Bolton-Hunter: anyone still use this?

[johnfravolda]

I am curious about isotopic methods for labeling proteins.

What I am looking for is something robust, well established and non-specific (labels all proteins/protein fragments more or less equally).

I see products for iodination using Bolton-Hunter reagents and believe this is what I am looking for.

As you can tell from my question, I have very little experience working with proteins (most of my lab time was spent in the DNA world).

Is there anyone reading this forum willing and able to share their experience with Bolton-Hunter labeling ?

I understand that radiolabeling itself is "out" of many labs - but perhaps some still practice the old arts?

[mdfenko]

this is what we used to use pierce iodination beads.

simple to use. gentle with the proteins. good results.

[relaxin]

Why do you like to use radioactive iodine to label protein while many labs have abandoned it? You will need authorization, training to handle radioactive material, and expensive disposal of radioactive waste.

There are many nonradioactive methods of labeling proteins. For example, fluorescent label or Biotin-label can be done easily.

[johnfravolda]

Thanks for your answers mdfenko and relaxin.

Regarding the question of why radioactive, the properties we are looking for are strong signal, small molecule (minimum "interference") as well as fewest possible steps in the procedure. The method is conceived for a lab that already has the necessary infrastructure for working with isotopes.

None the less, I am interested in understanding how non-radioactive systems might work.

Biotinylation of the protein mix followed by SA-HRP would be a possibility - and could be tested as an alternative.

Any other suggestions for non-radioactive methods?

[relaxin]

You may try fluorescent dye labelling. A number of these dyes are available.

http://www.piercenet.com/browse.cfm?fld ... 8931CF7E3B