RE: Trephine procedures

From:Tony Henwood

RE: Trephine procedures

I would love to know what everyone else is doing?
The following is our procedure:

Bone Trephine Procedure

There are many methods for the preparation and examination of trephine biopsies. This method was chosen for its speed and quality of section.


1. Fixative (Formal Alcohol)
40% Formaldehyde                200ml
95% Ethanol                     1800ml
Add 3 drops of 1% eosin to colour the solution

2. Formic Acid/Formalin Decalcification Solution (purchase from Pathtech Cat No ADF.J)
        (4% Formaldehyde, 33% Formic acid, 0.85% Sodium Chloride)


1. Allow trephine to fix in formal alcohol for 3 1/2 to 4 hours
2. Rinse the trephine in water and place in Decalcifying solution for 1 hour.
3. Rinse trephine in water and place in biopsy cassette.
4. Place in 10% formalin to await processing.
5. If calcium is present when cutting, then surface decal in RDO for 20mion, wash and recut

1. Blake, J (1970) "Histological Methods of the University of Melbourne" p15.

-----Original Message-----
From: []
Sent: Thursday, 29 August 2002 23:11
Subject: Re:

Hello everyone.  I have been hired to start a histology / immuno lab for specifically bone marrow processing.  I know that there are many different methods in use and I just wanted to pick some of your brains for some ideas. Currently, this lab is using Bplus fixative (zinc formalin) for at least 2 hours.  Then they use a decal sol'n called RapidCal(hydrochloric)for approx. 2 hours.  Next the samples are sent to Histology where they rinse and load onto their tissue processor.  They use 10% NBF 2 changes 45 min each, 70% etoh 1 change 45 minutes, 80% etoh 1 change 45 minutes, 95% etoh 2 changes 45 minutes, 100% etoh 2 changes 60 minutes, 100%/xylene (50/50 mix) 1 change 45 minutes, xylene

2 changes 45 minutes, then paraffin 3 changes 50 minutes.  If you were to have a lab where you only processed bone marrow biopsies and aspirates for routine h&e's special stains and immuno's how would you do it.  We want to stay away from the Mercury based fixatives and we want to keep our turnaround time the same.  So any suggestions will be

Trevin Skeens
Jeb Monasterial

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