In article <43733e0a.0306182130.73883db3 at posting.google.com>, immunoq at sbcglobal.net (Kiley) wrote:
>I need to radioiodinate a protein to use in immunoprecipitation assays
>for screening hybridomas producing MAbs specific to this protein of
>interest. I will be purchasing recombinant, carrier-free protein for
>this purpose and intend to have it commercially labeled.
>>I have never worked with radioiodinated proteins and therefore have
>some basic questions, such as: (i) will the structure and/or stability
>of the protein be affected by this process (and if so to what
>degree...what properties of the protein could predispose to structural
>changes/instability following radioiodination)?;
You cannot know it a priori. It depends on your iodination method
(Bolton-Hunter reagent coupling or direct iodination of tyrosines) and
properties of your protein. Generally however iodination is mild
enough to keep most of any protein active.
>(ii) given the
>half-life of I-125 (60 days?), at what point in time following
>labeling should I expect to see a decline in radioactivity to the
>point that sensitivity for the purposes of immunoprecipitation would
>be "lost"?
Assuming radiolysis is not a problem (also impossible to know
before hand), it will decay 8X in a year. Thus, it depends on
the original specific radioactivity of your protein and whether
you can afford exposing for ~ 10X longer.
DK
