hybridization time and temperature

[xenia.ser...@gmail.com]

Hi, I am trying smFISH in C. elegans in a mixed population. The Stellaris protocol recommends hybridization at 37ºC for 4 to 16h. I wonder if it can be a problem if the hybridization is kept for longer than 16h (long O/N), and if the temperature (30ºC or 37ºC) can also influence the results.

Thank you!

[stevens...@gmail.com]

Have you tried the turboFISH protocol?

[Arjun Raj]

Hyb longer than overnight should still be okay. I think we've even done 36 hours and it was the same. Temperature does matter. We usually use 37C now.