aitecls
the qualityof colorrenditionn colorpholos.
Thereiore,il s acfuisable
to useonlyman!1ac.
turer-specifed
oil,Inpadcularforlluorescence
examination,
useonlythe fluorescenceiree
oil provicled
wilhthefluorescence
micrGcopes.
clicksinlothe lightpalh.Makesurelhai
lhe objectivelronl ens is fully immerced
lntotheoil on lhe specirnen
slide.
(4)lftheirnageis notin vieq slowyrctatelhe
linelocusing
knoblll it comesintofocus.
I Makesurcthaltheobjectivefronltensdoes
notgel too closelo the specimen,
since
the imagewilldeteriorale
get
il air bubbtes
inlo the oil. lf a hazeseemsto coverparls
of lhe image,eventhoughit is in focus,
siwingthe nosepiece1 or 2 limesl.om lhe
clickslopin orderto removetheair bubbtes.
(3)Apply
oil10the specimen
surfaceand
rolatethenosepiece
untillheoiljmrnerson
objective,ils lip likewiseco\€redwilh oi,
li lhe image slill does not improve, remove
lhe eyepiece as illlstrated above, check
for air blbb es by viewinglhe back lens
ol the objeclive, wipe off the oil and
reapplyoil.
.i
.
,). I i. ./.;.
.i. oii.
--t.: ,. a..e 32 l
:ta
! -
!ta
q
'I
Selection
of a condenser
:rrpcse ol lhe condensers to eficenlly
'le ighl emanalng lrom the lighl
! on lhe specjnren.1rrcreale ightn!
i :.is malchng the oblecliveand lhls
r. de a be11er
image Dependng on the
: . a a b e . P a d c u l a r yw i l h L r l r r oa w m a g
.n obleclvessuch as lX 2X and 4X.
: n s i k e u n e v e ni u m i n a l o na n d i n s u l
f i c e n ta m o ! n lo f r g h l a 1 l h ep e r p h e r ya r e
key 1oocclr Thereloremake slre lo lse
lheseobjeclles n combnalon wtir lhe
! tra ow magnificatoncondenser
n orde. 1r]oblan betterphotoswtir the
S Pan FL 2X objectve.lse oi tire Lrtra ow
nragnilicalion
condenserBH2ULC s
:nbination of BH2 series condensersand LA obiectives
Condenser
:.r2aAc
:-2 SC
:-2 CA
:-2 ULC
procedures
Observation
Now we are Ina y gettng to observalion
bul firsl makesure that no dust or d ri s
on ihe obleclive,eyepiece,and specimen.
lvake t a habt lo check ior dirt beforeyoLl
Lrsethe mcroscope sinced rt prevenls
locusingand resulls n poor imagequaily
ol filters
1 Placement
Turf on lhe ma n swtch and adluslthe
voilagelo poston Pholo(ca 9V)
t Adiustmentof interyupillary
r,-
ob .
C
tt'@
I 3
r4 dislance
h^
a
=-1.
(3)Placethe specimenon the stageand focus
wth a lox objective
@_
I
( 1 ) P a c el h e i g h tb a a n c i n gf i l l e r( L B D . 2 N )
of the lighl ext wlndow.
(4)Adl!sllhe nterplpilarydslance |]nl boU.
leil and righl vew fieds mefge nio one.
(2)Pacean ND i ter in the slot close1olhe
amp housng Dependlng
on the objectve
r.agniiicationand the densly oi the
specmen.Lrselhis fltef so thal t provides
enoughbrghlnessfor easyexa..inaton.
{ Condensercentering
Diopteradlusiment
rc Jsllhe dioplerlo sul lhe observels
-,.esighl.
Ihe melhodd flersaccording
10
I rnlessthe diopieris adjusted,parfocalily
.r I nol be maintained' hen lhe objeclive
the
: r;_enusingWHKlox eyepieces,
.,:-s is adjusled
withlhe focusng knobs
..
^-€e observinglhrcughthe .ighl eyepEce
aiopleradjuslmeniing 6) on the letl
'd): s thenadjustedlor maximum
image
;'f
lor the lefteye.
I
I
(8)Alterlocuslng
on lhe specimen
withlhe
(7)Alrhattime,adjusllhe irnageso lhalthe
I 0X objective,rotatethe lie d slop A in
crossinesn lhecenterof lhelramemask
lhe d rectionof lhe arrowandreducethe
as twoseparaie
areclearydslinguished
fieldirisdiaphragm
d ameterio a minimum.
ines.Thenadjuslthe focusby roiating
Then
move
the
condenser
lromlop
slowly
cross
knob
so
that
the
linefocuslng
the
helghl
to bonomby usingthe condenser
imagearein Tocus
Inesandlhe specimen
posilion
knob
and
stop
at
a
adjuslnrenl
ghl_eye
I
mullaneously.
Alter
compleiing
s
wheretherieldiis diaph€gmir.age
alsoadjusllhe diopterlor lhe
adlustmeni,
is sharplydeiined.
(5).
as
n
by
rolating
secllon
lefl eye
A
mage1olhe
I Movethefieldirisdiaphragm
cenierof thevsuallied wilhlhe con'
knobsc .
densercenlerlng
@@a
I S ncethe \,pe of the lindereyepieceditlers
(g)Open
accoding lo lhe sizeoJlhe film used,
thelieldiis diaphragm
imagelntil
chooselhe typesuitablefor yolr patticular
it almosllouchesthe peripheryol thevislal
iield,andmakesomelinalcenlering
Fornormalobservation
adjuslmenls.
make
conditions,
thediaphlagm
slighly
EwlrKlox
largerlhanlhe visualfield.
E wFrhlox
I lfthefieldirisdiaphragm
cannotbe sharply
A'pholomicrG
us
ng
iinder
eyepiece
tor
a
focused,checkthe thicknessof the spec|
a
E wHkiox4 1 / 4 x 3 1 / 4 " P oa r o i d o
is madeby rotaling
menslide,Useslideswilh a lhickness
t=.y adjuslment
@wNrro{
-: :r B.of lheeyepiece
al therighteye.
belween
0.9and1.2mm.
I
-a
Handling
of specimens
Make I a habLltoc eantirespecmenreg! ary
bolh beforeand afler obsetualionJlsl as
wlh lenses 1is mosl mportanllo work wilh
a c ean specmen Nlakesure thal no dlsl
pairces stck lo the specimenwhen you
store I and do not toLrchthe gass slrlaces
H o l dl h e A l a s ss l i d e a l l h e c o r n e r o ny , o r h o d l h € g a s s s l i d €b e l w e e ny o u r r i n 0 e . s
, /
F i n o e @ a n l s o rnh e . o v e rg a s s
t ,
Noteson specimenplacementon ihe stage
when you pace a sp€cimenon the staqe
the specmen irolderis openedwide and i
t s re easedrapdly.i1wl h t the edgeoj the
slrdeand damagethe g ass.Aner coni rm ng
lhatlhe sp€cimenhasbe€nlirmlyput n pace
llre sprng loadedspecmen holde...u$ be
retractedsowLyso thal il genly lolches lhe
n casethe s de is damagedcarefuy
renrovethe tny glassffagmenls.I fragments
are eil on lhe stage.lheymay calse inilres.
or the specimens de may be paced n a
t t€d poston on lhe stage.causng one sde
o i t h e v i s l a i e l d t o b e o L roi i ' c c ! s
i .
T h e i m a a ei s b u i i e d a o n s t h e l r a c € s o l y o u .
I n9elpnnls
o n es d e o i r h ev s u a r { i e ri ds b
Useof the fieldirisdiaphragm
(with35mmJilm)
ljsing the Plvll0AD tunet masklocusingtelescope
. ' e d l r l sd a p h r a g m
s e v e s1 r ra o l u stlh e
- "ated areaon the specmendependn!
'',e obleclveoowe. lh s daohragmpays
r . a r o e d u rn 9 D h o i o nc r o l r a p h ya n d i
._:. ode thannecessaryr uninal n! gh1
. ecred..d scalteredrre_o!
a ly on the
: ren eslLlrngin a ossoi mageconlrast
: : i g d o w nt h el e d d l a p n r a g 1
m0l ! s 1
:1d the irame rel c e areav/i resu1 n
: rlreph c mageswih rmprovedconlrasi
s stopDe.jdo!,/n
:re ted rs dapl-rra_om
: c ose io the irame retc e tire ohoto
:!red ma_oe
may be cLrtai tfrecorners
! d a p h r a g n , s h o Li hr de r e T o b
r ee o o e n € d
trly morelhan 1l.ere:c e slro\ls
' ' . . r - ." ,
.l
I
l.
'
,..
: ' :,1 :i:i; 'r
I ' e r d , 3 d a p h f a g mh 3 s b e e n
Useof the aperture
irisdiaphragm
andits effeits
The diaphragmrnounledon the condenser
s ca ed aperturers diaphragmTheiunclion
oi this diaphragmis lo maintainopl mlm
condilionsoi image resolulon.conlrasland
ioca deplhby adluslingthe nurrericalapenure
of the lLrmnalionsyslemdependng on lhe
nur.ericalaperlLreof the objectivein use.
For most specimensopllmur. irnagequajly
s achrevedil lhe apenuredaplrragm s
adjustedlo between60% and 80% ol lhe
How lo adjusl the aperlure ids diaphragm
Thereare lwo methodsoi adlushent:
Pu oul lhe eyepecewith lhe specmen n
locus and then adlusllhe daphfagm!,vhlle
watchng lhe rs al the rear ioca planeoi
the oblecliv€as n pholo(1):usethe gradLra
ton markson tire condenser.as in (2)
0)O r€cl viewng wilh lhe eyepi€ce
-
s
nis darh€ap€r1ure
Reducino
prragmlo 6os0%ot lh€ numeica
apenureor lhe obleclve.
(2)Usnqlhe oraduaroi ma*s on rhe
Samp e Us nOaloxoblecrivewilh
a numercalap€duf€
ol
0.25andreducingll to
3 0 % , i h a q G d uoanr m a l k
on lhe connenser
should
b es e ra r0 2 ( = 0 . 2 s ' 0 3 ) .
I
Example Whsn uslng S Plan Apo 20)( NFK 2.5X
" *" "*n"*iris
draphrasm
I -*o
F€ge resolLilion
if lhe aperture
deleriorales
I
I
I
I
I
I
I
I
I
I
I
I
I
.: d aphraom
is sloppeddownloo much
r'h lheexceplion
ol specialyslainedlhn
.ec rnenslhe diaphragm
shouldnotbe
sDped doBnlowerlhan lo 60% ol lhe
nrencal apedLire
ol Iheobieclive.
I Ar effeclsrmrl lo stoppngdownthe
rDenLrfe
rrisdiaphragm
canbe achieved
:t movinglhe condenser
clownward
bul
:rislendstointerferewith
lhebasicillumina'
:o1 iunclionof lhe condenser
andresults
_ uneven
ilumination.
Thusalwaysuselhe
: aphragmandclonot rnovelhe condenser.
$#ffiffi
Fully op€nedposlilon
\i{
=t
&'/"
Conn4l b enhanced.
