May 09, 2023

TS Spurrs - FFPE block - 1mm3 cube cut out - tissue (TM - 013)

DOI
https://dx.doi.org/10.17504/protocols.io.kxygx968wg8j/v1
  • 1CSIRO ACDP
sandra.crameri
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DOI: https://dx.doi.org/10.17504/protocols.io.kxygx968wg8j/v1
Protocol Citation: sandra.crameri 2023. TS Spurrs - FFPE block - 1mm3 cube cut out - tissue (TM - 013). protocols.io https://dx.doi.org/10.17504/protocols.io.kxygx968wg8j/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: May 09, 2023
Last Modified: May 09, 2023
Protocol Integer ID: 81632
Keywords: ts spurr, parraffin block, face to spurr, spurr, ffpe block, resin block, 1mm3 cube, ffpe, resin block for em, tissue, tm, piece, resin
Abstract
Processing unstained parraffin block that has had a 1mm3 piece disected out of the face to Spurr's resin block for EM.
Guidelines
All step times are the minimum time required. Longer steps are acceptable.
Protocol materials
xylene
Ethanol
Ethanol
Ethanol
Glutaraldehyde aqueous solution 25% EM Grade ampoulesProSciTechCatalog #C002
Osmium TetroxideProSciTech
ethanol
HEADER
SAN:


SPEC No:


OPERATOR & STEPS:


OPERATOR & STEPS:


Dewax FFPE sections
15m
xylene or X3B Overnight at 60 °C

5m
xylene or X3B 00:05:00 at Room temperature

5m
xylene or X3B 00:05:00 at Room temperature

5m
Rehydrate
15m
100 % volume Ethanol 00:10:00

10m
95 % volume Ethanol 00:10:00

10m
70 % volume Ethanol 00:10:00

10m
Conventional - with reduced times for 5um thick FFPE section
2h 50m
2.5% % volume Glutaraldehyde aqueous solution 25% EM Grade ampoulesProSciTechCatalog #C002 in 0.1 Mass Percent Sorenson's phosphate buffer 07.2 300 mosmol/kg00:20:00 .

20m
Wash buffer 00:05:00

5m

1 Mass / % volume Osmium TetroxideProSciTech in buffer01:00:00

1h
70 % volume Ethanol 00:10:00

10m
95 % volume ethanol 00:10:00

10m
100 % volume ethanol 00:10:00

10m
100 % volume dry ethanol 00:10:00

10m
100 % volume Spurrs:100 % volume dry ethanol in 1:1 ratio00:30:00

30m
100 % volume Spurrs01:00:00

1h
100 % volume Spurrs01:00:00

1h
Oven polymerise 65 °C Overnight