I have been trying to get insitu hybridization to work on stomach cryosections and it doesn't seem to work. I am detecting a cytokine using a riboprobe I have designed. Has anyone worked on it and can offer suggestions about any processing steps prior to doing insitu?
Do you have a positive control probe like B-actin, do you use the digoxigenin system ?
did you try paraffin sections in stead of cryo-sections?
Hi, Which kind of in situ hybridization are you use? For RNA or DNA detection?
Rene, my positive control is proton pump which is present in parietal cells in the stomach.And yes, it is DIG system! After a couple of tries, my positive control worked, but now even that has stopped working!
Babak, I am detecting mrna and my probe is a RNA probe!
Hallo again Poshmaal,
In fact I had the same problems.
we used an extensive protocol from Thesleff Finland, it worked great in the beginning, but after a while I didn't get any staining at all, even the B-actin (commercial probe from Roche) didn't do anything.
Because there were so many different steps and solutions we didn't know were to begin.
What solved the problem in the end was going back to a more basic ish protocol, used by the Pathological Departement.
If you are interested I can send you the protocol.
I allso read some good suggestions in the forum concerning ish!
Good luck and greetings
Rene, if you could send me the protocol, it would be great! Thank you so much!
My email is: [email protected]
Rene, could you send me the protocol too? I also met some difficulities in ISH. Thank you so much. My e-mail: [email protected]
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