Blocking with Milk

U12201 at uicvm.uic.edu U12201 at uicvm.uic.edu
Fri Sep 22 13:10:04 EST 1995


In article <43t31n$klj at nnrp.ucs.ubc.ca>, hoekstra at unixg.ubc.ca (Sucking Gippy) says:
>
>Just a general question with regards to blocking membranes before
>probing. I generally use a non-fat milk solution in TTBS.
>Question: What are ideal dilutions to use? i.e. I've tried everything from
>          2%--> 10% Comments?
>
>          What is an ideal time period? Most "gaps" on the membrane (usually
>          nitrocellulose) should be blocked immediately upon addition of
>        blocking agent...it would be those so called "deep down pores/holes"
>         where a time period for blocking would be effective. I guess the
>         membrane pore size would also have an effect on the time period
>         required..right? Generally I use 1 hour at room temp with the 2% milk
>
Remember milk has biotin, so if you use an avidin/biotin
system stay clear of milk - it will cost you a lot of
avidin-enzyme conjugate to "titrate" out the biotin.
Each system has different optimal blocking reagent.
Pierce sells a "Blocker sampler kit" that may help
you select the right one.
Keld.



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