2. What kind of acid stability do these labels have? We can use either Boc
(HF) or Fmoc (TFA) chemistry, but were successful with Boc chemistry before
with the unlabelled peptide. Any advice on that?
I am aware that similar questions have been asked before from the relatively
few experts in the list. I am sorry for the reiteration. Thanks in advance for
your advice. Best regards,
Dear David: Try the carboxyfluorescien-OSU esters (Molecular Probes) in
DMF + DIPEA. About a 3 X excess over amino groups, overnight. For good
measure, I keep the reaction under low or no light, and flushed with Ar if
possible. These conjugates are stable to TFA cleavage. I have only used
Fmoc protected AAs. I am sure FITC should work well, but I cannot remember
why I have never used it.
NBD should couple readily as the ITC derivative, and is expected to be acid
stable. However, I have never conjugated this beastie during synthesis. I
have used it in soultion to label proteins and it works well. I think as a
general rule, a good teritiary amine in DMF should work for all these types
of coupling.
In case the peptides are difficult to synthesize by Fmoc strategies, try
HATU/DIPEA activation, works like a charm almost everytime. Although I saw
a new reagent for achieving high coupling yields for very hindered
couplings. I do not have the reprint with me, but I think it was in
Tetrahedron Letters. In any case good luck, Gautam Sarath
Gautam Sarath
N-226, Beadle center
Protein Core Facility
Department of Biochemistry
University of Nebraska-Lincoln
Lincoln, NE 68588-0664
Phone: 402-472-2928
FAX: 402-472-7842