[Histonet] RE: Brain Problems

[McMahon, Loralee A]

How big are these rat brains?  Are you processing by hand or with a machine?  
We routinely process pieces of human brain for about 10 to 11 hours on a VIP.  This seems like a really really long cycle.  No heat and no vacuum until the paraffin infiltration step. (formalin, 70% EtoH, 95% EtoH, 95% EtoH, 100% EtoH, 100% EtoH, Xylene, Xylene, paraffin, paraffin)
You might want to back it off to one hour in each step and then see what you get.  You may be able to get away with an even shorter cycle if the pieces of brain are smaller.

Loralee McMahon, HTL (ASCP)
Immunohistochemistry Supervisor
Strong Memorial Hospital
Department of Surgical Pathology
(585) 275-7210
________________________________________
From: histonet-bounces <@t> lists.utsouthwestern.edu [histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Poole, Kimberly [Kimberly.Poole <@t> drdc-rddc.gc.ca]
Sent: Tuesday, July 27, 2010 10:27 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Brain Problems

Hi there,



I am currently using rat brain for histology but something is not working. I follow the protocol below for tissue processing.



Station

Solution

Routine (mins)







1

Formalin

10

2

Formalin

10

3

70% Ethanol

60

4

80% Ethanol

240

5

96% Ethanol

240

6

100% Ethanol

240

7

Xylene

120

8

Xylene

120

9

Xylene

120

10

Paraffin Wax

120

11

Paraffin Wax

180

12

Paraffin Wax

180

TOTAL TIME



27 hours 20 minutes









I embedded my brains and then cut them afterwards. When the brain section hits the water, it looks like a blob of fat on the water. Then it spreads reall, really slowly. If a cut is made to the paraffin the brain bursts open and it falls apart. Am I doing something wrong in the processing stage? Is the tissue dehydrated too much. Any feedback would be greatly appreciated.

Kimberly Poole B.Sc

Casualty Management Section | Section de la gestion des blessés

Defence Research and Development Canada Suffield | Recherche et développement pour la défense Canada Suffield

Medicine Hat, AB, Canada T1A 8K6

kimberly.poole <@t> drdc-rddc.gc.ca <mailto:kimberly.poole <@t> drdc-rddc.gc.ca>

Telephone | Téléphone 403-544-5347 / Facsimile | Télécopieur 403-544-4714

Government of Canada | Gouvernement du Canada

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