Histotechnology Technical methods

Enzyme Histochemical Stain

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NON-SPECIFIC ESTERASE

SECTIONS

Unfixed Cryostat, rectal biopsy.


SOLUTIONS

Soliution a) Pararosanilin-HCL Stock

--Pararosanilin 1g

--Distilled water 20ml

--HCL (conc) 5ml

The pararosanilin is dissolved in the distilled water and the hydrochloric acid is added. The solution is heated gently, cooled, filtered and stored in a refrigerator in 0.4ml aliquots.

Solution b) Incubating solution

Mixture (1)

--Pararosanilin-HCL (solution a) 0.4ml

--sodium nitrite (40mg/ml) 0.4ml

Add the pararosanilin drop by drop to the sodium nitrite, shaking well after each addition until the solution is corn-coloured after leaving for at least a minute.

Mixture (2)

--a-naphthyl acetate (10mg/ml acetone) 0.25ml

--0.2M Phosphate buffer (2.83g Na 2 H PO4/100ml) 7.25ml

--Distilled water 2.5ml

Add these together THEN add mixture (1) to mixture (2) filter and use immediately.


METHOD

1. Incubate in solution b) for 1-20 minutes at 37ÉC.

2. Wash, counterstain in haematoxylin, wash.

3 Dehydrate, clear . Mount sections in DPX


RESULTS

Esterase activity - red/brown


COMMENTS

a) The solution are best kept in aliquots for urgent use, especially with rectal biopsy for Hirschsprung's.

b) The reaction is rapid and it is easy to overstain.

c) A very good marker for ganglion cells.

NOTE: No responsibility is assumed by The University of Nottingham or the Queens Medical Centre NHS Trust for any injury and/or damage to persons or property as a matter of products liability, negligence or otherwise, or from any use or operation of any methods, products, instructions or ideas contained in the material herein. It is the users responsibilty to ensure that all procedures are carried out according to appropriate Health and Safety requirements.


© Copyright 1997 University of Nottingham Medical School Division of Histopathology. This page was last built on Mon, May 5, 1997 with Frontier. Thanks for looking in. Comments to James.Lowe@nottingham.ac.uk