Hello all,
We are using EDC/NHS coupling to bind proteins to carboxylated quantum dots. However, after the coupling reaction we observe QDs aggregates that cannot be solubilized again, and therefore do not allow to continue further experiments. Anybody has experienced a similar protein and can give any advice? Do you think that we can solve the problem just by playing with concentrations or times for coupling? Or do you recommend trying a different strategy, such as biotin protein-streptavidin QD?
Thanks for your help!
Hi Laura, are you doing these reactions with QDs separate or all in one reaction?
The problem with using EDC is that it crosslinks all -NH2 to all -COOH creating a insoluble mess that cant be solubilized because they are crosslinked by covalent amide bonds.
My recommendation is to forget about the NHS coupling. Incubate the QDs with EDC. Use spin columns to separate the QDs from EDC and then incubate them with excess protein.
I do not know which kinds of carbxylic QD did you used for bioconjugation. Did you used commercial carboxylic QD or made by yourself. MPA capped QD might have some problems(insoluble problems) due to instability of MAP capped in low pH value (pH
Thanks for your answers.
We are using commercial carboxylated QDs. We have tried to do the coupling in 2 steps without using NHS, apparently worked better, although we still have to confirm the binding.
Thanks a lot,
Laura
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