Hello experts, *Some background:* I know that Congo Red can bind nonspecifically to non-amyloid components such as collagen and elastin under certain conditions (eg Carnoys fixative, insufficient differentiation, insufficient alkalinity, etc). However, everything I have been able to read on the topic suggests that over-staining is "easily" differentiated from true amyloid staining by using polarizing light microscopy. That is, true amyloid produces apple green fluorescence while non-amyloid components produce silver/grey color.
*My question:* I want to know if anyone has encountered false positive staining that *is apple green* in color? We had a few bone marrow core biopsies that stained bright green but were later found to be negative when stained at another lab. We subsequently threw out all of our working solutions and made up everything fresh and repeated the previous (false positive) specimens and they were indeed negative in our lab as well. *In order to prevent this from happening again, I need to attempt to understand what may have caused this to happen in the first place. * This is where the vast collective knowledge of this group comes in. :-) Can anyone offer some insight as to possible causes? *Our Congo Red method:* Deparaffinize sections and bring them to water. Stain in Hematoxylin for 1 minute Add 0.5ml of 1% Sodium Hydroxide to 50 ml of stock alkaline salt solution. Wash slides in running water Place in *working* alkaline salt solution from step 2 for 20 minutes Add 0.5 ml of 1% Sodium Hydroxide to stock Congo red solution. Start to filter *working* Congo red solution when 15 mins are left in step 6 Place sections in the *working* Congo red from step #8 for 20 minutes. Dehydrate the slides one at a time in 3 changes of absolute ethanol, 6 dips each. Dip the slide 10 times in a coplin of xylene. Continue dehydrating the other slides. Coverslip the slides. *Greg Dobbin* 1205 Pleasant Grove Rd Route 220 York, PE C0A 1P0 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
