RE: Myelin processing


We usually intracardically perfuse the mice using 4% paraformadehyde(rinse
blood vessels by saline before that) about 10 min, then postfix 1-2 hrs in
the same fix. The programs we use in Leica TP1020 are as follows:

Adult mouse brain:
STATION#	Reagent	Vaccum	Time
1	70%EtOH	off	25min
2	80%EtOH	on	25min
3	95%EtOH	on	25min
4	95%EtOH	on	25min
5	100%EtOH	on	25min
6	100%EtOH	on	25min
7	100%EtOH	on	25min
8	Xylene	on	25min
9	Xylene	on	25min
10	Xylene	on	25min
11	Paraffin	on	45min
12	Paraffin	on	45min

Small Large Nerve(rat nerves or mouse sciatic nerve)		
STATION#	Reagent	Vaccum	Time
1	70%EtOH	off	10min
2	80%EtOH	on	10min
3	95%EtOH	on	10min
4	95%EtOH	on	10min
5	100%EtOH	on	10min
6	100%EtOH	on	10min
7	100%EtOH	on	15min
8	Xylene	on	15min
9	Xylene	on	15min
10	Xylene	on	15min
11	Paraffin	on	30min
12	Paraffin	on	30min

Usually the specimens are hold until the "DELAY" program runs in the late
night.Because this processor only has 2 stations for paraffin(previously the
same protocol which has three paraffin stations,30 min each, worked well in
other brand processor) we modified to 45 min each. We found some area in
brain and spinal cord still have problem of keep the myelin structure. Any
suggestions about this?

Xiangjun Chen
Department of Neurology
University of Chicago

-----Original Message-----
From: Pamela Marcum
Sent: 4/17/2003 7:43 AM
Subject: RE: Myelin processing

Good Morning,
Could you please send or publish your current processing schedule with
reagents, times, temperatures and vacuum?  This will help understand
the problem may be and why it is happening.  Just from your question I
cannot do anything except hazard a guess and that is not a fair way to
If you just put the program on HistoNet with you question we can move
faster.  Thanks,  Pam Marcum

> -----Original Message-----
> From:
> []
> Sent: Wednesday, April 16, 2003 9:14 PM
> To:
> Subject: Myelin processing
> Does anybody there know of the paraffin tissue processing
> protocol for mouse
> CNS and PNS using Leica TP1020 automatic tissue processor? We are
having a
> hard time to preserve the myelin structure of mouse tissue. Any input
> highly appreciated.
> Xiangjun Chen
> University of Chicago

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