rocedure for Sucrose Coat|ng and Cur|ng on G|ass S||des

SLep l reparaLlon of a 1M naCP SoluLlon and a Sucrose SoluLlon (lf already

prepared proceed Lo SLep ll)
(a)1M naCP SoluLlon

1) ulssolve 10g of naCP pelleLs (Slgma S3881300C) lnLo 230mL of dlsLllled waLer uslng a
1000mL beaker 1hls soluLlon ls a 1M naCP soluLlon 1he number of grams of naCP Lo
prepare a 1M soluLlon ls calculaLed below

I E
2

. o oE
I E
2

I E
2

I E
2

. oE
o oE
= oE

2) Soak Lhe vW8 Mlcroscope Slldes (CaL no 16004422) ln Lhe 1M soluLlon prlor Lo
sucrose coaLlng Lhem 1he soluLlon can be poured lnLo a glass dlsh Lhe slldes placed ln
Lhe soluLlon and alumlnum foll placed on Lhe Lop of Lhe dlsh

(b)Sucrose SoluLlon

1) uslng a 1000mL beaker comblne 200mL of dlsLllled waLer and 280g of sucrose (Slgma
S0389300C) Add a sLlr bar Lo Lhe beaker (found ln Lhe SLage l plasLlc bag under Lhe
fume hood) and place Lhe beaker on Lhe magneLlc sLlrrer
2) Ad[usL Lhe seLLlng Lo 400300rpm and allow Lhe sucrose Lo dlssolve for 310 mlnuLes
1he sucrose wlll noL fully dlssolve slnce Lhe soluLlon ls saLuraLed
3) 1urn off Lhe magneLlc sLlrrer and obLaln a vacuumdrlven dlsposable fllLraLlon unlL use
Lhe small vacuum pump by Lhe lamlnar flow hood
4) our Lhe soluLlon lnLo Lhe Lop parL of Lhe fllLraLlon unlL and aLLach Lhe Lublng LhaL's
connecLed Lo Lhe pump Lo Lhe slde arm of Lhe unlL
3) 1urn on Lhe pump Cnce Lhe llquld has empLled ouL of Lhe Lop conLalner and ls presenL
ln Lhe boLLom conLalner Lhe pump can be shuL off and Lhe Lublng deLached
6) 1he Lop conLalner can be dlsposed of ln Lhe garbage 1he blue cap should be fasLened
Lo Lhe Lop of Lhe conLalner Label Lhe conLalner and provlde Lhe daLe lL was prepared lf
Lhe soluLlon ls noL belng used lmmedlaLely Lhen place lL ln Lhe refrlgeraLor for fuLure
use
SLep ll CoaL Lhe Slldes wlLh Sucrose SoluLlon (lf already prepared proceed Lo SLep
lv)
1) 1ake ouL Lhe sucrose soluLlon from Lhe refrlgeraLor and obLaln a 3mL syrlnge
2) 8emove a glass sllde from Lhe glass dlsh conLalnlng a 1M naCP soluLlon ury Lhe
boLLom of Lhe sllde wlLh a klmwlpe and place Lhe sllde on Lhe sLlcky Lab locaLed on Lhe
cenLer of Lhe fan
3) LocaLe Lhe yellow wlre and aLLach lL Lo Lhe baLLery Allow Lhe fan Lo spln for 1 mlnuLe so
LhaL Lhe sllde can parLlally dry
4) AfLer 1 mlnuLe dlsconnecL Lhe yellow wlre from Lhe baLLery
3) Apply approxlmaLely 2mL of Lhe sucrose soluLlon on Lhe sllde 8econnecL Lhe yellow
wlre Lo Lhe baLLery and allow Lhe fan Lo spln for 30 seconds Lxcess sucrose soluLlon wlll
be removed from Lhe sllde leavlng a Lhln layer
6) AfLer 30 seconds dlsconnecL Lhe yellow wlre from Lhe baLLery
7) Carefully remove Lhe sllde from Lhe sLlcky Lab and wlpe Lhe boLLom of Lhe sllde wlLh a
klmwlpe lace Lhe sllde on Lhe Lop or boLLom shelf of Lhe vacuum oven noLe Make
sure Lhe shelves ln Lhe oven are covered wlLh alumlnum foll 8epeaL Lhls procedure for
mulLlple slldes 1hen proceed Lo SLep lll
SLep lll vacuum Cven CondlLlons for SucrosecoaLed Slldes (lf already prepared
proceed Lo SLep lv)
1) Cnce all slldes are placed ln Lhe oven close Lhe oven door and secure Lhe slde laLch
2) 1urn on Lhe oven by fllpplng Lhe green swlLch
3) Cpen Lhe venL valve all Lhe way by Lurnlng Lhe knob Lo Lhe lefL Close Lhe vacuum valve
all Lhe way by Lurnlng Lhe knob Lo Lhe rlghL
4) SeL Lhe LemperaLure Lo 120SC and allow Lhese condlLlons Lo remaln for 18 hours
3) AfLer 18 hours Lurn off Lhe oven Loosen Lhe laLch and keep Lhe oven door a[ar so LhaL
Lhe lnslde chamber can cool down
6) Cnce Lhe lnslde of Lhe oven ls near room LemperaLure remove Lhe glass slldes SLore
Lhe slldes ln a whlLe sllde box for fuLure use or proceed Lo SLep lv
SLep lv CasLlng CS on SucrosecoaLed Slldes
1) 1he amounL of CS prepolymer used Lo creaLe a membrane wlLh a Lhlckness of 230m
on a glass sllde was calculaLed

o o on o s = . = 8.
3
= .8c
3


p
PuS
=

- = p = . c
3
/ .8c
3
= .89 s /

2) 1ake ouL a [ar of prepolymer from Lhe deslccaLor and measure ouL close Lo 03000g of
CS on a sucrose coaLed sllde uslng Lhe ML11LL8 1CLLuC welgh scale
3) 1ransfer Lhe sllde over Lo Lhe hoL plaLe/sLlrrer near Lhe vacuum oven MelL Lhe pre
polymer aL 160C and smooLh ouL Lhe llquld over Lhe enLlre surface of Lhe sllde
4) Carefully Lransfer Lhe sllde Lo Lhe vacuum oven and place lL on Lhe Lop or boLLom shelf
noLe 8emove Lhe alumlnum foll from Lhe shelf prlor Lo placemenL of slldes 8epeaL Lhls
process for addlLlonal slldes and Lhen proceed Lo SLep v
SLep v vacuum Cven Curlng CondlLlons for CS
1) Cnce all slldes are placed ln Lhe oven close Lhe oven door and secure Lhe slde laLch
2) Make sure Lhe oll level ls beLween Lhe hash marks on Lhe vacuum pump lf lL lsn'L Lhen
flll Lhe pump wlLh enough oll unLll Lhe level ls beLween Lhe hash marks 1ake off Lhe
plasLlc elbow [olnL lnserL a funnel and pour Lhe oll lnLo Lhe funnel
3) Apply vacuum grease Lo Lhe Lop porLlon of Lhe [olnL LhaL lnserLs lnLo Lhe condenser
4) llll up Lhe uewar flask wlLh llquld nlLrogen from room 812 lasLen Lhe blue blankeLs
around Lhe Lop of Lhe flask and secure Lhem wlLh a LwlsLy Lle
3) 1urn on Lhe oven by fllpplng Lhe green swlLch
6) SeL Lhe LemperaLure Lo 120C or 160C based on Lhe followlng curlng Llmes (as of
7/13/10)

C
16 hours
12 hours
10 hours
8 hours
6 hours

7) Close Lhe venL valve all Lhe way by Lurnlng Lhe knob Lo Lhe rlghL
8) 1urn on Lhe vacuum pump and gradually open Lhe vacuum valve by Lurnlng Lhe knob Lo
Lhe lefL Cnce Lhe dlglLal dlsplay lndlcaLes a pressure of approxlmaLely 283 ln Pg close
Lhe valve by Lurnlng Lhe knob Lo Lhe rlghL 1hen open lL agaln wlLh Lwo 180 Lurns
9) Allow Lhese condlLlons Lo remaln unLll Lhe deslred curlng Llme ls compleLed
10) Cnce Lhe deslred curlng Llme has been reached Lurn off Lhe vacuum pump Slowly
open Lhe venL valve by Lurnlng Lhe knob Lo Lhe lefL Cpen Lhe valve all Lhe way unLll 00
ln Pg appears on Lhe dlglLal dlsplay
11) Close Lhe vacuum valve by Lurnlng Lhe knob Lo Lhe rlghL
12) lllp Lhe green swlLch Lo Lurn off Lhe oven loosen Lhe slde laLch and keep Lhe oven door
a[ar Lo cool Lhe lnslde chamber Lo room LemperaLure
C
24 hours
48 hours
13) 8emove Lhe blue blankeLs from Lhe uewar flask Lo allow any remalnlng llquld nlLrogen
Lo evaporaLe
14) Cnce Lhe lnslde has reached room LemperaLure remove Lhe samples from Lhe oven
lace Lhem lnLo separaLe eLrl dlshes and puL a plece of Lape on Lhe lld of Lhe dlsh
WrlLe Lhe sample name curlng LemperaLure curlng Llme and daLe on Lhe plece of Lape
SLore Lhe eLrl dlshes ln a deslccaLor unLll Lhe membranes are needed
SLep vl Cleanlng up
O lf Lhe shelves conLaln any polymer or sucrose resldues Lhen Lake ouL Lhe shelves and
clean Lhem wlLh 70 eLhanol Make sure all of Lhe eLhanol ls removed from Lhe
surface afLer wlplng Lhem
O lf Lhere ls any waLer presenL ln Lhe condenser Lhen carefully remove lL from Lhe Lop
parL of Lhe unlL and pour ouL Lhe waLer 8lnse ouL Lhe condenser wlLh waLer and seL
lL on Lhe lab bench Lo dry
O Wlpe off Lhe vacuum grease on Lhe lnserLlng componenL of Lhe condenser unlL