Detailsarealso
cr€anyvblble,.nd localdepthls
incr6ased,
r€sullingin an oprimum
g./o
Re.oluriond.r.rioralesas a resull
I
Basicfocusingmethods
Focusing lor obseruation
A lOX obleclve s usedas lhe slandardfor
locusng.thef the objecrlves changedlrom
r(]X lo 4X and rrom rOXto 40X and i!(her
lo 100XDo nolchangeabrupilyiromiowmag
n ficaton (2X and 4X)to h gh (40Xand 100X)
As a resLr
t of lhe imled eye oi the obsetuer
and the largeioca deplhoi low magnilicatan
objectves(2X and 4X) lhe ironl mounll)1
the h gh magniticat
on objectvecouldtouch
the specrrnensurtacewhen the revovng
Focusing lor pholomicrography
Focusng dlring photographys done ether
throughthe locusinglelescopeof the photo
graphlcaltachmenlor lhrolgh the eyeoieces
of the binocLr
ar tube.When locusngthroLrgh
lhe eyepieces.a jnder eyepiecemLrstbe
used Pror to pholomicrography
the fnder
eyepecehas 10be foclsed by meansoTi1s
f o c u s n gl r o n tl e n s .t o m a k ec e a r l yv i s b e
doLrbe cross linesas 1wopara el nes ln
lfre cenlerot the lramingrelice
1
l g
iI
I
- b
AJleriocusng on the specmen $,th lOX
obieclivesel the upper mt ol the coarse
adllslrnentexcursonwillr the preioclsing
Movethe specmendetai 1{)be examlnedto
the centerof lhe vsua leld and increase
maqnilicatonby rolalingthe nos€pece.
1
2
--)F
l-.
i-rl
*r+
/n
a'
"i]".
I
Focusing lelescope ol lhe photographic
For pholoqraphy
the specimenlocus mLrsl
be adllsled wth the same eye wilh wh ch
lle .ross ines were foclsed
{
Er
Finder eyepieceol lhe binocular tube
Focusngthroughthe binocLr
ar tube s
possbrewlh microscopesVanor BHS.BFT
and BtsTU
I
l 3
s;
1. Focusing when using 1x,4 and 4x
obieclives
Foclsingerrorsocc!r qu le irequentywth
low nragnflcaton oblectves Tierelorelse
a l o c u s n gm a g n j e ra n d a d l u $t h e i o c L r s
by iolow ng the procedureslsled beow
shoud
Bui beiorethat the pholographer
adjustdioptera1the cross nes oTthe
J o c u s n gi e e s c o p eo r t h e f n d e re y e p e c e
4r!
2. Focusing when using 10X and 20X
rs achievedby adjustnglhe
FocLrsrng
c r o s sI n e s s o l h a tt h e ya r e c e a r y v i s b l e .
and then rolatng tire lineJocusngknob
Lrnt both lhe cross nes and lhe mage
0 1t h e s p e cm e na r e c l e a r yv i s b e s m !
laneousy.By sLghly mov ng yolr eyes
oi lhe
to a sdes. see i the posnLons
cross inesand the specmen mage'1onol
s h i l f t i r i s s t h e c a s el h e P c 1 ! r es
n focLrs t they do sh it. the tocls rs not
l,^,*
rff)
'rage is s ghlly oul oi iocls aner
. : o j m a g n l c a l i o nl s e o ny t h e f n e
_j knobior refocLrsingn partcLrar
rse the coarselocusngknob for
_! a h gh-magnlflcat
on objectve. s nce
. a dangerlhal the obleclve w rLn
- specmen ana uer oamageo
fs
p r o p e r ya g n e da n d m l s l
w tl-rtire lne jocusn9 knob
3. Focusing when using ,lox and 10{x
Adl!$ the cross nes so tfral lrreyare
c eary v sibe. llren s owy adlustihe locLrs
ot the speclmenimageu71hlhe l ne
locusng knob lnli cross Ines and lmage
oi tl-respec..en are c eary vsDle
a o c a tn 9
: : ' e y e p l e c ei s o r o v d e d w t h
: . n must be firmy nse(ed nto the
r ' t h e e y e p i e csee e v e f l h e p n l s
_-_t rnserted.correcliocLrsngbecomes
: e a n dl h e m a g e w b e o L t o i f o c l s .
Camp the iocusingma!nlfleron the
'oclsing leescope.afd side the top sec
the tocls
ron n or olr therebyreadluslLng
al lhe cross lnes. FocLrsis co(ecl t
bolh the cross lnes and lhe specmen
m a g ea r e c e a r y v s i b e s i m u l a n e o u s y
Basicfocusingmethods
Focusing lor observalion
A lOX obleclve s usedas lhe standardior
locusng.then the objecllves chanled from
rOXlo 4X and irom 10Xlo.10Xand ilnher
lo I 00X Do noi changeabrlpl y iromlow mag
nirical0n(2Xand lxl to hrgh{40Xand t00X)
As a res! t 01llre lmled eye oi lhe observer
and the argeioca depthof ow magnificaton
obleclives(2X and 4X] lhe Tronlmountoi
the highmagniiicalionobleclivecoud louch
lhe specimensLrrlacewhen lhe revolvng
2
1
3
I
I
-''b
,
,l
Focusing lor pholomicrography
Foclsrngdlrng pholographys doneeilher
throughthe iocusng leescopeoi the pholograprrc attachmenlor tlrrouqhthe eyepeces
o t t h e b n o c u a rl u b e W h e nl o c u s f g t h r o l q h
l h e e y e p e c e sa. f n d e re y e p e c em u s tb e
used Priorlo photomcrographylhe inder
eyepecehas 10 be focLrsedby meansof ls
focls ng iront ens to rnakecteartyvrsbte
d o u b ec r o s sl i n e sa s l w o p a r a l e l n e s n
l h e c e n l e ro l t h e l r a m i n gr e l c e
After ioclsing on lhe specimenwith l0X
oblectve.set the lpper mil oJ lhe coarse
adjustmenlexclrson wth lhe preiocusn9
fvlovelhe specr.fendela to be examned 10
lfre cenlero1the vsLra fied and ncrease
maqnlcalion by rolalng the nosepece
1
2
-)E
Ir-'
la
l**r+
il1
:
:
lr
(
----
i
Focusing t€lescope ol the photographic
For pholography
the specimenfocusmLrsl
b e a d l l s l e dw l h t h e s a m ee y ew l h w h i c h
l"e .ross lnes were foclsed
Finder eyepieceol ihe binocular rube
Focusnglhroughthe binoc! ar lube s
possbrevilh microscopesVanox.BHS.BNT
ard BrITU
,
fs
1. Focusing when using 1X, 2X, and 4X
2. Focusing when using 10X and 20x
F o c u s n ge r r o r so c c u rq ! i l e j r e q u e n t y w i l h
ow magni cal on obieclivesTheretoteuse
a focLrsng magnilierand adlu$ the iocls
by fo owingthe procedlres sled beow
should
Blt bejorethal llre pirolographer
adlusldopler al the cross Lnesof the
tocls ng leescopeor lhe fnder eyepece
Focusng s ach evedby adlustnglhe
c r o s sl n e ss o t h a tt h e ya r e c € a r y v s b e .
and lhen ro1alingthe fne ioclsng knob
unt borhthe cross ines afd the mage
o t l h e s p e c l m ea
n r ec e a r y v s b l e s i m u
taneousy By s ghlly moMngyoLrreyes
to a sides see I the posilbnsor the
cross lnesand llre specimenlmagedo noi
shil li th s s lhe case the Pclure s
n iocls li they do shjl. the iocus is nol
-')I
:,:,'"Hll"l
lfn
: male is slghty oul oi jocus after
t e 0 1m a g n i c a l l o nl ,s e o n y t h e l n e
: -! knobior reiocusingIn pancuar
: lse the coarselocusng knob for
. r g a h g l r . m a gfnc a t o no b l e c t v es. n c e
s a dangerthal the oblecl&elvr rln
: specmen and get damaged
-
qt*i-j
fs
b-t
properya gn€dand musl be readlusled
w t h l h e f n e i o c l s i n gk n a b
3. Focusing when using 4OXand 10ox
Adilsi the cross nes so thal lhey are
c eary v s b e thenslowy adlustlhe locLs
ol the specirnenimagewlh the r ne
jocusng knob un1 cross lnes and image
o f t h e s p e cm e na r e c l e a r Yv s b l e
\
r: r eyep€ce s provicied
w lh a localing
_. pln musl be I rm y inserledinto tl-re
::r y nserled.cotrectlocls ngbecomes
: o e . a n d l h em a g e w i b e o l t o f i o c u s
Camp llre iocls ng magnifieron lhe
IocLrsngieescope and s de the lop sec
lon n or o!1.ih€rebyreadjustngthe tocLrs
at lire cross lnes Focus s correcl 1
boththe cross nes and the specmen
i m a g ea r e c e a r l yv i s b e s m L r l a n e o L r s l y .
Howto spotdirtandspecksof dustin the
opticalsystemsof the microscope
and
the photomicrographic
attachment
D ( and dLrstpanc es are sor.el mes noted
dur ng observaliofor phologrpahy.
but pin
pontingthe r exact localionnraybe diiiicuit
Whenphotographing
imponantSpecmens
which cannolbe photographed
again11can
b e r e a l yi r u s l r a l j nigd u s tp a d i c e sa r ev s i b e
on tlre pclLrre.An eliectivemethod s therefore requred thal w hep you deiecl
I S i n c e dr l a t o c a t o n sn o r _ a n di m a y c o m e
irom paniciesoj lhe n m lisei (oadng. j m
transpo().regularchecksard necessary
I lf the aperlLrre
daphragmis.edlced below
30%. dirl and dusl Specksbecornecearly
Obser.
grapny
\frf' oescrirrion
Dlst speckson the pr sm insde the obser'
valionlube.on lhe insideof the objeciive
and on lhe jnsde oi the rn croscopebase
cannob
t e w p € d o J le a s y . i c e a n n g
becomesnecessaryconlactyourOympls
Melhod ol verilication
Rernove
theadapterior large{orrnal
cameras
and check ior dirt by peerng ln lhrouglrthe lop
li you spot dilr unsc.ewand removellre re ay ens
Rotalelhe lop lens eement as you observelhe
z 199!li9
:91 Cameraprism
a
Selto,Timemode,openlhe shulterandpeerin
Eitherremovephotoeyepleceand checkior dusl
rotate
q"lig919l r".9""phgto9lepecein prace,
t andcheckfor movngdustparlces.
-_
r
Oplicalpath
O
Tubeenglh
correcton
O
objectve
cha^ge rfe op_icalpa f 4- le alernarelyooqetu g
lhroughlhe foclsing leescopeor lhe fnder
a
a
Remove
lhe observation
tubeiromthe mcroscope
bodyandchecklhe prisrnsu.faceior fngerprnts
or conlaminalion
-
checkt lor d rt or conlaminalon.
Specmen
_O
!0 Condenser
^
-
; ; *
.'
ce ovF the obectNp roa l\e nos6prale d o
Observelhe specmen and move lt in lhe fied.
li dust is on lhe specmen,il wi also move.
Removelhe condenserlrom the mlcrcscopeand
look for dlir and oi deposits.
Checkthe iilef after removingit lrom the m.rG
scooe oase
Swilchon the lluminalion
andexamnelhe lens
l?
Lghl exil
8
gtass
Frosted
j.i
Co lecior lefs
a
a
Bulb
^
'
-'
check lhe irosledg ass
Aemovelhe amp houslngand checkrhe enses
in lhe colleclorassembly.
Bemovethe bulb trom its sockeland check
sgns of backenng fingerprnts.dirr etc.
frame
Howto cleanthe microscope
:ii _son the rnicro€cope
lramearelirsl
Mslerials us€d lor th€ mlcroscooe
..€d wltha pieceol clolhwetedwitha
i:
amounlof neutraldelergent,andthen
..€a cleanwltha pieceof clolhlhal has
)ir- ,mmerced
ln luke-warm
water Butmake
L-. roi io louchlhe lenssection,rvhile
:er_ ng the microscope
ifame.
