FW: FITC peptides

Everson, Anita [PRI] (AEverson@prius.jnj.com)
Tue, 10 Aug 1999 16:31:19 -0400

We coupled numerous peptides with FITC (as the isothiocyanate)and not had
any trouble with them using 0.1% TFA. Coupling with FITC takes place
post-synthesis because it is soluble in water. Often, however, the
fluorescein group is hard to couple onto the peptide and it is not unusual
to see peptide with and without the fluorescein group. We've also tried
using the Sigma fluorescein (as well as fluorescein from other vendors) to
couple before cleavage and they all seem to have a great deal of lot-to-lot
variability in appearance and solubility, and also success in coupling.
Currently we use carboxyfluorescein to couple on the resin (cleave with HF)
and we've not had any trouble, other than usual steric hindrance in
coupling.

Anita Everson
Peptide Lab
RW Johnson PRI, La Jolla

> -----Original Message-----
> From: Roger Murphy [SMTP:Roger.Murphy@ludwig.edu.au]
> Sent: Monday, August 09, 1999 9:33 PM
> To: Recipients of ABRF List
> Subject:
>
> Hi folks!
>
> Any comments from anyone on the stability of an FITC-peptide conjugate at
> low pH (say ~ 2)? I've labelled a decapeptide at the N-terminus with FITC
> (i.e. as the isothiocyanate) but it looks as if it's fallen apart on
> purification under standard RP-HPLC conditions (i.e. 0.1% TFA). We're
> going to look at HPLC at pH 6.5 and at using succinimide ester derivatives
> as well, but just wondered if anybody else has had a similar experience
> with FITC and peptides.
>
> Cheers,
>
> Roger
>
> ------------------------------------------------------------
> Roger Murphy, Ph.D.
> Biological Production Facility
> Ludwig Institute for Cancer Research
> Austin & Repatriation Medical Centre
> Studley Road,
> Heidelberg, Vic. 3084
> Australia.
>
> Tel 61-3-94965463
> Fax 61-3-94965436
> Email Roger.Murphy@Ludwig.edu.au