Hello,
I am currently setting up IP from cells overexpressing a protein tagged with HA. But I am not able to pull down my protein.
Could you tell me which HA antibody you are using for an IP? And what protocol do you follow? lysis buffer used? etc.
I use transfected primary neurons. (Transfection at 7 DIV, and cells lysis to do the IP at 14-15 DIV)
Many thanks in advance for your reply.
https://www.researchgate.net/post/Where_could_this_non-specific_pull_down_of_a_target_in_Co-IP_come_from
Hi Manon,
You can try different lysis buffers first to optimise your protein fractionation i.e. if your protein comes in a good amount in soluble form. You can start from basic RIPA buffer or simple Tris, NaCl.
For IP I have used HA antibodies from both Thermo scientific and CST and both works fine. I have used Protein AG beads from Thermo for antibody conjugation for IP.
You can find the protocol in the publication below:
Article Characterization of Plasmodium falciparum NEDD8 and identifi...
Hi Divya Das ,
Many thanks for your answer, and this information! This is very helpul.
Could you please give me the reference number of these HA antibodies?
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