\. : usingorgancsolvenls
whichmay
--:ge paslicparls.
::r duslconlamnationthaladheres
10
= _ - pansand is difficultto remove,wipe
r: . pieceof clothor soll tissuepaperthat
-3: :een soakedin a mixlureoi 7 partsether
r: I parlsalcohol,
Keeplhe m xtureaway
+:- claslicpanslo prevenldamage.
E
E
E
Howto cleantheopticalsystem
'p6ora
. 6 oor.ot
. ran . pa. o dt.
.
s. o..o
r e r . o r . -o "
surtacet canformay berenroved
wth a
bower Yo! shoLr
d irowever.make a habl
oi coveringthe mcroscopewth a dlstprool
cover after eacn lse
,- ll:e i\
Cleaning melhod
Tduslspolsonoptca glassessuchas lenses
prisms and i ters are e11unallended.the
dLSrbecomesd ficLri ro renrole and may
calse mold By alwayskeepngoptca glass
s!rfaces clean.yo! avojdma nlenanceprob
ems ard prolonglhe ie of yolr mcroscope
Cleanng oj lhe ens suriacesapp es ony 10
exposedareasoi obleclives.
eyepeces lirlers
and condensersIf lnlerna or malorcean ng
becomesnecessary.pease conlactyoLrr
O ympusN4icroscope
deaer
ls
2
|
I
+'.
'il 'fl 'n
.s.b |i
To preventscratcheson coalingsand oplical
! ass removedin and dlsl lhat stcks 10
the rsuriaceswth an ar gunor b owerbrush
Wrap lhe lens lssLrearoLnda woodenor
bambooslck as ustraled.
)t
5
Requiredlools
-
e
,
-
t
..ii-'
'€
i A r gLrnor bower brlsh
2 c e a n n g m x l u r eo t 7 p a r l se l h e ra n d
3 pai(s acohol.or efs cleannq f! d
3 Ol ps wood sl ck
a Sojl qauze.lens 1ss!e
! Magnryng glass An €yepiececan aso be
lsed n pace ot the magnf e
t
-------"'-
'"
__
Whencleanng largeg ass slr{aceson both
sdes ot an accessorysuch as a j t€r fod
two or three ayersof lens lssle soakedn
t h e c e a nn g m x t u r e h o d t h e a c c e s s o rayl
ils edges and wipe lrom the cenierlowards
llre perpheryas yo! s owly rotale I
Whenc eanng tl-resuriacesoi the condens:
and oJ the iighl exil gass hod a pieceoi
y o l r m d d e a n di n d e r
l e n sl s s u e b e t w e e n
ingers lod I afd $/rapi1arolnd yoLr ind:
ringer Thenhod lhe lissuedownwth yo!
lhlmb wlrie vr'rpnqllre lens sudacescea.
tL, -=t
\
w
--
v
.:, -
YJ
:.:. lhe lens by pulilnga smallamounlol
? : .eaning ilud or cleaningmi*ure on the
. :'a lenstissue.Dlscardeach ens tissue
wipe
Whencleaning
a largelenssurface,
if
fromthecenlerlowardslhe periplrery
T,
*-'l
ti
]- : eanng,examnethelensslrfacewlth
. -.:t. [rng glass.]i colorrcfectedirom
t- -,_ssurfaceooksuneven,
it is an ind.
- :. :nattherearesl I duslspecksand
.
" . ewngihrough
thebonomoi an eye
,oLrcanuseit as a magntying
gass
i.,
Qtt'
Whencleaning
a argelenswilhlenstissue
afoundyourllnger,yoLrshouldwipe
wrapped
in a
Iromlhe centertowardslhe periphery
Aways,usea cean pornori
circuarrnolion.
of the ensilssueas yoLrrotaieyourindex
Aftercleanng,breatheghllyon the ens
surlaceunli lhe wholesurfacehaslurned
whle,thencheckwhelherihe hazedis
appearsunlormly.Spotswherethe haze
d sappearsony slowlyare not yet wiped
Howto cleanan oil-immersion
objective
w pe rhe llonl rens oi llre oblecl've r\1
Cl€an the oil-immersionobieclive dunng
Aner in sh ns obseryalon wth an oi
mmersonobjectve.wel a pad oj cottonwoo
or a pece oi ens lissuewilh a sma amoLrnt
of cean ng mxlure conlanrng7 pans ether
and 3 pans acohol,to femov€ol adherng
lo the objectve S nce an oil 1 m w I oilen
adhere1olhe obieclivefronl ens t sholld
be wipedclean l\/ice after eaclrJSe
.)
!
)
llse only lactory recommendedimmercion
oil and r€moveit atter examination
i the o femainson the objectveior a ong
p e r o do i t r n e . t w l l h a d e n ( e g c e d a r o )
makng t diiiicLrllto removeeven if yoLr
wipe the ens repeatedy Tlre lens may be
l n l h e p r o c e s sU s eo n y s p e c i € d
damaged
oi. and afler]]se wlpelhe o irom the mmef
sion surlaceof the objectve.keepng t
lj you lrequentyuse an oll rlrme|sonobleclive olmay contaminatethe sLrrjaceoi a
dry t/pe oblecl ve *hen you changeoblectves
To preventlhe o irom adherngto llre
obleciivecareiullyrolalethe nosepieceafter
lowerng lhe slage.so thal the o does nol
I lhe mage of a dry lr gh-magnifica|on
obleclve appearsiuzzy.check for o thal
nriglrthaveadhefed10lhe 1ipoTthe
\
Lower rhe slaoe slowly so lhal no oi l
louch lhe rronl rens oi the dry objecr '
I
6
I
r-
,Lt
a
t
Alle. owernslhe sla0e carerury.ola
nosepiece
and crickrh€ dry obleclNe
Howto cleanspecimens
r,:ke il a habl lo cean eachsp€cimen
bolll
:r'cre andaflerobservation.
OlheMise,
dirt
-r dusiihal youlailedto noticedlrng
riervalon mghtappearon llre pholo.
::- cleaning
thespeclmen
a sonc oth,gauze,
:, seceo1lenslissuemaybe usedwiihoul
:.aing liqrid.Bll ii ihe conlaminal
on s
,-.Lrll lo clean,brealheon the specmen
ri:re wipng il.Whenceaninglhespecimen,
*,nts to note duringcleaning
' iemovalof oi as wellas roulnecleaning
:an b€ donemorceasilyil lhe specimen
; removedlrom the stage.
: .ihenusingceaningmxturcor lens
: eaningillid, usea moistened
clolhor
:?aniaglissue.Becerlainnotto apply
:.aessivefluid,as il mayseepunderneath
-. coverglassanddarnage
lhespecimen,
--:re
areMo lypesoi sp€cimens:
thosewiltr
a ::.ergassandlhosewlhoulacovergass,
rJ:- as blood smeaG
.b
+
cleanlncsp€im€ns wilh coverqlass
Cleanlng
specimsnswith coverglassos
?swhencleaning
lefses,wpe ol1lhe
:'3 dirlwilha pieceof lenslssue ighlly
.'enedwilhcleaning
mixlure.Because
: cannolbe complelelyremovedwith
roe repealwipingunlillheol film
Cle6ningspcimens wilhoutcover
tElss6s
::-eing to uncovered
speclmens
cannol
a 3edofl. Youcan, however,removelhe
:. Tmelsnglhe specimen
for 5 to 10
-.s n a xylenebath.Therearecontaners
:'::. horizonlal
andverticairnmerson,
:..o€r selecllon
of whichdepends
on
t/t
cleaninsspecmenswrthoutcov€rorass
@
Section2
Whentaking piclurcs
in photomicrography
and
OperatlnoInstructlonsfor modds PM-1oAD
(with36mmcameraback). . . . . . . . . . .36-37
PM-1oADS
. . . . . . . . . . . . .38
Magnification
of photographlc
equipment
Dlfferencesin re6olutionaccordingto the combinations
of objectivesand ey€pieces. . . . . . . . . . . . . . , . . . . . . . 36
Settingof photographicmagnlllcation(effective
magnification)..................................39
Framiog
of the specimen.... - . . . . . . . . . . . . . . . . . . . . . 39
. . . . . . .,O-41
Checking
ot the linishedphotomicrographs.
E x p o s uar d
e j u s t m et snct h n i q u e. .s. . . . . . . . . . , . , . 4 2 - 4 3
. . . . ., ., .44-45
L,seot theAElock
failure... . . . . . . . .,16
Compensating
for a film'sreclproclty
, , , , . , . . . . . . . . . . . . . 47
Ocoforfilm
.............. ',|8
aTyp€offilters
on the
a Dlfferences
In colorrendition
dgpendlng
of film andlightsource. .. .. . . . . . . . .. .,$
comblnatlon
a Dlfferencesin color reproductlondepending
.. . . . . .. . . . . . ., .50
on differences
in colortemDerature
a Diff€rencesin color r€nditiondependingon the type
ot tilm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .51
O Purchas€
of colorfilm. . .. . . .. .. .- . . . . . . . .. . . . . .... .52
[Jseof colorrompensating(Cc)filt€rs whentakino
color photos
a T e s tp h o t o g r a p h y
. . . . . . . . . . . . . . . 53
.,...........54
...........56
a B l a c k € n d - wfhi li m
te
a c o m o a r i s oonf d i f f e r efnl ltmb r a n d s. . . . . . . . . . . . . . . . . 5 6
. . . . . . . . . . .57
a Filters. . . . . . . . . . .
P h o t o g r awp iht yhP o l a r of i |dm . . . . . . . . . . . . . . . . . . . . . . 5 8
tD
Basicintormation
in photomicrography
formodelsPM-1OAD
Instructions
Operating
(with35mmcameraback)
andPM-10ADS
Loadihe film propody
1 Setingthe lilm lormat
2
Plsh lhe buton ior 35mm camem.
Loadthe ii m nto the cameraand makesure
rhat the end ol the iim does nol protrLrde
beyondthe spoo groove OlheMSe tire
Jramespacingmay not be Lrnlorm.
7 Centerthe specimento be
, photographedin the visuallield
andadjustthe apeiure
R
v Fgcus
msotapnragm
.
r
l
thewinding
ortnerirm
3 confirm
f
Wlnd the fim onlo rlre emptyrakeup spoo
and advancei110irame m. I To conflrm
that rhe fim s winding be cedain thal the
f i m f e w n d n g c r a nsk f o l a t n gt l s n o t .
rolale the crank 2 or 3 tmes n the d recton
oj lhe dotledarow lo pick up lhe slackol
the I m nsde the car.era. and aga n check
lhe rewndng crank to coniirmthe wndng
of lhe i rn
V
Adjustshullsr speed
X
Movelhe area ol llre specmenyo! wanl 10
pholographinlo the cenler of the visua ield
wilh lhe stagecontros.WhenyoLruse mode
PM-1oADS.
cenler rhe area10be pholo.
graphed n the spot mete.ingseciionoTlhe
Focuson lhe specmen and adluslthe
condefrserapenureirls diaphragmso lhal
sutabe cofirasl s achievedThe aperllre
.laphragm s ordinary sel at 60% lo 80%
or the objecuvenLrmercaapedure
Checklh€ shunerspeed.Adluslllre speed
to between001 and 05 secondsby using
ND i itels
4
Color phologEPhy
Set lhe colorlemperalureto malchthe type
of color film used.Measurelhe temperature
at a blankareanot coveredt'v the specimen.
I!,De of fflm
Daylbhi film
Tumsi.n lllm
S€rlhespeedfor lhe lih Lrsed.
