Home >
Community >
Enrichment of AHA-tagged proteins: Should I use DBCO- or...
Upvote
21
Downvote
+ Click chemistry
+ Proteomics
+ Chemistry
Posted by
Lore Murdock
Enrichment of AHA-tagged proteins: Should I use DBCO- or...
Dear colleagues,I would like to seek your kind advice on click chemistry enrichment of azidohomoalanine (AGA)-tagged proteins.From literature, I learned that there are 2 main avenues, either (1) copper-free, DBCO-agarose bead pull down method or (2) Click-it enrichment kit (e.g. Thermo C10416) based on copper catalysis and alkyne-beads covalent capture.Ref 1 (DBCO): https://www.ncbi.nlm.nih.gov/pubmed/28104718Ref 2 (Click-it): https://www.ncbi.nlm.nih.gov/pubmed/28928394From your experience, which method would provide the greater enrichment efficiency without intensive optimization?As a newbie, it may feel safer to consider using an enrichment kit (e.g. the Thermo Click-it kit), but the manufacturer's protocol recommends an input of 5–20 mg proteins. Any experience on scaling it down (e.g. using 100-200 ug protein input)?Any feedback would be much appreciated.Thank you,Kay
