I have biotinylated my antibody and mixed with the magnetic beads. But after mixing I have blocked the conjugated magnetic beads with 1% BSA . I was seeing aggregation of magnetic beads even before and after the blocking .
Before answering the question it is necessary to know which kind of magnetic beads you used:
- What kind of reactive groups do they carry?
- Did you couple proteins to the activated surface before or should the biotinylated antibody be coupled?
- How did you 'neutralize' remaining activated groups?
Peter
How exactly did you biotinylate your antibody? I've used Lighting Link kit to biotinylate my antibody and found that the product caused my streptavidin magnetic beads to aggregate.I suspect the kit reactants and side products in the product mixture were to blame; there was no clean up step, as the manufacturer states the product is ready for use without cleanup.
Or...maybe you have too many biotins on your antibody and this is causing it to cross link your particles?
Hi Alamgir, a fact that is often unknown or underappreciated by many researchers is that serum albumin (BSA) has a free sulfhydryl.
These sulfhydryls can combine with any reactive groups on your magnetic beads, depending on their surface chemistry.
Thank you for answering . I have reduced the amount of Biotin and it improves the beads aggregation.
- Badges
- Science topic
- Similar topics
- Biological Science
- Immunology
- Antibodies