What we are looking to do is to activate our surface with EDC/NHS chemistry, and then dry the surface for future binding of target proteins to the surface. Is this possible? Or, does the surface need to remain in an aqueous environment through EDC/NHS activation to protein conjugation.
Hiii you can activate your compound with EDC/NHS chemistry and then dialyzed in PBS or any other buffer to remove the unconjugated EDC/NHS and lyophilize for storage before conjugation with protein..but before conjugation with the protein re-suspend the lyohilized activated compound in the buffer in which you want the conjugation reaction with the protein and dialyzed the activated compound in the same buffer to remove the salts before conjugation if you had used different buffer for lyophilization of your activated compound for storage...if you had any query you can ask me any time..
Hi Francie, previously, i also activated my cooh-mwcnt electrode with edc/nhs to immobilize wth aminated-ssDNA. What i did was i immersed my mwcnt electrodes in the solution containing edc/nhs and PBS. Then i kept them in fridge at 4 degree C for about 2-5 h. Before i immobilized with aminated-ssDNA, i wash the activated electrode first, with several drops of PBS to removed unbound edc/nhs. then ready to immobilized. I also keep them in fridge after that for few hours before further characterize.
if u want to keep the activated electrode for few days before immobilize with ur target protein, i suggest u to wash the activated electrode with PBS , put them in a clean container like small petridish, wrap them well, and keep them in fridge until u r ready to immobilize. But not for so long, a few days might be still ok.
NHS can form relatively stable esters that can be stored away. EDC, on the other hand, degrades quickly in the presence of water (and worse in acid). If there is an amine in the water, the degradation of EDC to EDC-urea drives the amidation (a good thing)! Without amine present in water, the EDC will begin to degrade, resulting in reduced derivatization.
EDC can also form inactive N-acylurea by-products, in the presence or absence of water (even in hydrophobic organic solvents). These N-acylureas are resistant to aminolysis (not good).
If you are careful with things, keeping water away until you need it, and don't wash the surface with anything reactive to the EDC and NHS activated molecules, I think you should be fine.
All the best!
Hi to all,
I have functionalized my carboxylic nanoparticles (previously activated with EDC/NHS) with PEI (polyethyleneimine). I just want to know if it is possible to remove excessive EDC/NHS with multiple washing and centrifuging or we should dialyze it.
Thanks
Dialysis is the common method! All of nanoparticles will be remain by dialysis!