Protein free blocking agent

Stephen Dahl stayve-and-irayne at worldnet.att.net
Thu Nov 11 14:46:05 EST 1999


Fred <dynamin at hotmail.com> wrote in message
news:382A3EC8.9A7FED38 at hotmail.com...
> Funny.  We use NC and PVDF both.  Exclusively use chemi.  Never really
> noted a problem, having used it for about 6 yrs.  Still using it now.
> No other changes to standard blot protocols.  An advantage of the PVP40
> is you can amido black (or coomassie) stain after chemi, rather than
> before.  We find this very useful (still works after milk blocking, but
> not so well).
> whats' "A/B"?  We switched to PVP40 from skim milk.

Sorry, a little jargon there.  A/B is simply comparing something side by
side.  When I did my test I ran a mini "curtain" gel (More jargon,!  Put the
comb in teeth up--not too deep--and after polymerization you have one well
that extends the width of the gel)  I ran a cell lysate, blotted and then
sliced strips with a razor blade.  This yields identical blots for
comparison.  Then you can either use the plastic bubble package from Qiagen
mini columns or the cases from coverslip boxes or anything you've got that
works with small strips individually to compare blocking agents.  I checked
out milk, gelatin, BSA, PVP40  and got best results with milk.  You've
piqued my curiosity though so maybe I'll try it again with a different
primary antibody.

Regards,
Steve Dahl
JHMI
stebby at jhmi.edu






More information about the Methods mailing list