Re: Antipeptide antibodies

Gottfried Proess (g.proess@eurogentec.com)
Tue, 03 Aug 1999 16:28:03 -0700

Dick Wicks wrote:
>
> Dear ABRFers:
>
> After seeing the post on the anti-peptide antibodies, I was
> wondering if anyone could give me feedback on our experience with
> making these. We made 12 mer peptides from 85 different proteins
> (selected for hydrophilicity, antigenicity, etc using DNASTAR
> program). Coupled to BSA via the terminal cysteine that we added
> using SMCC, then injected into rabbits. The resulting antibodies
> were purified on peptide columns. Only about 20% of the
> antibodies reacted with the respective proteins in Western blots
> under reducing conditions. Any idea whether this is unusual?
> Thanks very much.
> Dick Wicks
>
> ---------------------------------------------------------------
>
> Name: vcard.vcf
> Part 1.2 Type: text/x-vcard
> Encoding: 7bit
> Description: Card for Dick Wicks

Dear Dick,

it is quite unusual to my opinion. We get statistically around 70-75%
success rate for Western Blotting under denaturing conditions; our
peptides make generally 15-16 aa. This rate is based on about 300
feed-backs from some of our customers. I even remember even one customer
who asked us to synthesize 12 peptides from the same protein, all of the
sera worked nicely in Western.

Maybe we use other parameters to determine the choices than you do?

We choose the peptides with the same program, using the algorithms for
hydrophilicity (2), antigenic index, surface probability, flexibility,
secondary structure predictions and charge distribution. The comparison
of all these parameters determine our choices.

If you need further informations, contact me. I do not know if
additional discussions will be interesting for the whole list?
Best regards

-- 
Gottfried Proess
Tel.: + 32 4 366 01 50  Fax : + 32 4 365 16 04
E-Mail: g.proess@eurogentec.com
Web-Site : http://www.eurogentec.com  <==== visit it !