Sel lhe characleristicsior reciprocityfailurc
for lhe filmused.
tldrl
Dlal pG!
helarclng thn
CIn
ffllor
t.BD-2N
I-BT
D
T
Thevollageposilionwhennot usingthe color
temperaturemetercTF is:
8.5-9.5V.for modelsBHSand
4 or 6V lor modelsBHI and BHTU
Elackdnd'white photography
Sel the voltageabove6V Normally,a gleen
lilteris used.
(Fat enphaslzing padicular colors rcler
ta page57)
Focusis adjustedeitherthroughthe tocuslng
or the
oi the PMI0AD/PM_I0ADS
telescope
binoculartubeof the microscq*,(rcfer to
pages2627)
oADS/
I Thefocusingtelescopeof lhe PM_1
PM-1oADincludesdifferentphololrames
lor diflerentlypesof film.
I l}le lype oJJindereyepiece\?tes wilh
iilm sizeand eachtype of nnderelepiece
incluclesseveralpholoframesindicating
oi NFKphoto
diile€nl magnilicalions
Presslhe shutlerreleasebutlon.After
complelingexposure,checkior the sound
of ilm wnding.
aesure adjustmentis basedon the lolal
lo
ion oi the specirnen.freler
qt
Magnification
equipment
of thephotographic
Photomagnlcalon on the flm surfacevaries
wth the prolectonlenglhoi lhe photographc
eqLrpmentand lhe type of pholoeyeoi-oce
lsed ln a casespirolorna_on
fcaton s
c o m p u t eb
d y m u t p y n ! t h e o b j e c t v em a g
n I cat on andtheprrotoeyepiece
magniTicat
on
v r t h t h e c o e i i ce n t s l e di n t h e t a b e o n
rne flgnr
errotogaptricequipment
Photoevepiece
FKNFK{ype
P.type
PM.lOAD
P]\,4.1OADS
0 5x
P[.410M
0.5x
PN,l,6
09x
BH2 PM 6
08x
0 4x
Whef usnq modelsPf.'ll0ADor PNrl]OADSrOXoblecliveand 2.5XNFKpholoeyep€ce
P h o t om a n1 c a lo n = r 0 x 2 . 5= 2 5
Differences
to the
in resolution
according
combinations
andeyepieces
of objectives
Eventholglrlvhenlhe overalpholo magni ca
Example:Pholo magnilicalion ol 100X
l o n s l h e s a m e .r e s oL r l 0 n! a r e s d e p e n d n g
on the combnalon of obleclveand pholo
l n o n t h e1 m p a n e
e y e p e c eT h er e s o u o
s mprovedj lhe magnflcaton oi the photo
eyepece s ow and thal oi the objeclve h glr
tl
[
.
As the photoshows.a combnalon of oblec.
l
a
Uve40X and NFK2 5X provdesa c ear
dspay of evenminLle magedela Foca
deplh.lrolrever.becomesslra ow
-
at'
t
, t.
a
'
't
. t t
tt
i '
,'l?
' o
' ;"
?1 'tr'-t
.,...1 lt
a
I
i
l..
1r
'
z l{
:''
.''l . a r
t
.e
..tt...
t-
.."tr
,1'.''.
' .o
',
1
".
.,.a\ I
^1
Oble.l ve S Pan 20X and prrolo eyep ece NFK 5x
Obrecrve s Pran40X and pholo eyepece NFK 25r
,"- --.'
Settingof photographic
magnification
(effective
magnification)
n'en takingpctures,youhaveto Jirstdecide
ar rhedesiredenlarqemenl
ralio.Themag.
r_calionon ihe film paneis generaly
:E:erminedby objectivemagnilicalionlimes
9oro eyepiecemagniiicaiion.But the eliec
*Theetfeclvemagnilicalion
llve magnilicallon
ol the piclurechanges
is basedon the
wilh the nume cal apenureol the objective, assumpllonthal ihe picturcis viewedwilhin
and ls normallybasedon lhe lollowingrealhe closesldistanceatlordingdislinclvision,
The values istedbelowdo not applyllten
tionship,which mustbe lakenintoaccolnl
when enlarginglhe pholomicrcgraph.
tryingto proiecl35mrnlormal slides.
[email protected].<M<1qnN.A.
Si6ctiYe
.agnilication
tF ol
ai.tclive
t-4"
EHive
atnilication
't0x
tx
N.A =nune,iczta\dup at th. obta.iE M
mx
'tmx
40x
S Plan
S Plan
S Plan
S Plan
S Plan
S Plan
S Plan
S Plan
S Plan
S Plan
0.13
0.16
0.30
0.40
0.46
o.7D
0.70
0.95
'|.25
1.30
65- 130
80- 160 150-3m 200-4m
230-Affi
350- 700 350-700 475- 950 625- 1250 650- r 300
Framing
of thespecimen
-! pholography,lhe specimen ollen
operationot lhe microscopebecomesmorc
.rollil inlolhe ohotoframe.Thisoroblem d ficull.Herc,theiramingof the specimen
is carriedout the p€ferrcd way,by rotating
!e overcome by rotaling the camera
the slage.
in relalion
lo the specimen,
though
'nE
of the finishedphotomicrographs
Checking
Phobrnicrography
inuol!€sscientjficpt|otc
gradry,which makesit impolatn/ethal lhe
dDl€rapher accuralelyrecor(bhis fndings
on film.Addliooally,lhe pholographshorid
conv€ya stror€ estrlelicimpressimto lhe
vis/ver Valuablerecordsshdid mt be
documeitedwith runof-theflill dtolo6.
Tofurlher inpro/e your rcsultsin photc
micrography.
lt h importantlhal you aler4/s
checkyo|lr o n pholo6.By relerdngto the
follo/vingchecl@inF,liou c€n pinpoinlany
problenEconcemingliour photo6.Tll€n prc
c€edlo the secondhalf of this section
lilled Photomicrcgraphy
Techniques.
3. Bfackrndwhlb photogt $if teler to
1. Ouosdonar€lallng to all phoiography
pages 5557)
(l)ls lhe imagein focw f/eter to pagesb-27)
(])Do€sthe tinishedphotographsfrori,
and lhe e&o6ureprcperlyadjusted?
gradedblackard whiletoneswilhoul
(refet to pages 4246)
(aHave din or dust speckson the sp€cimsn
sivelydark(solidblack)shadowareas
washed{ur(corndelely$/hite)highlights?
foundtheirway onlo the photograph?
pages
to
2&33)
telet
(3)lslhe specimenproperlystained?
(4)lsthereunevenilluminalion?
ffetef fo
pages225)
Z Coforphologr.phy frefer to pages 47.54)
(1)lsthe baclloround(emptyspace)whiteor
ol lightgrcy lone?
(4ls lhe color of he specimenaccuralely
reprcduced?
25
nrmKodachrome
1984Dec.l
Purpos€
1
a
a
3
6
7
8
I
10
tl
12
13
t4
11
0
1 77
lq
18
18
rs
19
ry
21
i
4
a
a_
n.
a
4
a
I
31
3
.v
5
Photomicrography
techniques
techniques
Exposure
adjustment
Whentakng pholomcrcgraphs,exposure
compensalon
is necessary
depending
on ihe
dislibUlonof the specimen
in lhe iield.
Butii the specimen
is evenlydistrbuted
ZP
at
al
or spoimeieingrcnge,
withinthe nlegrated
is requred(incaseoi 1X).
no compensalion
1. PM.lOAD
(60% Averagemeierlng wlth mod€l
PM.lOAD)
$
$
#
Z PM.IOADS
rMod€lPM.1oADS
can b€ u3€dtor inleg6ted melering ol 3ool"anctspol melenng
ot 1oh.\
Measuringalaa
Soecimencondlllon wlthin
the l% meledngar€a
dotted
Brightf
ieldbackground
wilhfairlydensesp€cimens
0.25X
con'
ieldbackground
Brighlf
specrmens
lainingscattered
0.5x
d slribLned
Specimenis evenLy
area.
wlhin lhe I % metering
1X
Sellhereclpro
lilmbelngused.
Aboulhalf(50%)01the
wiihln
specimen
is distribuled
the da background
*t
Abouionefourlh(250lo)of
lhe
specimen
ls distributed
n the
darkbackground
Thedarkbackgrcundis dotled the lSo/ASA
dial
sensitivily
Useof theAE lock
By prcssingthis buttonduringthe automatic AE lock operetion
exposurernode,you can adjustlhe exposure
P -10lDSfnr6grat€dmetoringg,%,
lime indicaledon the displaypanet(expected
spot msteringl%)
exposurelime, actualexposuretime).
_lhe
Wlen pressing AE 'ock hJtton,the
wuning light abo/e it nashe6to indicate
AE lock mo(b. The unil is oow readyfor
ptDlog€phirE.Tocancelthis tunctton
pressthe AE lock buttonagain.This
sl|itcheslhe lightoft andthe de\ricere
verlsto nonnalautornatic
oocure anode.
gimlnaling density yadatlooeby AE lock
1. Takingpanoramlcpictues uslng ih€
AE lock
For panoramapictureswheieany numberoi
copiesis lakenfiom differenlseclionsoi the
samespecimenor whereseveralpholosare
patchedlogeiher,pertectpanoramicphotc
graphswilh uniformdensitycan be lakenby
locking-ina lixed exposurefor all the pholo
PanoEmaphotography
Plr.loAD (averlge mei.riig d}%)
Aller poGiliorng lhe specimenlo be
photogmphed
and pressingthe AE lock
button,lhe $/amirulightabo/e it tlashes
to indicateAE lock mode.Then,after
laling the frst photograph.
eecr.ire
lirne is locked.in.
Tocancelthjstunclionpr€6slheAEbck
butlonagain.Thisswitchesthe waming
lightoff, and lhe devicerevertslo the
norml automallce4osure mo(b.
z Erampleol uslng lhe AE lock on model
PM.'IOADS
4)ln conbination with spot neteing
' specirnenareasrequiringspoi metering
of faminqprob
arenol cenlered
because
in the cenler
€ms,lheyarefirslposilioned
andthe AE lock is activated.Thenthe
soecimens
are movedbacklo lheirorlginal
aosilionand pholographed.
1 Th€sperm€ndetaills al th6 locallon
bul
whereit is to be pholographed,
exposurecannolbe m€suGd
2 Aller movhqth€ spocinendelailto
areaand aclival,ng
the measuring
the aE lock,lhesp€clm€nd€tailcan
now be movedbacklo iis o.loinal
postllonand rh6 pholooraphs lak€n
2)!!9!9gEP!!!9vt!!i! lJ
to naximum exposurctime
' exposure
displayls c oselo the maxrmum
duringfluores_
eroosure
lime,lor example
the pholographic
:ence photomicrography,
an underexposure
mighldisplay
eouipment
Thiscan
exposure.
^?rningduringautomalic
by usingtheAE lock,resulting
.€ prevented
. ongrme exposure.
Th€sAFEry lampcomerro r€d .nd
an audlblewarningsounds.
