[Histonet] Sirius red with low contrast between cells cytoplasm andstained fibers

[Tony Henwood]

Including hydrochloric acid in your staining solution might help (0.25ml per 100ml).

Regards 

Tony Henwood JP, MSc, 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: Guillermo Palao [mailto:gpbnas <@t> yahoo.es] 
	Sent: Thursday, 6 April 2006 1:51 AM
	To: Tony Henwood; Histonet
	Subject: RE: [Histonet] Sirius red with low contrast between cells cytoplasm andstained fibers
	
	
	Thanks, you advice going straight from sirius red to 70% ethanol or maybe to 100% as suggested in other reply, but how do I get rid of the excess red stuff on the slide?
	 
	Best regards,
	 
	Guillermo
	
	Tony Henwood <AnthonyH <@t> chw.edu.au> escribió:

		Drop step 3 and 4 (picric acid is soluble in water)
		
		Regards
		
		Tony Henwood JP, MSc, 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: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.eddu] On Behalf Of Guillermo Palao
		Sent: Wednesday, 5 April 2006 4:59 AM
		To: Histonet
		Subject: [Histonet] Sirius red with low contrast between cells cytoplasm andstained fibers
		
		
		Hello all:
		
		I would really appreciate any suggestions to improve sirius red staining of kidney samples. Although collagen fibers in tissue nicely stain red, cells cytoplasm do not stain yellow, but sort of pinkish, making it very difficult to separate from red-stained extracellular fibers for quantification purposes.
		
		My protocol is as follows:
		
		1. Deparafinize and hydrate samples
		2. Stain 1 h in 0.1% Sirius Red in saturated aqueous picric acid
		3. Wash 2 min 0.01 N HCl
		4. Rinse in water
		5. Dehydrate and cover slides
		
		I have tried shorter staining times and rinsing with water before HCl washing without improving results. Any help would be greatly appreciated.
		
		Guillermo
		
		
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