Slide preparation

1. Date.3RaaAL
xpt. No. PageNo.. *r*nssseara*****esrnsa"
ICROTOMY REPARATIONDE fERMANENTSLIDE UE MaMMALIAN ISSUERAT
INTODDCTON
biecHo bestudie2undea micioscope nee prepouadim.f
theohject issmallencughthroughahich ightcanpond,anðtha.
Studyintinest detailk isnat uenpiseda whale moun prep.asuatimuil
beSuficientln caae 0f laMge ojec they ae do_becukinthin
Sectioastor the study his iscalled histolosica prepauatins
ha objectiedhistological pHepauation is toundenstnnd
th micro.anatomy 0t celli, TisSues,andongand and teCaralala
Hha Structune dth suncdin
Haematoayun A ~Asin Stoining pxctecol_is weameaant-
tn histoloqyto examine Thin SectimA,Ahismehedknownas
Double-Staining Method
Teachers Signature..

2. Date
No PagyeNo
REQURED MATERLS
Jfssues collecte Axom live specimen
Bouin's uid (aqmeousonalcoholic-
Distilled wcte
ades of alcakoleg 5o,30%,q0, L100/)
Xylal
Para-tin
:Haematoxylin and tosin Stains
Kectanqulo block making maulds
APX OH Canada balsam
7
ylam slides
ove slips-
Teachor's Sinnatu

3. Date
pt No
ROCEDUE
A.CoLLECTION OEJissuE
Fo Sectfoninq, issues are to be collectedfrom ive specimera. In
Small vetebrates,the animal is paralysed by damaging te brair.
Sn aqen vextebrates,the dame. Hesult isachieveo by staikinq
heae aqainst a hrdobfec ha tssues anu cokécies immediate
afte killing tho animal.
FIXING
Kfnq the tissue aithout anyStuuctuual ah chemical changa isknauan
O&xing.
Jhetissues ate Cut inte small pieces,lashed with phystological soluho
eauiue, immersedinthe xativeandlost thea soradeiniliE
boniodhemast (ommon fixative une3isBouin'sHuidauesuslalcehoi
C ASHING
Ohio is uDually doneaith Juate til t aeaqent ú empletely Hemaveà.
SThepe1centage otwaten in the xative ia belous50,itghouldbe
JAlashedonlyîn solutien Hecom.mended fothe puupose
D DEHVDRATIONL
he small pieces ot tissue aa cateied through dittxent guadesd
alcahal,eg 5010,90,2 loo,HoDloLoen quae to ppe g9uade ond
TM
en ultimately toahsoulealcohol he_poxiod o tegbmentuih euchgade
clependingondhe size oftissue luechangeAaa ivan inabsoluula olcahol.
Teacher's Signature..

4. No
E CLEARING
Jhe tisu îs transfemed to Cecde. uoo oi, utMin 2 ous ha tiu
ns thandpasentE and ana 1eady {or î'ntifratidn
F INEILIRATION
Jhis is to Heplace oil with paiatin. The intecelluluu Spare ane
Completely occupied bypaattin
he tissua. is ocahedinxylol andtranahehed to a
taveed pn celain
CYucible Containing Xulol and mo lten paraftin ar aHotit 1 and ha
C3utbleis putinatempetature Contanalled poartin badh or o 1S
minuteaAHerthopanio, thdissu fs transhe1ed to ancthen ave2
Caudecontaúning molttn tadfin anly, kep in hasome bath
G. BLocK-MAKINGa PaaceDu2E
Molten paxafin ispousredin_ablack_makingmould_andihetisauaixpu
nth midale dt AHe dhaidening,extra poratin is treameðtn mave
a Hectamgulan block_keeping the tis.sue in themiddle
h SeeTioN CuTTIN
hinis clone witha microiomeSectionson otine ulonKarauLally
Ccutb-6Lmthick A queanefuee suide is tuken,oneauace is
wmaxkedwitHaadiamona_pentl Adoplet ot Mayersalbumenissnat
onthe mcnkedsduace. Athinfilm otdotenisput anthe slide andthu
P.aa tin uibbon _aith sectionsaxaplaces on it.ke side isheated
ona hot plala and the sechona ane propenly stmetched
TM
n
h wate isdraned otf andthe ide isdried on the hot 8lal
Teacher's Signature... ** * * * * * * * * * * * * * *

5. Date
Page N
1 STANING OF HisToLOGICAL SLIOES
ections of tissues ae
usually Stained wi t tuao dye to
oxdny
CcntscasbE betuoeen dithetent histologca stsuuctudu.Ihis rna
detciled study ecoie
SAcining i tuoo dyes s knaen an doubla daining hi mos
COnmon double SAaining practised in clam woHk 1s th heemaloryin
and ensin. Jhe haematbryliw oeinqbasic dye împaul blua Caleuu t
acidc mateyialb,i.e.,nucleicacids, whichaxo Cencentsucla in tha
nucleuo andonly scaHeued 1nthe uoplaon
lhe eocin isan acia due and the toplaomic mateials being
bapic innatu a Stained hy it.Nho ceult a,he nucles
andonu a smal 4uaciion Othe toplaom oppeav du,
hil tha Heat d the ceall takesup s2colown.
TM

6. Date
No. Page No
STAINING ECHNIQUE
wha slde wit the panaffin aibbon Contodning Sections of .
is dipped in xylol fon cabout 5 minuls.
CAte the panatfin getcdissolvea,tho slide is transherdhed
absoui alcehal
Sh slide isthen Cannieo through qHades oaloho) from upper to lee.
gMadeie joo,9o,t07A501
imeineach guadeor_altohal isabout 9-5 minuli.
hen thaalide is transheroned to tasdistilledwate and net to tha
haemataxylin Stain o_albout 30seconds to min
At aDip_in. haemaiaxylinu,the uideiswashedln tapkat
) aenagaintha slidefscasthiedthraug gMadesofalcahal,hremlonert
highe qrades orabaut 5-0minutes ineachgxade
in Atenihat,the slide is then dipped in eosin fom too noua
han theslide.is washedin90? aloholfollouedbytuoochangesinabzauli
alcehol
Afte Reeping the slidein Xylol foHabout minule,Hhe blideu Heady
oH mounting
kDASmal_amauntot Canada.balsom_o àPxisputan thaulide,depending
onheSizececavenslip tobeuned JhiCouehslip s put oniE
gi air bubbles ax lockedbetoeen thecoveneslipandthe slde,itmaybe
Hemoved by leaving the slide oveHniGht ona hat plala.
ail)hen the gide lsheady and istaatched unden compound mícrascap£e
Hostudy thedetailed siudune ofhesue.
Teacher's Signature..
16

7. AvnaffinSection
Mounhng
on slide
ylol
Xylol
lo aloho)
5min.
qo% aluhol
Casn)
die
loo/ altohol
0 min
osin
90/o alcohol
qo% odcoho)
10miw
o alco hol
40,alcoho
1ominv S0 alcohei
50% alcohol
tomin Distilled wa
Distilled walu
Alkaline wal
Snin fou niny)
Haematoxlin 3sec laP wal A A etJoaten
Cuoas Citferentiatiar)
FiG- Bouble staining, e hod (Suwmart)