Peptide Solubility

gheavner at NETAXS.COM gheavner at NETAXS.COM
Sun Feb 12 19:05:44 EST 1995

You wrote:

Help! I am performing MHC/peptide binding assays by FACS but 
terrible problems with peptide insolubility following HPLC 
(elutes @ 30% acetonitrile) and freeze-drying (sequence 
 I have tried dissolving in PBS, DMSO,and acetonitrile. Initial 

dissolving is in
Guanadinium prior to HPLC, but this doesn't agree with cell 
culture (20h incubation
with EBVs). Any suggestions?


p.s. a similar peptide biotin-GLEPGVVAEKVRNLS is no bother at 
all. Ho hum.



We synthesize, purify and assay about 1000 peptides/year.  The 
solubility senario you describe is nothing unusual.  Peptides 
can be as insoluble in aqueous systems as any other organic 
compounds.  You might expect that the presence of charged 
residues would confer solubility, but this is not always the 
case,  Our experience is that solubility is as much a factor of 

sequence as composition.  Scrambling the same sequence to 
create a "control" peptide can give a compound of drastically 
different solubility.  Under such circumstances, you have to 
question if such scrambled sequences are good choices for 
negative controls.  The addition of prosthetic groups such as 
biotin, dansyl, etc. can also have significant but 
unpredictable effects on solubility.  We have used several 
approaches to dissolving sparingly soluble peptides.  One that 
you may try is to prepare a saturated solution of the peptide 
in 25-50% aqueous DMSO and cautiously dilute with your assay 
buffer.  You should also run a DMSO blank in your cell culture 
to determine compatability.  Dilute buffers such as 25 mM 
acetate or bicarbonate also can be used, with subsequent 
dilution with assay medium.  More concentrated buffers do not 
appear to work as well.

George A. Heavner
Senior Director, Peptide R&D
Centocor, Inc.

More information about the Immuno mailing list