Good morning ,
I try to do formaldehyde cross link between purified tagged protein from E.coli and total extract of bacteria : sinorhizobium melliloti to identify interactants proteins of my purified tagged protein , the goal is to see by western blot , a complex of interactants proteins. I'm looking for a protocol to do it.I tried one protocol with different concentrations of formaldehyde and i have problem , a white precipitate appear ; i didn't know wh??
You need to optimize the conditions of your reaction medium/buffer. pH, ionic strength and the use of additives are very critical factors to maintain your desired products in their soluble and stable form in solutions. You can use a high-throughput screening platform format for identification of reaction media/ buffer conditions.
Hi Alemu Tekewe,
thank you for your answer , i would like to have a protocol to try again to do it if you have something to suggest me .
Best Regards
Saida
Here is an example of Formaldehyde cross-linking optimization (among many) with protocols J. Biomed. Biotech. 2010, 927585. doi:10.1155/2010/927585.
Maybe the white precipitate comes form formaldehyde polymerization into paraformaldehyde? This can be avoided by decreasing formaldehyde concentration, preventing storage in cold, or using stabilizers like methanol.
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