[Histonet] H&E water contamination[Scanned]

[Kemlo Rogerson]

Plus xylene will tolerate low levels of water (very low) without going
cloudy; the contamination only is seen microscopically as a fine 'mist' of
droplets. Knew a Chief Technician that kept anhydrous copper sulphate in his
last alcohols to absorb any water carry over. 

If you try staining in very cold and damp conditions then a mist can form
from one alcohol to another. It's something to do with the eutectic point of
water or I may just be making that up. All I know is, even with fresh
alcohols, staining in cold damp conditions can result in water from the
atmosphere contamination the xylenes. The only way I could get round it was
to use a hair dryer to raise the temperature a bit, and hope I didn't catch
fire. I hasten to add I did that before CPA, etc., and not in my present
employ.  

Those were the good old days of sharpening knives with diamond paste and a
bit of wood, and making up your own stains and ripening the haematoxylin in
the window......... I must be old. I AM old!!!!

Kemlo Rogerson
Cellular Pathology Manager
East Lancashire Hospitals NHS Trust
DD. 01254-294162
Mobile 0774-9754194
 

-----Original Message-----
From: Tony Henwood [mailto:AnthonyH <@t> chw.edu.au] 
Sent: 28 November 2004 23:00
To: 9msm8 <@t> qlink.queensu.ca; histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] H&E water contamination[Scanned]

A few ideas.

Check your xylene for water.
Keep the alcohol and xylene containers covered when not in use. Alcohol will
absorb water from the air (possibly xylene as well - though of this I am not
certain).

Regards

Tony Henwood JP, BAppSc, GradDipSysAnalys, CT(ASC)
Laboratory Manager & Senior Scientist
The Children's Hospital at Westmead,
Locked Bag 4001, Westmead, 2145, AUSTRALIA.
Tel: 612 9845 3306
Fax: 612 9845 3318




-----Original Message-----
From: 9msm8 <@t> qlink.queensu.ca [mailto:9msm8 <@t> qlink.queensu.ca] 
Sent: Saturday, 27 November 2004 9:12 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] H&E water contamination


Hi histonet
I have been performing H&E stains on sliced parraffin embedded mouse
hippocampal slices. I am consistently getting water contamination in my
slides and have been unable to rectify the problem. My procedure for
staining is: Xylene I for 7.5 minutes; Xylene II for 7.5 minutes; 100%
ethanol I for 2 minutes; 100% ethanol II for 2 minutes; 95% ethanol I and
70% ethanol I for 2 minutes each; Water I for a second; then "H" for 20
minutes; Water II for 10 minutes; "E" for about 13 seconds, Water III for
about 1 minute; 70% ethanol II for 50 seconds; 95% ethanol II for 5
minutes; 100% ethanol III and IV for 5 minutes each and then Xylene III
and IV for 7.5 minutes each.
I beleive the "70% ethanol II" is the key step for removing water after
the staining but increasing the amount of time spent in the solution
hasn't worked yet (though I have yet to leave it in for longer then a
minute).
Any tips would be greatly appreciated,
Mike



_______________________________________________
Histonet mailing list
Histonet <@t> lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet


**********************************************************************
This email and any files transmitted with it are confidential and
intended solely for the use of the individual or entity to whom they
are addressed. If you are not the intended recipient, please 
delete it and notify the sender.

Views expressed in this message and any attachments are those 
of the individual sender, and are not necessarily the views of The
Children's Hospital at Westmead

This footnote also confirms that this email message has been 
virus scanned and although no computer viruses were detected,
the Childrens Hospital at Westmead accepts no liability for any 
consequential damage resulting from email containing computer 
viruses.
**********************************************************************


_______________________________________________
Histonet mailing list
Histonet <@t> lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet