Antibody's specifity...

Mr. Hat misterhat at sbcglobal.net
Wed Jul 23 21:59:30 EST 2003


In article <bfnant$qf3$1 at reader1.panix.com>, iayork at panix.com (Ian A.
York) wrote:

> In article <3F1F1222.9000403 at NOSPAM.ucalgary.ca>,
> Bryan Heit  <bjheit at NOSPAM.ucalgary.ca> wrote:
> >Mike Clark wrote:
> >
> >>Antibodies may or may not be "specific" depending upon how they
> >>are selected and how in practice you define specificity. Specificity is not
> >>
> >But how many times have you run a cell lysate blot with an antibody and 
> >seen greater then 1 line?  It's rare, and when it does occur it's 
> 
> Many, many times. 

Yes (esp. with polyclonals).
 
> 
> >usually human error or a closely related protein.  In all of the blots 
> >and elisa's I've done (that's a lot) I've never once encountered an 
> >antibody with cross reactivity for a non-related protein, and that 
> >includes some "dirty" polyclonal serums I derived myself.
> 
> Many of the polyclonals I've raised, and many of those we've got from 
> various collaborators, cross-react.  Rabbits see *lots* of antigens even 
> in their constrained little lives in a lab cage, not just the ones you 
> inject.  Those may not be formally cross-reactive (i.e. the same clonal Ig 
> may not be reacting with two antigens) but the result is the same.

(Exactly...I should have read this response from you first).

> 
> I've also seen a few monoclonals that cross-react with unrealted proteins, 
> though not very many.
> 
> >Can you cite a single occurrence where antibodies show cross-specificity 
> >with a protein only 50% identical, unless it is recognizing a shared 
> >epitope?  I did a quick check of pubmed and could not (not to say there 
> >isn't, I didn't look too closely).
> 
> Most of the cross-reactive antibodies are simply thrown away, they're not 
> worked up to the point of eliminating shared epitopes (an artificial 
> constraint you've added to the original question).  By the time they're 
> published, you're generally only seeing the best (i.e. most specific) 
> antibodies.  Depending on the published literature for this question is 
> not particularly useful.

Absolutely, because most people do not have a use for these and therefore
they are disposed of. In the cases where a broadly cross-reactive
monoclonal is DESIRED however, like I said before (a previous post?) it's
all a matter of what you choose to pressure the selection in the course of
screening. I happen to be developing hybridomas of this exact sort at the
moment...it makes screening rather tedious, but you get what you seek.

> 
> >Or you could look at it as a group of antibodies all recognizing the 
> >same antigen.  Idiotype antibodies are a special case, as it is hard to 
> >identify what is the "antigen" and what is the "antibody".  As I said 
> >previously, I am unaware of any study showing cross reactivity between 
> >proteins with little/no homology.
> 
> I've certainly seen cross-reactive monoclonals.  Were they reacting with 
> shared epitopes, with closely-related proteins, with unrelated stuff?  I 
> have no idea.  I threw them away, because they were useless for my 
> purpose.  That's what happens to most of the cross-reactive antibodies.  
> 
> Ian

best,
M.H.



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