I am working on identification of a small peptide in brain tissue. We have modified the peptide with alkyne modifications for azide-based clicking. For Click reaction we have used 50 uM CuSO4, 300uM BTTAA, 1.75mM Na-Ascorbate and 5uM Biotin-azide/ 647-Fluorophore-azide/800-Fluorophore-azide. We have tried with different concentrations of azides as well. The problem we are facing is we are getting non-specific click labelling all throughout in all conditions when we are checking it on a gel (20% Tris-Tricine) or in a plate reader.
We have also tried with peptide purification followed by click reaction, that also is giving us non-specific labelling.
Any lead in troubleshooting or explanation would be really helpful.
Thanks
Try to use CuBr because the Cu(I) may have less interaction with peptides than Cu(II).
Good luck,
Laszlo
Hi Pushpa, i'm having the same problem. Did you find a solution? Thank you
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