3 replies to this topic

[Claudia67]

Hi everyone,

Do you know if I can trypsinized the cells after fixation in order to obtain RNAm for RTPCR?

I really appreciate your help.

Thanks in advance.

[Mrtn]

Claudia67, on Mar 11 2009, 02:08 AM, said:

Do you know if I can trypsinized the cells after fixation in order to obtain RNAm for RTPCR?

Is RNAm a different notation for mRNA (messenger) ?
Why do you fixate cells before extracting (total) RNA?
I've always isolated total RNA from cells with a MN nucleospin column and then used a one- or two step RT kit with oligo dT primers for q-RT PCR.

[bob1]

Fixing crosslinks the DNA and RNA so that it will be very hard to extract.

[DavidJ]

It kind of depends on the fixation. I believe you can do it from (snap) frozen cells. Can you try lysing the cell directly. I also do that for non fixed isolations.