----------
	From: 	Edna_J_Gonzalez/Powderject@powderject.com
	Sent: 	Tuesday, September 28, 1999 10:23 AM
	To: 	vbaker60@yahoo.com; valleygal@aol.com; HistoNet Server
	Subject: 	staining



	I was staining sucessfully my paraffin block with pig skin samples, the
	color of the hamatoxylin and the eosin was perfect. I s\don't know what
	happened but know is different. After staining in the hematoxylin,
rinsing
	in running tap water, I dip the slides two times in 1% HCl in 70%
ethanol.
	At this point the slides will turn light red or pink, but now they are
not
	turning to light red, they stay purple. I haven't change the procedure
at
	all. I just replaced the reagents with fresh ones. This hasn't happened
	before to me. Can someone help me with this? Was is going on?

	I was using the AFIP method:
	2 changes in xylene for 2 minutes each
	2 changes of absolute alcohol for 2 minutes each
	2 changes of 95% ethanol for 2 minutes each
	70% ethanol for 2 minutes
	distilled water for 30 seconds
	Harris' hematoxylin for 6 minutes
	running tap  water for 2 minutes
	2 dips in 1% HCl in 70% ethanol
	running tap water for 1 minute
	.3% ammonia water for 30 seconds
	running tap water for 10 minutes
	70% ethanol for 2 minutes
	eosin for 1-2 minutes
	2 changes of 95% ethanol for 2 minutes each
	2 changes of absolute alcohol for 2 minutes each
	2 changes in xylene for 2 minutes each.

	This happened to me once. I checked everything and even got rid of good
reagents. What it ended up to be was the HCI was old. So get a fresh bottle and
remake the Acid Alcohol and everything should be fine. Run a control slide( just
some unstained slide) first and recheck.

	Ruth Yaskovich 
	N.I.D.C.R. N.I.H