PepSyn:Problems labeling with Fluorescein

lisa bibbs (bibbs@scripps.edu)
Tue, 14 Sep 1999 14:25:19 -0700

we have been trying to label the following peptide on the n-term with
Fluorescein.

Y G R K K R R Q R R R G K P P A P P M R N T S T M

Before labeling, we did a partial cleavage to make sure we made the intact
peptide and it looks great.

We tried using FITC followed by washing and cleaving. We end up with ONLY
the peptide - Y. I figure I must be going through an edman type cleavage
during cleavage and deprotection even though we keep it pretty cool. (I
know some are saying DAH!)

We then used Fluorescein-NHS. Most of our product is unlabeled (90%) and
then there is a product that is that is -17 amu.

What is your favorite protocol for labeling with Fluorescein?