Compensating
fora film's
reciprocity
failure
W:thnormallyusedphol€raphicemutsions Exrnd.: Dai. ior rhe afirracl€r&llcs ol r€clprodty law teilu|e {Kodrkcotortflm DKD-|/|1)
thereis a rule(the reciprocilylaw)thal
determinesthe luminanceof he lightstriking
the lilm surface-Accordingto this rute,lhe
tolal amountof e&osure is delinedas the
produclof the lumimnceand lhe exposure
time. For example,the anoont of oocure
wita 1,60sec. o@surc at E is the iame
as lor I B0 sec,at f11. &Jtfor longerexpoGure
limesthis ruleno longerapplies,leadingto
unclerexposureand changesin color re
producljon,Thisphenomenon
is knq n as
reciprocilylaw lailure_Bul sjnce,in d|oto
exposure.lJnevencolor reprcdrctionmuat
compensation
by lilter are necessary.For
mrcrography,
be compensat€d
expoGure
compensation
for with a CCflter.
cannot 'll|e
additiomldah on recipnccitylaw tailure
be carriedout via the aperlufediaphraqm,
abore chan [sts lhe compensation
data characterislics
conlactthe lilm manufacturcr
exposuretime is lenglhenedor shonened
lor rcciprocitylawfailurecharacterislics
when
lo obtaina suitableexpmurelevel.lt the
usingKodachrome
25 ilm lor geneElptrolo
reciprocitydial is set on rnodetsPM-1OADS micrography.
The chan showslhat for tong
and PMIoAD, compensaiion
is carried
expo6uretimesexceeding1 sec.,bottl
oul automatically,
resullingin pfoper
exDo6ure
time @rnoensation
and color
Ex8mpleol comp€rcallon(u,t|€nUstngFulhhromeROl00)
No corn.
p€nselion
Compeft
sallon
Colorphotomicrography
Colorfilm
'i.v differenttypesoi coor i m are oiiered
:.e markettoday eavnu tfre Lrser2t a
:: rs 1')lrhich brandto use tor photomrcro
:rhy Normaly. day ghl typ
'- an ISO/ASAspeedof 50.100 s lsed
e microscopesreq! re a lghl baancing
' i r L B D - 22. N )
EKachrorne
64
EKachrame
100
Dayighttyp€
Agfachror.e
CT18
Agiachrome
50Type
S
Agiachrome
100
Fulchrome
50D
Fujichronre
r00D
t4
g
Eldachrome50
Agfachrome50 lype.L
::quiremenls forthe selectionof colorlilm
I m w t h h q h r e s o l v f gp o w e rs. f c e p h o l o
_ .rograplryrequres disoiayoi dela ed
-_!
d i s p a ym u s tb e a b e t o d s c r m n a t e
:.lween the f ne coor dilferentalonscon.
: .i e d i n a s n ! e s o e c m e n .
: : l h l L rr e p r o d u c t ooni s p e c m e nc o o r s
inoul backgrolndd scooraiion
l. vew or theselactors the iolowng condi
tons can be sel ior pholomcrolraohy
! l!!qq1!
(2)Good
colorcontrasl
l?lqlgqJ9!9r!9!!Iro!
For noma photomcrographyn lrans.
m l r e d l g hbr r g h l f i e d u m i n a l i ocno l o ri m
wth an lSo/ASAsoeedbelween50and 100wi
ass!re salisiaclorvqla i1y.For specal cases
s J c ha s p r n t e dp i r bc a t o no r e n l a r g e m e n i s
<cd:chro'.e fim oijers lood qla ly
ng dark specmens(phase
r.,/.e. oholograprr
conlrasl poarz€d ght. 1l!orescence)
exmsurelime ncreasesAtiroLrghtirere
.nay be ercessve gra n for cases reqLrr n-o
iasl shullersoeedslse flm willr a h gh
rSO/ASAspeed
Typesof filters
Fill€rs
Seectionoi filers lor usein phoiomicrc
graphyis basedon lhe tlpe ot I m used.
lor daylight
UseLBD-2N
lilmsandLBTI lters
lor tungsten
typelilm Forchanging
lighl
inlensity,
useof a neutraldensiiyiiler (ND)
is recommended.
Thenumberon the lllter
rirngiveslhe transmssion
valle In case
oi iillerND6,94% of rherotaliluminaling
lighiis absofbed
andony 6% of lhe ighl
is lrafsmiired.
Typesof lilter and their lunctions
F tersior lse n coor pholomicrography
comprise
ihe io lowingmaintypes:
I Lighl balancinglilter
Toconverllhe colorlemperalure
oi ihe mlcro
scopelghl sorirceto the lih beingused.
eND (neuiraldensity)lilter
.F
a
To feduce the lighl inlens ly without atiecllng color
lempelalure,when llluminalionis too b.ighl.
3 CC (colorcompensaling)
liltor
suchas slghldi
Forcolorrendlonproblems
pholos.
lerencesin co or hueor ladingin developed
a4-Didymiumlilter (markedFF
on Olynpusmod6ls)
€ Heatabsobinglilter
Toeriphaszecolor,i.e.wirentrying1ocompensate
tor the insullicient
inlinsicredcolorrendition
oi
a llm or lo enhance
theredcolorofthe specimen
or lhe coiorcontraslol Polaroid
coiorfim.
To absorbheal raysemanatingtrom lhe jghl
sourceof lhe m croscopelo preventdamageto
or deslruction
oi livespecimers.
Sincelhisfiler
lransmils
a smalqlanlty of bluelighi,I may
becomenecessarylo usea color.compensal
ng
lilter (CCl0Y or CCl0M)\rheniaklngco or photos.
depending
in colorrendition
Differences
of filmandlightsource
on thecombination
-.F are tro types0i coor i n's d.y 9r1
sm .r _vrh.n !s n,othe llngslen lype I m
- 1 b ra n c e dt o r s l n g r i l : n d i L r r l s l e f
o l t c o o r sa b ! s i r . l ' o t o qa t t s o b t a l ^ e o
The ta! I r Lrclrc:ses s ta Lrr. 10lse
: . 1 b a a n c e cI o ' a r t i c l a g h 1 ). h es e e c lo n
:he pr..e- I r. .r r:lcf .he ,ori sc! cF
. . i d n g u p o ni h c L l l n ls o ] ' c e V r l l a y
. s e r sh a v ee r p e r e n c eich ea s a c p o n l m c n i
r e r o h o t o sa p p e a r . ! ? l c e s s l e y r e d
_
' ! . l h e yl s e d d a y
Slr_ m lnder n.andes
lo or rendilion dependingon the combination ol lilm and lighl source
Daylishl.lype(sunlighl)
I
Tungsten.type(adificial lighl)
incolorreproduction
depending
Differences
in colortemperature
on differences
Color l€mp€ralure
Dillerenc€s
in color reproduclion
d€pendingon dillerencesin colorlempefature
Coor temperaturedesignatesthe propenles
of the ight source.A blackbodyradator.
whenhealed.emanalesighl of dfferenlcoor
dependrng
on the temperatureThe properlies
oflhe ightsourcecan be nd caledby reiering
to lhe lemperalLrre
oi the backbodyat a fxed
ter.peratureevel.Ttiis ieature s ca ed coor
tenrperatLrre
the numericaLrnlbeingex
pressedether rn absoute lemperalureor
in degreesKevin (K)
C o o ri l m f o r u s e n p h o l o m c r o q r a p h
sy
normaiydaylghl lype bul sincethe ight
sourceoi lhe mcroscopeis a tungslenlype
ls coo. temperatures low (2800.3400rc
and as such il is unsuilableior daylighlfllril
wlh ts color lemperalureoi 550C6000K
In orderto achlevethe propercoor rendilion
a ight baanc ng i ler is used,and llre
lungslen ghl s convened10dayight
l
j , ',.
'
s h qh l exhbilsa'blue s
lli colortemperalur".
s low lhe photoshowsa redsh11 and .doi renoeralure
E
'
>
in colorrendition
depending
Differences
on the typeof film
Colorr€r'dilionfor diflerenttypesot film f.om
lhs sam€manufacfurcrlendsto vary,ard
evenditferentorcduclionlots of lhe same
typeshov/sljghtdtferences.
Whenyou slan to lake color photo6,lirD
drouldflrst d6l6mlnelhe oFffmumcondilibns
to thenlmthrought€stphotogkphs,matchlng
flem wih boh micrGcooeand soecimen
(Fot the methoclol taking t$t photos
rctet to page 54)
Offl€r€nce3In color davalopftGit
&pddlr! dr lh. m. ol lllm
I}re 6lx ptroloGon the rlghtweretak€nwllh
flms ol dlfler€it brancls,and wflfi thE €xc€p
tbn of lhe ISOASAspeedall drotographic
conditions
wereidentical.lhis €vampledearly
*lo € that propertiessuchas color rerditior!
contrasl,cladtyof baclgrerd, etc.are alldlfftrcnt becauseof lhe variousbrards of tilm.
Photognphlc condldort
BHS, PM.ioAD, LBD,{ (color tompontu|l 5500f0,LBT (color tampor.ture 3,mr+
duii.. 3pe€d0.1+l s€c
D.tlglrt tyF
llrngd€.r lpe
Purchase
of colorfilm
1. Whatlo walchlorwh€nbuylngcolorlilm
I Coorlilm is a hghy sensitive
subslance,
andenvircnmenta
laclorssuchas heal
andhumdityeasilycausechangesn film
speedandcoor rendition.
Whenbuyng
coor film,avoidcamerasioresin which
the sheves slorng color t m are exposed
to sunlghl.
I Choose
onlycobr r m wiiha sufficienly
longpeiod beforethe explraiion
dare
as markedon lhe package.
I Coor fenditionoi lhe samelilm tlpe may
diiier,ii il cornprises
dltierenlprodLrction
lots.li youlrequently
lse coor film,you
photographic
canuseil underideniical
conditions
andavod varaiionsin thecoor
ol the t lrn if youbuy argequanlilies
irom
the sameproduction
lol.
EE:
;:!
1602/54
sg:
im
of Yalldlry(ddEnon dal€)
3. Poinlscalllngtor sp€clalatt€ntlon
2. How to slore lilm
whenusinglhe lllm
Forthe basicpinciplesol composlon and
colorrendition
checkthe lechnicai
literalure I Donotleaveihe Jimin ihe cameralonger
on lhe subjecl.As hasbeenpoinledout,
perforrnance
changes
according
1othe con.
dilionsunderwhichthe f m is slored,
ColorI m is normallykept
n therelrigerator
lo protectit irorn the etlecls ol heatand
humidjty.
Removelhe
lilmtromlhereirgeralor
t hourbeforelse andalor t 10reachroom
lempe€lure.
Do nol remove
ihe fim lrcm
lhe rc immediatey
aftertakinqit out ol the
/-f--\
*,
[ (,r
,
/-F\
I Developexposedf m as soonas possible.
@M
z
%
o
reirigeralor
lo prevenlcondensalion
on lhe
I
I
;:E
ii
I Eventhougholherpholograph
c condilions
colorrendilion
mayvary
maybe idenlical,
wlh lowor highambient
temperalures.
I Do nol usethe fim in a gaseous
en(suchas lorman),sincecolor
vironment
rcndilion
wil be aclversely
aflecled.
(CC)filters
Useof color-compensating
whentakingcolorphotos
liltersarehighly
S:ncecolorcompensaiing
sensilive
lo heai,do nol usethemclose
:c lhe lighl source.
ICC filter is the abbrevialionfor colorfiler,andlhe coloroi ihe
compensaling
parllcularJilteris markedin capita letters
on its rim.Sixdillerenlcolorsare available,
eachcolor beingofferedin six dilferenl
oi densily.
No.5 andNo.10 are
degrees
mostwidelyusedin pholomicrography.
lcolor
fltersare madeby severananu'lacturers.
