Dear Rajesh, please at least give a reference for your "old protocol", if you don't have, at least the whole steps of procedure you performed,

best regards, Wolfgang

You can find the staining protocol here:

http://www.ihcworld.com/_histologic/pdf/73_july.pdf

and here:

Theory and Practice of Histological Techniques

John D. Bancroft und Marilyn Gamble

sir

i have followed this article for my special staining.For chrome alum preparaton i have used ready-made solution of haematoxyline  (harris) as one of my friend told it can be used  as it is 1% solution.other preparations are done as per the reference.my main problem is , during blueing or after phloxine treatment , tissue comes out from the glass slide. is there any mistake in slide preparation?. But for H& E i am getting results

wating for your response...

Rajesh R

thank you for your help and support ma'am....i have read bancroft but i didnt find much description abt chrome alum stain

Do I understand right, that you took ready-made Harris instead of 1% hematoxylin? Or did you take harris instead of the chrome-alum-hematoxylin-solution?

It is a huge difference to take pure hematoxylin-powder or to take ready-made hemalaun like harris for preparing the working-solution. Harris hematoxylin consists of additional reagens, that are likely to influence your staining. http://stainsfile.info/StainsFile/stain/hematoxylin/aluminum/harris.htm

And Harris hemalum has a low pH. If you mix Harris with the chrome alume recept you get something very different.

To save sections from coming off, you can use adhesive slides. But if your preparation of the working solution is wrong, it is possible, that the pH of your solution is very, very low. This enhances the danger of tissue-lost. Be sure, that you "bake" your slides for sufficient duration in the 60°C-ofen (30-60 min).

Dear Rajesh, I only can second the opinion and fact finding of Gudrun. I personally emphasize - if you are reproducing a method or technique first time -  to follow a given original recipe step by step, using all the solutions and chemicals given there. "Modifications" in/of solutions and chemical always may have some impact on outcome, yielding sometimes wrong / false results. Losing / loosing tissue from the section may have to do with the strong acidic pH of the Harris' Hematoxyline, as Gudrun pointed out, but may have also other causes: Unfortunately you didn't tell about the tissue you are staining (and more interestingly, how you processed that tissue before staining...i. e. fixation, processing into paraffin etc.). Good luck and best wishes, Wolfgang.

 Just as an information to readers:  S.Raja Sekhar has started also a separate thread on this matter: https://www.researchgate.net/post/Pancreatic_beta_cells_not_able_to_take_up_the_blue_color.