Recombinant protein - (May/16/2007 )

Dear ALL,

I got a recombinant protein which is from insect cells and purified to single protein band. I was told it can be used for Western. My question is: Is this protein a antibody? Why call it a recombinant protein? What is the difference between the recombinant protein and polyclonal antibody? Thanks!

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Well, I'm confused.

I'll assume you mean that the recombinant protein can be used to detect other things in a western, not that it itself can be detected (as all proteins can.) Is it an insect protein (unlikely) or was it cloned in for expression? A human or other mammalian antibody will dissociate into two bands (light and heavy chains) in a reducing gel (if loading dye had BME). A single band could be a heavy chain or a single-chain Fv (variable fragment, not used in westerns so much, since it has no constant region). It's recombinant because it was cloned (into baculovirus, I assume) and may have had fun bits tacked on to it.

A recombinant antibody will be monoclonal.

Where is the protein from and what is it supposed to detect?

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I think a Far-Western was suggested; instead of an antibody, you "overlay" a Westernblot with your purified protein; you may find protein-protein interactions;

the big problem is to monitor these protein-protein interactions; your purified protein should be labelled f.i. biotin, fluorescence tag. radioactivity a.s.o.

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I had wondered if it were something like that too. Perhaps if there is a lot of protein, there is enough to be labelled without too much loss.

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recombinant protein is called for every type of protein which is produced either artificially in unnatural host (ex insect cells for mammalian protein) or a modified protein (flag mammalian protein in mammalian cells).

I think that if you have enough protein you may use it in a polyacrilamide gel to purify it and immunize a rabbit to get antibodies, in supplement with other ideas previously mentionned, which i agree too.

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