CheckyourOlwpus dealeror
camerasloreior avaiiability.
a repuiable
Color to b€ r€duc€d
BLUE
CYAN
GNEEN
Color€mp€n6ating lllt.r rcqulr€d
ccY
Red
Magenla
YELLOW
Blue
RED
CYan
MAGENTA
Green
ccB
ccM
ccB
CCG
H
Testphotography
1. The need lor lest pholography
equp'
Evenii you |]seaulomatcpholographic
menl and set bothlhe ISO/ASAspeedand
lhe rec proclly aw iaiure characteislcs,
you cannolbe sure that yo! wil always
ior everytype
ach eve the perfeclexposLrre
ol specimenThe reasonior llrls s that
phoiograplrlc
equpmenl s mafufactu.edto
malch dstfbLrtonstate,coor and exposure
slandardoi averagespeclmens.ln orderto
obralnbelter resulls,you shold I rst lake
tesl pholographslo match lhe conditionsoi
iilers etc
the sp€cimen,fim, nricroscope.
2. Method lor test photography
Charl2: Datacharl
(when using nadels BHS and PM-12AD
Color
Colorcom.
and reversal filn)
lemperelure Exposure
(1)Prepare
one rol oi illm (36 exposlres)and
1
a irequentyusedspecmen
0.8x
No com' (Z)Setlhe lest condilions(seechan 1) and
2
1X
pensar
on
preparea data chait (seecharl 2). Listng
1.25X
a the condtons gven n charl 1 wil resLrll
in a dala chan 2. TakeyoLr photosbased
0.8x_
1
bgooK
on this chair and delermnelhe optmum
1X
5 o r
CC5G
pholograph
c conditionson the basjsol lhe
10v
resulisoblaned on lhe developedI m.
a
1.25X
it
7
8
9
cc5M
0.8x
lx
1,?sI
J9 ssoox
sv
1'
0.8x
'lx
'| 25X
Charl l: Testconditions
170mired,180m red, lmmred,
5900K
5500K
5260K
Volase 10V,9V 8V
0.8x, 1.25X
Measuingwilh CTR
No CTR
ng I lter
Color compensatlon Coorcompensal
3. Evaluatlnglhe test photos
observalon oi reversali m transparenc
es
can be caried oui eillrerwilh a lighl box
or a slideprojeclor,bLrtihe dispiayof cotor
vares wde y wilh lhe colortemperalureot
lheselighisourcesAs a slandardior evauaf
ing the pfopef coor Lrsea ghl box wilh
a color
'Tl]e lemperalurecloseto 5000K.
io lowinglp€s ol lghi boxesare
avaiableon lhe market
C Fuji film color box 5000
O Dlrotesl ColorClasse.50
O GeneralEecrric Chroma50
4. Organizingphotographingdata
[.4akeI a habil1o recordal dala rearing
io bollr lesl phoiographyand reguarphotc
graphy.lf the pholographiccondilionslhat
resulledin goodpholosare relaned on i e.
lrolbeshoolng and correclon of probems
can be caried oul quicklyand efiiclenly.
Cc5Gnocorapensat
on,c,cs\,tr
5. Slodng developd film
Whenstorng I m ior a ong perod oi tme.
the most imporlantprecaulon s proteclion
frornlight Siice mold is prevalentin hot anc
humd paces, t is advlsablelo keeprhe iim
liglrty seaed, togetherwilh a desccant
We recamnend storing film in an envnon.
nentwith a relativehunidiry of 1U0% and
an ambient tenperaturebelow 21oC(71'f ).
(fron Kodak Colot OKP 141instructions).
photography
Black.and-white
film
Black-and-white
: nce the condtons diier ior plrotomcrcr
phoiography.
i1 s
: aphyand generaL
:cessaryto seecl lhe properiim Joreach
'.oe oi pholographyIn photomc rogfaphy.
! h c o n l r a sjl m w t h r n e g r an i s u s e di n
-.der to documentm nule slrircluresol
: oo! ca specmensand to achievesharp
:_ololraphic reproduclon.Examplesoi lhs
n typeare KodakPanatomc X. Agiapan25
.rd Pan F. and Fll NeopanF
I
I
€
,:jl r#
Jse ol black-and.whileliln
':re are manylypesof blackand.whle ilm
j the keytoobla ninggoodpholographses
r e e c t n gt h e r i g h tf l m l o r a p a r l c u l ajro b
Specinren
Normaor hgh conkasl
ior tlreslanedsecllonoi
generapathoogical
Agfapan25
KodakTechnicalPan
palno09 ca speclmens.
2415
as we as thoseol other Agfaonho25
Wlren
shape
s
areas.
more mpo(anl llran
Agiapan 400
Dark specmens.and
when ong srrltter speed l l o r d H P 5
SO/ASA
32
50
25
32
on the coor 01lhe
1
Srce the conlrastoi the film
is very lrigh,the rangeor Pro
64 per exposureis very narrcw
expos!reis sel wlhln
100 Uriless
+l/3 slep oi lhe opi murn
25 lhe pholow I be ether over
32
*r,"nvo,
X33 u""t eseryp""
400 want10shol(enexposlretme
ef flm glan
5 G 1 6 0 0a 1 l h ee x p e n s o
400
Comparison
film brands
of different
Typesoi fim orovdinga wde gradalon iom
shadowro h 9lr _ohts
and featurnggood
conlrastare Fll NeopanF and Kodak
PanalomcX. TechnicaPan oiiers s ghty
h gher contrasl wh le N4n Ci]pyerimnales
NeopanSS and Tr X on lrre olher hand.
havefanly o!\rconliast
Kodak Techncar Pan 24t5
t
Filters
Ellects ot verlous conlrasl lillers
Us€ ot contrast lillors
Conirastfllers are usedio conlrollhe conlrast
pholos.
or b ack.and'white
green
Forblackandwhilepholomicrography,
i tersare normally
used,bul choceoi lhe
mosleffective
flter depends
on lhe tlpe of
Filterclhatenhafcethe colors
specimen.
andconlrastof the specimen
are:
Color ol lh6
Colorol rh€ tllt r
Green
Blue
Yellow/orange
Oranqe
Blue
usea Ulleroi lhesarne
Forreducing
conlrasi,
.oloras lhe specmen.
Reasonsfor using a gr€€n lill,ar
-iere
aretwo reasonswhyconlrastimproves
shef a greenllter is used.
T)Snce objectiveabe(ationsare rnosl
effectivelycompensalednearlhe green
wavelenglh,
lossoi imagec arityduelo
chromaticabefialionis avenedby a green
I ller.
2)Dyessuchas hematoxyln
andeosin
in higher
absorbgreenlighlwell,resulting
conlrast
whena greenlllleris used.
the red po.lion is €nhanced
lhs r€d Porlionis reduced
Photography
with Polaroid@
film
Oneof the acl€ntagesof Polarod litm is thal
I canbe quick]yvlewedas a iinished
pholo,
bul il is seldompossible10oblaincotor
reproduclloncomparableto thal oi 35mm
ol lllm us€d ln
r.Pny
t 1E' xS th'
(actuelslzo73crnx 9.5cn)
I
k
I
a
n
I
Type
107
67
tso/AsArackcontainin! Tlpe
spe€o
8 photos
3m0
3000
o
o
ISO/ASA
speed
52
Sheel
4(J0
o
30m
o
rack coolaining
I photos
o
4m
665
(PdnU
I negalave)
50
108
80
80
I
,l'x5" (actualda gdnx11.5cm)
668
669
14'x5'
o
55
(P nU
negative)
o
o
o
80
50
58
59
80
559
80
80
2. Photogrrphlc t€chnlqu€s
l1JBIackand -whi te photogh phy
Usea greenfilter to obtaingoodcontrast.
(2)Colotphotogftphy
Sincelheoverallcolorhuetendsto be either
ighlgreenor blue,useCC10-20Mor
CC10-20Ylor compensation.
ll youwanl
io enhance
colorconlrast,
usethe FFiiller
availabletromyou. Olympusdealer.
SincePolaroidiilm is morc easityatfected
by reciprocilylaw Jailurethan normaliilm,
exposurclime shouldbe adjusted10belween
0-05and0.5seconds.
o
o
o
o
filmhoLclers
dilleraccording
to lhe typeot titm
sh@i lllm-:(s typei m hotd€r
Packedfr/m-550 typ€ tllm hotdef
by'haPora|o]dcor@ralioi'camb.ido€.MlEs'U's.A'
4500K-LBE!2Nolus FF tllto.
cororlemoeraiure
SectionS
Howto obtalngoodprinis
. . @-61
H o wt o o b t a i no o o dc o l o rp r i n t s. . . . . . . . . . . .
aPreparingblacLand-whitebrlnts. . . . . . . . . . . . .. . . 62
Maiks on the Dhoto. . . . . . . . . . . . . . . , . . . . . . . . . . . . . . .63
... ......64-65
Howto avoidmarksduringdevelopment
Howto obtaingoodcolorprints
ThelollowlngdiagEmshowshow to get
good color pdnts {positiyepdnb).
Color negalivelilm
ln pholomlcrography.
lhe colorreprodlction
oi lin shedpr nts takenwllh negalve fi m
differsirom the observedcolor-a lact many
usersare probaby very muchawa.eof.
reasons
forthisarethalthetechnician
Possible
who madelhe pinis d d notviewthe speci
mencolorlhrougha microscope
andthus
didnol knowlhe actualcolor,oathe pholc
gapher madea mistakewhenhe lsed a f ller
andlailedto noticeihal lhe iilm quality
deteriorated.
In ordeflo gel beter resulls,
you shouldlirst photographlhe specime.
tromwhichyouwanl lo get prinlson reversa
R6wtsalfllm
photoas cojor
llm, andatlachlhe iinished
reterence
sample1othe negative
lilm.Pinls
canalsobe madelromcolorrcvercal
{ilm,
bul lhe processdoesnot matchthe pint
qualityas obtainedfrom negativelilm.
a
Color pdnl with Poor color r€produclion
Color pdnt with good color reprcduclion
Preparing
prints
black-and-white
Therearemanytechnicat
pubtications
on ihe
markelllrat exptan in delaithowto devetop
iilm. li you planto do yourowr iitm pro
cessrng,uselhesebooksas reference.
Butsinceconlrastin photomlcrog.aphy
is
owerthanwithnormalphoiographlc
subjecls,
youshorildpaypanicular
attention
lo under.
developmenl.
ll youmakea positive
print
iromanunderdeveloped,
/ow-density
negalive,
an evengrealerreduction
in contraslwitl
occur,badnglo a poorresuti.
1. How to obtain a good negative
(])ljse a devetoper
thalmalcheslhe iitm
(2)Usethedeveloper
allhe lemperalure
specilied
by the manuiacturer
(3)Pdherelo the speciiieddeveopmentiime
lhal malchesllm sensilvity
(4)Agilate
genllyandfreqlenllyto eliminaie
unevenoeveopmeni
i5)Makesurelhal lhereis neltherloo much
nor loo Itl e it ng
(6)Followthe paperspecificaliolswhen
washing
the prinl
carelutty
andpfolect
OHandleihe negative
1 lromfingerp|nts,
scraiches
anddusl
2. Selectionof phorogEphlc pap€r
Whenmakingpints, youcanvarytlte contrasl
dependng on lhe t 9€ of pinting paper.
selectthe lpe that bestsuitsbolhthe
densityof thenegativeandsp€ctmencontrast.
Theexamples
on ihe fightshowprinlsmade
iromlhe samenegative
whenusingdifierent
tlpes 01pinling paper.
Gradeno ol
photographlcpap€r
t€
Grade3 nomalcontrast
Markson the photo
1.Ma*s as a esult ol negaliv€d€veloFnent 2 Ma*s occuriog dudngprinl devglopment
Markson the printtendto slandoul more
prominenlly
in pholomicrography
thanon
nomal pholosol p€opleand landscapes.
empoyslilm with
Sincepholomicrcgraphy
faiiy highconlrasl,
eventhe rosl minutedif
ferencesin brighlnessshowup in the piclure.
