in-vivo cross-linking?

Emir Khatipov khatipovNO at NOuchicago.edu
Thu Apr 11 15:03:55 EST 2002


What kind of approach exactly you are thinking about? Immunoprecipitation
might be a better way to study protein interaction. How you are going to
separate proteins that after crosslinking will form a net inside the cell?
Intracellular protein concentration of protein is >100mg/ml, and other
proteins will be crosslinked to your complex of interest.

I for your question, I would try paraformaldehyde or formaldehyde. These
chemicals are generally used to fix (in fact - crosslink "everything to
everything") yeast and mammalian cells. PFA (usually 1.5-4% solution in PBS,
sometimes supplemented with ~1% sucrose) works faster - within a few minutes
(<5min). Formaldehyde requires longer times of incubation (30min-overnight).
formaldehydes should work same well for bacteria, as for eukaryotes.
I guess detailed protocols are published for bacteria. However, I don't have
any reference.

Emir

PFA works faster "Yoram Gerchman" <gerchman at Princeton.EDU> wrote in message
news:3CB4606C.5D51 at mailserver1.Princeton.EDU...
>
> Greetings Netters
> Does any one know about a cross linker suitble for in-vivo cross linking
of
> soluble protein in bacteria? We want to look for complex formation.
> Thanks Yoram
>
>
> ---





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