I do Weigert's staining for 10 minutes and use it for a week at the most. I, too, have heard conflicting times about the stability of the Weigert's, but have the best results with using it no longer than a week.
Deanna Rhoads HT (ASCP) ________________________________ From: Andrea Marion <amar...@uic.edu> To: histonet@lists.utsouthwestern.edu Cc: itai.mo...@mail.huji.ac.il Sent: Fri, September 17, 2010 10:31:16 AM Subject: [Histonet] Re: Masson's trichrom - problem with nuclei staining Hi Itai, I am interested to hear if you've resolved this problem. We use the same kit to stain mouse heart and embryo sections, PFA fixed. Our protocol is similar to the one you mentioned. I cannot get nuclei staining with this method either. The nuclei are well stained (ie black) up to the PMA/PTA step, but during that step the nuclear stain is completely removed. I cannot shorten the PMA/PTA step without negatively effecting the collagen stain. I have in our original protocol that the Weigert's working solution is good for one month, but I cannot recall if this is from Sigma's specification sheet or a personal observation from a lab member. However, from reading online some say it is good up to 4 months, others say it needs to be prepared fresh each time. I have tried fresh preparations with the same results. My instinct is that something is off - the staining is just not stable enough to withstand the subsequent acid steps in Masson's trichrome. Can an expert weigh in on this? Is there a way to strengthen nuclear staining from Weigert's? Sigma's formulas and usage recommendations are: Part A: 1% w/v certified Hematoxylin in ethanol Part B: 1.2% w/v Ferric chloride, 1% v/v Hydrochloric Acid Combine equal volumes Part A and Part B, stain sections for 5 minutes. Andrea Marion Graduate Student University of Illinois at Chicago amario3 /at/ uic /dot/ edu Itai Moshe itai.moshe <@t> mail.huji.ac.il Wed Sep 15 11:34:51 CDT 2010 Hi Histonet's I'm using Masson's trichrome to stain paraffin mice diaphragm fixed with PFA. I'm using this protocol: http://www.bcm.edu/mcb/rosenlab/index.cfm?pmid=12997 <http://www.bcm.edu/mcb/rosenlab/index.cfm?pmid=12997>With sigma masson's kit (#HT15) and Weigert's Iron Hematoxylin Set (#HT1079) The staining is beautiful, but i can't see the nuclei good enough. 1) Is there a way to enhance the nuclei staining ? (the nuclei is the only reason that i"m not using the simpler sirius red and fast green staining.) 2) What is the meaning for washing in running tap water washing, is it done by putting the slides in a jar with simple tap water for a few minutes ? Thank's Itai P.S By mistake I've post this before in another message with a wrong title. please respond to that message, so the title will be o.k for future archive searching. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet