[Histonet] Dirty H&E after break

[Tony Henwood (SCHN)]

Hi Allan,

With the quality of the eosin dyes today, filtering should not be required. The same for xylene and ethanols.

You will need to monitor the usage and solution levels (evaporation being the issue here).

There are no hard and fast rules but a suggestion is one slide per ml of solution. Therefore a bath of 400ml of haematoxylin should stain 400 slides consistently before a noticeable deterioration in staining is noted. The same with the other solutions.

Regards
Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA)
Principal Scientist, the Children’s Hospital at Westmead
Adjunct Fellow, School of Medicine, University of Western Sydney
Tel: 612 9845 3306
Fax: 612 9845 3318
Pathology Department
the children's hospital at westmead
Cnr Hawkesbury Road and Hainsworth Street, Westmead
Locked Bag 4001, Westmead NSW 2145, AUSTRALIA

From: Allan Wang [mailto:allanvv at gmail.com]
Sent: Saturday, 7 January 2017 4:01 PM
To: Tony Henwood (SCHN)
Cc: histonet at lists.utsouthwestern.edu
Subject: RE: [Histonet] Dirty H&E after break

Yep, filtering the hematoxlyn fixed it. Thanks for the tip.

Is it worthwhile to filter the eosin, alcohols, and xylene substitutes as well?

Allan


On Jan 4, 2017 1:04 AM, "Tony Henwood (SCHN)" <tony.henwood at health.nsw.gov.au<mailto:tony.henwood at health.nsw.gov.au>> wrote:
Hi Allan,

The flakes are alum precipitates.

I would suggest filtering the haematoxylin before use.

Regards
Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA)
Principal Scientist, the Children's Hospital at Westmead
Adjunct Fellow, School of Medicine, University of Western Sydney
Tel: 612 9845 3306
Fax: 612 9845 3318
Pathology Department
the children's hospital at westmead
Cnr Hawkesbury Road and Hainsworth Street, Westmead
Locked Bag 4001, Westmead NSW 2145, AUSTRALIA

-----Original Message-----
From: Allan Wang via Histonet [mailto:histonet at lists.utsouthwestern.edu<mailto:histonet at lists.utsouthwestern.edu>]
Sent: Wednesday, 4 January 2017 4:45 PM
To: histonet at lists.utsouthwestern.edu<mailto:histonet at lists.utsouthwestern.edu>
Subject: [Histonet] Dirty H&E after break

Hello all,

Does someone know what would cause dirty flakes of light purple that showed across the whole slide?

http://i.imgur.com/1nIXwAy.jpg

Reagents are relatively fresh but the H&E stainer hadn't been used for a week over the holiday break.
Is it surface film on the hematoxylin? I didn't notice anything in the container. We are using Mayer's, followed by differentiation with HCl acid alcohol, with 2-3 minute long wash steps in between.

Thanks,
Allan
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