MarksresuI ng from improperdevelopment
Thissecliondeals
arealsoveryconspicuous.
wth marksresulting
lromdeveloprnenl.
Bothuppd and lowerponbns of lho photoare
bnohr* thanlhe centralsection.
Markswith iregularb qhlness.howup.
How to avoid creating ma*s
(1)Confirm
ol lhe
thedegreeol exhaustion
processorto develop
It youusean aLnomatic
lhe prinlingpaperno markswillocclr,but
slncemar!€caneasirysho/ up when
deveopingin a t€y, rhe io lowingpoinls
Carelully
noteproducl
on daleandlre.
quencyol useof the deveLoper
(2)Agilale
y beioreLrse. How to avoidcrcatingna*s
thedeveloper
lhorough
(1)Agilale
lhoroughly.
lhe develop€r
(3)Usethe developer
al lhe specified
(2)Keepthe lemperalureof lhe developerat
20'C.or as speciiied
by the manuiaclurer.
(4)Donot workwlh an exposirelhat resllts
markscaneasilyoccurif deveor
ina developmenl
limeof Less
lhan5 mlnules (3)Since
(lheresullwil be exlremely
clesensitized menttime s too short,set the exposure
of lheenargeral sucha evelthatdeveloP
developmenl).
menttirnes betweenI min30 secand
(s)Agilale
duringdeveoF
lhebaihthoroughly
2 min.
(4)Agitate
thebalhthoroughly
duringdevelotr
(6)Mainiain
lhe speciiied
lixng lme.
y.
f)Washlhe filmthorough
(s)Conl
nueto agilalelhorclghyevenwhile
andiixing.
stopping
(6)Wash
thepinl lhorcughly
in runnlng
waler.
paper 30€0 min
Regularbase
paper-4.5 min
Resin-coaled
Howto avoidmarksduringdevelopment
DoYslopmentol tllm
1
2
lmmerselhe ree in lhe developer,and aiter
turningii 2'3 limes,lap il anolher2 3 limes
agansl lhe botlomol thelankto remove
a r bubbles
sllcknglo the fim surface.
Complete
thisprocess
wilhn 56 seconds.
and
Nowremove
the reellromlhe developer
immerse
il again.Aflerrcpealng
mmedialey
iurnlhe ree immersed
in
lhs procedure,
23 times,thenpausefor
lhe developer
30 seconds.
Sncema.kson lhe lilm occureasilywhen
uslnga ree-tlpelank,developmeni
should
be donein lhe io lowingmaanern the
Development
ot pdntlngpaper
t,
'@@@
.l
2
I
Thisseclon dealswithdevelopmenl
using
X
a tlay.
/ ---
-
-
-
-\-
Whie holdlnglhe edgeol the exposedpr nling lAs shoM in lhe piclure,do noi immerse
paperlighllywilh a pair of longs,quickly
lhe prinlng paperinlo the developer
parallelto lhe developersurface,bU till
place il inio the developer.
il nlo the deve oper balh.
lollowlhe
Alter completingdevelopmenl,
iixlng,andwashng
sequence
of stopping,
nol b€sut
Thodeveloperwill
ficientlyaoilaredsimplyby
tuning lhet@|,andtheresull
will be sporson ihe lilm,
lh€ rcelfromlhe dev6lop.r,lhe
By @movlng
d€v€lopscanreachlh€ spac€sbetweentheroll6d
p€vontlno
lilm,
lhe oon€nlingofspols.
nlervals
I I the compielon
Pedormlhe operalon shownin piclure2 of ihe leftin 3o.second
work.
ol developmenl
paper
I Makesurethallhe photograplric
doesnol float!p lo the surfaceof lhe
ro agilale the phologGphic paper, ho d il
at a cornerwlh a pair oi longs and move
r back and lorlh whlle I is submergedn lhe
aeveloper.Repeatthis operalion in 15-second
ntervas unilldeveopmentworkis compe1ed.
workon lhe
Aftercomplelifg
development
prntingpaper,qucky pe.lormthe sequence
fixing,andwashng.
ol stopping,
sheelsot prifilng
I Makesurelhatseparale
paperdo nol sticktogether
if thefxer
I Do nol usethe samepar of iongsduring
andlixinglhalyourseddu.ng
slopping
developmenl
work.
thedeveloP
Makesurenolto codaminale
ef wth e lherslopbaihor I xer qud.
Section4
Trouble-shooling
P|obloms in linished pholos and lheir con€ction
Poorcolorreproduction
. . . . . . . . .. .. . . . . . . . . . .. .68-7'1
B l u r r ei m
d age
..,,........72-73
Theimageis in focusbut not sharp.. . . . . . . . .. . . . 74-79
Objectsother than the specimenimageappeared
on the tilm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .80-81
U n e v ebnr i g h t n e s s
........82-83
Prcblemsin finishedphotosand theil conection
-Poor colorreproduction
1. Th€ beckgroundis coloEd (.€dibhF)
Caul6
Cor'rctlon
Rama*s
Rel€r lo
Daq€
The color temDeralu@of the lliuminatlon ls not malched to that of
the fikn.
.Lioht balanclngfllter specified by
manulacturerls not us€d.
.ljse flltels sp€clfled by the manu'
lacturer.
.Daylisht type: LBI!2N, 5,5mK,
D settlng
.Tungst€n type: LBT,34mK
T setting
Page48/
page 50
Lamp \oltage ls too low (loo high).
.Raise (low60 lamp voltag€
.BHS above&5V
.BHT and BHTUat 5-6V
.lf you have reachedthe desired
color temp€ature, do not chanoe
the voltage position. lf you want
to chango lrieBity, u6€ an ND
fllt6r.
.!!b recommendthat for iest
photograplrythe voltage should
be set lo the standad Dosition
+1V whgnfi6t detednining
lhE ootimum color temDeEturc.
Pag65(y
page 54
6v-rcltege ls t@ low {Bd hu6).
l1v-rcllaoo l. too hloh{bluohu€).
Blackenlngof lamp due to pre
longed Uae
.Replace wlth a now lamp
@
2. The backgroundis color€d (gr€€dmagenta)
R€marks
R€f€rto
Deq€
Caus6
Con€ction
.A iilm dilferentfrom yourusual
film lype was used.
.Since colorrenditionvarieswith
differentfilm typesevenfromthe
samemaker,choosea lilm with
coloringsuitablefor yourtype
.A iilm of the sametypebutwith
difterent
emulsion
number
was
used,
.li possible
buyfilmwiththe
numb€r
in laryo
sameemulsion
quantities.
.ln orderto maintainthe performancequalityof the film, slore
it in a relfigoralor and remove
it one ho0rpriorto us6,allowing
it to reach room lemperature,
.Even for film of the sametype
therewill be slightvariationsin
colof reproduction
dependingon
labofatory,developmentcondllions,
and stainingof the specimen.
Pag652
.l.Jsea color€ompensating
(CC)
lilter
'Color€ompensatin0
(CC)tllters
Buyfromyourpholodealer,
.Too muchgr€€n
UseCcM {mag€nta)
.Toomuchpink
UseCCG(gr€en)
Page53
a
a
t
a
rt
a
I
oC
O
ta
t
t l
O
-
t
t
t
t
ar I
e aB
t l
a
+
t
ra
rt
cc05M-optlmumcompensallon
a
a
O
rt
:i.
r.l
r
a
a
*
.a
cc10M-ov6r{ompensared
Page 51
usecc
Forphotomicrography,
filters0+10.ForlurlhercompensaandCClologether
tionuseCC05
3. Inconeclcolor rendilion
Caus€
Con€ctlon
Remarks
Relerlo
pag€
.An excessively
longexposure
time hasbeenusedandthe characieristicsof reciprocitylaw
failureleadio incoftectcolor
rendition.
.Set shutterspe€dat 0.01{.05sec.
Adjustshutterspeed
uniJormly
as
fal aspossible
byusingan ND{ilter.
.When usinglongexposurelimes
(above0.5sec)wilh models
PIV'1oAD
and PM'1oADS,
set the
characterislics
numberof the film
usedon ihe dial for reciprocity
law failure,
.With longexposuretimes,even
wiih the exposurctime compensated,thecolorfenditionchanges
as a rcsullof film properties.
Usea colorcompensaling
filter
speciiiedby the film manu'
.Automaticexposurehas been
usedwithouladiustment.
.When usingmodelsP[,]-1oADS/
PMJoADset the exposureadiustmentdiallo:
0.8-0.25X
for bighl background
specimens
andto 1.2t4Xfordark
backgroundspecimens.
.Carry out exposu€adjustments
Brlghtbackgrou
nd:ISO/ASA
100to 50
Darkbackground:
ISO/ASA
100to 200
.ll you changethe magnification Page4Z
of the objeclive,the lightdistribu- page 43
lion withinthe visualfield will
atsocnange.
.Aclual and nominalsensitivity
ol lhe iilrn differ
.Vary the ISO/ASAspeedsetling.
.Nominal speedmay diiJerlrom
actuallight sensitivityas much
as 1/3to 1/2aperture.
Page 216
Page 42
4. Poorcolorol a prinl enlarg€dlrom a negative
.The printingtechnicianin the
laboraloryis nol sureof lhe
subiect'scorreclcolor.
.Add a lransparency
of the same
specimenwith correctcoloras
a sample.
.Better coior is achievedwhen
with
shootinga transparcncy
reversalcolorfilm (butthe cost
of pints goesup and quality
1
5. Poorcolorol Polaroidlilm
Cau3e
colortemperature.
.Abnormalcolor characieristics.
.Term of validilyof the lllm has
.ll roomtemperature
is abruptly
lowered(raised),
sensitivityis
alfectedand printcolorturns
towardsblueted).
Coneclion
Remarks
R€ferto
.The film is a daylighttype,but
becauseof the film characteristics colortemperature
shoutdbe
set lowerthan for 35mmfilm.
.lf you s€t colortemperature
b+
tween2rc00K
and 5000Kand
use an OlympusFF filter,color
contraslwill b€ enhanced.
Page58
.lf the abnormalilies
are within
the adjusimentrangelor color
temperature
and withinthe
operalionalrangeol the colorcompensatlng
lilter,determine
ihe conditionsbytest photography
.Use film whoseterm of validity
has not yet expired.
.Since Polaroidfilm is easily
Page54
aftected by adversestorage
page58
conditionsas far as colorcharacleristicsare concemed,protectit
irom heatandhumiditybystoring
it in a refrigerator.
.For developmeni
time and color
reproduction,
referto the film
instructions,
Blurredimage
l. The ov€rall locus ol the oicturc is blun€d
Cau3€
.Finder 6yepisceor focusingt6lescopeis nol prope y adjusted,
Conlction
.Adjust thediopteruntilthedouble
crosslinesare clearlyvisible.
Remarks
.Slnce most peopledo not have
the samevisualacuityln both
eyes,d€l€mlnewhicheyeyou
alwaysuse for focusing.
page
Pages
2e27
lt
ll
.Blur anddrittresulting
from
vibrations.
.Uss a vlbration.proof
table.
dF
lm.o..fi6t.d
by vlbEttons
.Uss an ND fllt€r and lncrease
shutter speed (1/2sec to 1 sec).
.Use a standfor the photographlc
equipmentto separatephotographlcequipmenl
andmicroscope.
