Hello, I’m performing RIP experiment recently and I run into some issues.

Below is simplified version of my protocol

1. collect cell (I’m using 1/2 of a 6 well plate of 293T)

2. Pellet cells, add 1ml lysis buffer (with 0.5% NP-40)

3. incubate on ice for 5 mins

4. add antibody, rotate at 4C for 3hrs, then add beads, rotate for another 1hr.

5. wash, store sample for RNA (Trizol) and western-blot analysis.

The issue is, after 4C rotation, there’s white undissolved material present, and the buffer become so thick and slimy, can’t even been pipetted, or load into SDS-page gel.

I knew there’s protocol indicated that should use sonication for cell lysis, but my professor tells me not to since it’ll break RNA as well.

Is there any solution for this type of situations?