Hi!
We are trying to use surface functioned microbead for protein conjugation. One major way is to use NHS functioned-bead. The manufactor also recommend the epoxy functioned bead. Both functionalisation can directly immobolise protein covalently. (aren't they?) Which conjugation reaction is more efficient?
Thanks!
It depends on your target protein. NHS will attach the protein via amine groups ( amino acid Lysine in the protein). This is a very efficient and targeted reaction and lysine itself offer a spacer between the microbead surface and the protein. Epoxy groups are usually used to link with hydroxyl groups on the protein (amino acid Tyrosine) though it can also be used to link thiol, and amino groups on the protein. So, the choice depends on which amino acid in your protein you want to use and what impact it will have on subsequent use of the microbeads
NHS ester groups are very reactive and also very unstable. Epoxy groups are very stable but less reactive. For batch consistency I prefer epoxy chemistry but the rate of reaction is likely to be slow unless you have quite high pH. This is not always possible to do with proteins.
NHS reaction will be easier via lysine but be careful how you store NHS activated materials (must be very dry).
However, if your protein has accessible free thiols then epoxy chemistry becomes more attractive.