Thanks, I did, with 50% nitric acid. I would have thought that if there was a problem with the glassware it would have shown up in the silver nitrate overnight. The silver solution was clear after incubating. The milky mess didn't happen until after washing in 95 ETOH then placing in the developing solution. Just to be sure though I'll make sure to clean the glassware longer than last time. I'll also run a couple of controls and take one out after an hour to reduce just it really is over impregnated.
Thanks again, Amos Message: 16 Date: Wed, 1 Dec 2010 20:58:20 -0600 From: "Ingles Claire " <cing...@uwhealth.org> Subject: RE: [Histonet] Hellerstrom & Hellman anyone? To: <histonet@lists.utsouthwestern.edu> Message-ID: <f2f030053f9b7345831bed293a6d57e103a1a...@uwhc- MAIL01.uwhis.hosp.wisc.edu> Content-Type: text/plain; charset="iso-8859-1" Did you acid clean your glassware before hand? Claire ________________________________ From: histonet-boun...@lists.utsouthwestern.edu on behalf of Amos Brooks Sent: Wed 12/1/2010 12:07 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Hellerstrom & Hellman anyone? Hi, I just did a Hellerstrom & Hellman alcoholic silver stain for pancreatic D- cells. I followed the method described by Bancroft & Stevens. When the slides were placed into the reducing solution they instantly precipitated into a toasty mess, the solution turned milky brown and the slides (even around the sections) picked up a lot of silver precipitate. If anyone has done this of could offer suggestions about it, I'd really like to trouble shoot the stain a bit. A bit of background about the stain that I thought I should add. According to the published method I used a 10% silver solution in ETOH with 100ul of 1M nitric acid. The pH of the solution was 5.0. The reducing solution was alcoholic formalin with 5% pyrogallic acid. I thought the silver solution seemed really concentrated (It hardly even disolved). Incubating such a solution overnight at 37 deg C (as indicated in the method) seemed a bit much. Has anyone heard of modifications of this stain? There is no sense reinventing the wheel, but I certainly don't want to repeat it knowing that it will innevitably do the same thing again unless there is something glaringly obvious that I did wrong. Thanks, Amos _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet