TS Procure 812 - primary fixation Karnovsky's - transwells (TM - 013)
sandra.crameri
Abstract
The method is used to Karnovsky's EM fixed transwells to Procure 812 resin blocks. The presence of 4% paraformaldehyde is only used to satisfy microsecurity requirements.
Steps
HEADER
SAN:
SPEC No:
OPERATOR & STEPS:
OPERATOR & STEPS:
Conventional
All solutions are exchanged above and below the membrane, up to step 10.
Karnovsky's fixative: 2.5% volume 4% volume 0.1Molarity (M) Sorenson's Phosphate buffer (pH 7.2, 300mosmol/kg) for at least 0h 40m 0s or overnight
0.1Molarity (M) buffer 0h 15m 0s
1% volume 1h 0m 0s
70% volume 0h 20m 0s or overnight
95% volume 0h 20m 0s
100% volume 0h 20m 0s
100% volume 0h 20m 0s
Remove membrane
Remove membrane from transwell scaffold using a 45o scalpel and transfer to a glass vial with 100% volume
50% volume 50% volume 0h 20m 0s
100% volume 0h 20m 0s
50% volume 50% volume Procure 812
100% volume Procure 8120h 20m 0s
100% volume Procure 812 1h 0m 0s
Embed & polymerise overnight in oven at 60°C , section with diamond knife as usual.