Page12
paqe 48
2. Focuslngenor occurs wh€n you u96 a low.magnlcallonobiectiveol less than 4X
Remafts
Ceus€
Coar€cllon
.lf magni{ication
is low,tocal
depthat the lllm planebecomes
shalloq easilycausingerors
in focusing.
.Mouni a locuslngmagnifierto
lhef Indereyepiece
or thefocusing
lelescope,and afterfocusingon
the doublecrosslines,fine focus
the specimen.
.lncreasingthe magniticatlon
eliminate6tocusingerors,
Reiar
pa96to
Page27
3. The p€rlph€ryls unilomly blun€d
.An Achromattype obj€ctive was
us€o.
l,r,i';.,r',
1.,iil;.'
R6ma|ks
Coreclion
Cau3€
ausea PlanAchromat
type
obiective
Reler to
paq€
Pag€14
;;1..i.'i:.1
;i:'-:: l,i;"li ;1
/:::).\
.'-
-::'
.):tt.
::
::.:':itl:i: -r11.;'ij;:. : .ij,
':i
;';iiii,''''r,;ii'':.:r'::;ji liir'i'.1.t...'1,.:
:',:,1:,11i,;i;,,.,. .;.,.
;t::'.
. - . _ :J : - - i : . . ! : - : r .
.: .i!:::.:'1:.:):.t .
D Plan1oXNFK2-5X
.Objective and photo eyepiece
were usgd in th6 wrong combination.
.LB series(longbanel)-NFK
photo eyepiece
ashort-barrel
serles-FK photo
eyep|ece
Page13
Theimageis in focusbut notsharp
1. Inadequat€
resolvingpower
.Use ol a combinationof lowmagnification
obiectiveand highmagnification
pholoeyepiece,
.ln orderto obtainhigh resotuiion,
usean objectivewith large
numeicalapertureand a pholo
eyepiecewith low magnification.
-?
,1
.Stop downthe
apertureiris diaphragmto 60€0%
.Use of the condenserwith lhe
apenureiis diaphragmfully
opened.
.The field iris diaphragm
was fully
opened,
.Use of thick coverglass.
straylight,slop
downihe field iris
diaphramto an
areaonly slightly
largerthanlhe
.Use a coverglasswith a thick.
nessof 0.17mm.
':
'
-:-. ,,
.'''.
' - : .= ' .
.. '.t
.ln orderto obtaina magni{ication
;i ioot i; tniirr. pti ", ,iJ-
eitheran objectivezloxwith a
photoeyepieceof 2.5Xor an
objective20Xwith a pholoeye
piece5X.To increaseresolving
power,the combinationof obiective40Xand photoeyepiece2.5X i
(Focaldepth,
is pfeferabie.
however,
will b€comeshallow)
.Vary the amountby wh;chyou
reducetheaperlureirisdiaphragm
accordingto the magnificaiion
of the objectiveand the contrasi
ol the specimen.
.Do not reducethe diameteroflhe
field kis diaphragmto such an
extentthat it louchesthe frame
reticlebecauselhe aclualarea
photographed
is alwaysslightly
largerlhan the areawilhin the
lramereticle.
.Olympusobjectives
lor biological
specimenshavebeendesigned
in sucha way ihai optimumresolvingpoweris obtaaned
when
a coverglasswith a lhickness
of 0.17mmis used.
Pase
38
Cause
Cor€ction
.The specimenslain is too weak,
resultingin lack of conlrast.
(Photographing
a specimenthat
cannotbe stained)
.Use a denserstain,
.Use a contrastlilter (fof black-and.whiie
pholography
only).
.ll the specimencannolbe slained,usephasecontrast,differentialinteF
iercncecontasl or darklieldto createconirasloDticallv.
Beler lo
pag6
Page57
gee
G*,il
_Effiil
ST
oiii€Gnlial inlorfercnce conlrasl
Cause
Remarks
ConEctlon
Colorphotography
Ref€rto
peg€
Page51
.Use of a low convasifilm,
.l.Js€a high conlrastfllm.
:',
r-'d):.:')*.:;i;
tt))b,;nite-:i
i:);'.':ii;i;?;":i
photography
Black'and.whlte
.Variationsin lhe so€clrals€nsitivityof the film ifiect ihe
Page57
.Normally,a greenfilter is used,
but if you want to emphasize
a
sp€cificportionof the sp€clm€n,
use anothercontrastflller,
.lf a filtercomplementing
thecolor
of a specimenis used,it will
emphasizelhe contrast.
2. The image appearshazy
Caus€
ConEctlon
aTheconectioncollarof the obiectiveis not adlustedto the
lhicknessof the cov€rglass,
,i
, .
r, 'Ja.
'rt
.
.
j
.:-1 'l:
i
,' '-r',!! ba
.
' . r
. ' l r .
.Adjust the conectioncollarwhile
€xaminingthe specimenand set
it at a positionprovidinga cl6ar
image.
R€marks
.On the LB objectivea correction
collaris mountedon the S Plan
Apo 40X S Plan100Xdry and
D PlanApo 60X.
Rolorto
pags
Page16
Cause
.An objectivenormallyusedwilh
cover{lassedspecimenswas
ussdon a specimenwithoutcover
glass(orvice vefsa),
Bema*s
Con€cllon
.Use a no{overobjective.
' ji;t,
) \.'1t
-.J
Relgl lo
pags
LB objectives
.NC S Plan40X
.NC D PlanFL mX
.NC S PlanApo 1mX oil
.NC S Plan100Xdry
Page17
.lt you use dry 40Xor 60X
objectivestogetherwiih an oil.
ths oilon ihe
immersion
objective,
specimenmay soil the fronl
lensesol lhe dry objeciives.
Pages
26- 32
'\
)/ '-. J .'1\)/' U) . . ^ \ J \ - ' ,. 4
)-
''(t
-\
!.-
!i
- - - , ,- ',--'(.
r ' - - \-v:'
- 'J
.Finqenrints,an oil lilm or dirl
particlesin the opticalsyslem
(objectivefront lens,photoeyepiece,prism,specimen,etc.)
..,t,
.
1:
,', .'
.-.t .
.Clean lhe oplicalsystem.
.Alwayscoverlhemicroscope
with
a dust coverwhen not in use.
3. No sharpimag6is obtainedwilh a 10OXolflmmerslonobiective
Cau36
Conection
Remarks
.No immersionoil was used,
.Use oil specifiedby the manufacturer,
.Unsuitableoil was used,
.Use specifiedoil, sincethe types
andnomal
usedfor fluorescence
white light oftendiifer
.The oil containedair bubbtes.
.Apply th€ oil aft€ryou hav€
removedbubblesln the botlle.
.Usingoil in a roomwithunsuitable
(toohighor low)
temperatures
.lf roomtemperaiure
is eithertoo
highor ioo low,o. if the air is too
humid,the diffiactionindsx
changes,causingchangesin the
imag€,Usethe oil at a room
temperatureof 22-25"Cand at
a humidilyof about56%.
.The specimenis too thick.
.lt is advisableto usea specimen
with a thicknessof 2'3r.
-
l,
r1S:.
'.o'*.
-j- p
!.'l
R€l6rto
page
Page18
Page18
.Removelhe eyepiecebefore
examinalion
andlookthroughthe
Page18
Page12
lllm ls nol sharp
4. Entireroll ot black.and.whlle
Cruse
.Possiblecausesare:type of film,
emursrcn,
overexposure,
overdevelopment,
improperhandling
or accidentsduringdevelopment,
Con€ction
Rema*s
Relerlo
oaQ6
Pages
.Check lhesepossibililiesand
B{- e,5
5.Th6linishgdpdnl app€arsgrainy
Caus€
.A fllmwlthcoarseorainwasosed,
R€mark5
Coreclion
.Use a finegrainfilm.
.Kodak Panatomic
X
.Kodak TechnicalPan2415
.Agfapan25
.lliord PanF
.A slandarddevelooer
was used.
.lJse a linegraindevelopet.
.Kodak Microdol
D-23
D-25
.Fuji MicrcFine
.Magniticationratiowastoo high.
aL,sea largsformatfilm.
.4" x5" sheetfilm.
Referlo
paq€
Page55
image
Objects
otherthanthespecimen
appeared
on thefilm
'1.Shadow.llk€
lmag€
Caus€
Con€ction
.Optical palh selectorot the
pholographlc
attachmgntor vinc
cularlube was intenupledat
.Engagethe opticalpathselector
at its prop€rposllion,
.The lield kis diaphragmwas
stoppeddowntoo much.
.Open the field kis diaphragma
littlewiderthanthe photographed
areaot the lindereyepieceof
the focusing telescope,
.Tlny bits of film, dirt,etc. stuck
to the p sm of lhe photographic
equipmentor to the largefomat
.Ch€ck for, and remove,dirt from
the prlsmot the photographic
atlachmentwhilethe shutleris
open(Iime setling).
.Flemovethe largefomal relay
lens and cleanit.
Remarl(s
Feterto
Daqe
Page23
.Periodicchecksare recommendedif a largenumberof
photographs
is taken,
Pages
2931
I
Cau3e
.Dirl In the opticalsystem
Corectlon
alocate the dirt and removeit.
Cause
.When movingthe film or when
unrollingthe backingpaperof
the fllm, staticelectricitycauses
spafis.
Con€clion
R6ma*s
.Yo0 can locatethe dirt by moving
and rotatingeachcheckpojnt,
alt€natelylooklngthroughthe
blnoculartube,the focusing
telescopeof the photographic
attachmsntand the lllm plane
(by placlnga pieceof frosted
glass in the camsra body.)
R€lerlo
Page
PageI
Pages
30-31
R€l6rlo
page
.Do not rewindlhe film too rapidly.
.Keep humidityat zl5%minimum
in the roomwhereyou handle
the tilm.
aMakesurethat the camen back
and the darkroomars lree of dust,
3. Rellocllonol windowor roomilluminatlon
Cause
.Stray light gntsrs from the ey€"
piecesor lhe focuslnglelescope.
Coneclion
.Move lhe oplical path selector
of the tdnoculartub€io lhe
Camera1m7oposilionandcover
the locusingtelescopeof the
photographic
atlachmenlwilh
a cap.
aPut capson both eyepl€ces
and
the locusingtelescopeof the
photographic
atlachment.
.Set up the mlcroscopein a
dlllerentlocation.
Referto
Pag€
Page12
Uneven
brightness
1. lJn€yenareas occu. on one sld6 of th6 fram€, in lhe c€nt€r,and under the pertoratlonof th€ film
Cause
R€ma*s
Con€clion
.Th€ microscopelight sourceIs
not prope y centered.
.Prope y adjustthe light source.
.The field iris dlaphragmis off
axis.
.Clos€ th6 fleld his diaphragmso
that it appearsin the visual field
and adjust the conden6erto
center the diaphragm.
.The optical syslem is contaminated by dirt.
.Clean the opticalsystem.
.Dgvelopment problems(on blackand.whltepint)
.Developthe film prcperly.
@
R€ler
paqelo
Page21
Pages
29-31
.8e aware lhal a change In l6mpe€ture occurs betweenthe
center and the pedpheryof the
tank as a resultof hgatconductlon by th6 sl€€liank
Pages
64-65
2. Ma*s on the nogative
Caus€
.Fixing time was too shortor exhausted fixer was us€d.
ConEction
.lncrea6etixingtime or use a
new lixet
Roma*s
RaLr
lo
paEe
OLYMPUS
Matc[ yout skil]s rrith ours.
OI.YMPUS OPT|CALCO,
LTD'
OTYMPUS OPIICALCO{EUFIOPA)
GMEIH
OLYMP(IS CORPORATION
P' nied ln Jap.n M132E.123,1I