We will trying ti isolate marker positive extracellualr vesicle(EV), from pre-enriched EV sample using antibody conjugated bead.
The used bead for antibody conjugation is aldehyde/sulfate or carboxylated bead.
We confirmed that EV attached to these bead thorough remaining EV amount in supernatant (using protein amount) and bead based ELISA.
Now the problem is how disassemble or detach these EV and antibody-conjugated bead complex.
The commercial product just lysis the EV in state of EV and bead complex or just used in FACS.
In researchgate question section, pH adjustment (low pH) could be used it, but it require additional wash step (maybe ultracentrifugation step).
Is there any method to disassemble the EV-antibody conjugated bead complex, without additional wash step? (except the bead elimination step, the low speed centrifugation is enough to do)
To Fish
Thank you for your kind answer.
The washing means, if we added the buffer for disassemble the antigen-antibody complex, we have to eliminate that buffer. To eliminate the buffer, it require additional ultracentrifugation step or dialysis, and these process maybe reduce the amount of the EV. That's why we require disassemble buffer that doesn't require additional washing step.
And thanks for advice. Maybe Mg++ ions could be a additional buffer. I would check it.
Hi William,
Thanks for the clarification. For elimination of dissociation buffer you can try one of the two:
1. High temperature (if your reagents can withstand 60C and above) - no buffer is involved.
Or:
2. Weak acid (e.g. acetic, citric) for dissociation then add concentrated TRIS Base to stop the reaction and create the buffer.
Good luck,
Falk
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