RE: AAA-GLP

Sheila Magil (smagil@bioninc.com)
Fri, 28 Aug 1998 09:12:14 -0400

Nadine -
At my previous job we used the HP1090 with their on-line OPA-Fmoc
derivitization for AAA with great results. While they no longer sell
this option preloaded (it is just method software and loading the
chemicals which are available). However, I agree that hydrolysis is
crucial. I was using vapor phase hydrolysis in a Teflon vessel in a
dessicator with KOH trap in an oven. I used both 110*C for 16-24 hours
or at 165*C for 1 hour. The high temp method was particularly good for
quantitation of a protein of known purity and sequence without having to
extract it from a polymer matrix.
Sheila Magil
BION, Inc.
840 Memorial Drive
Cambridge, MA 02139
617-873-0963
Fax: 617-722-9772
Email: smagil@bioninc.com

-----Original Message-----
From: Ritter,Nadine
[mailto:Nadine.Ritter@add.ssw.abbott.com]
Sent: Thursday, August 27, 1998 3:42 PM
To: Recipients of ABRF List
Subject: AAA-GLP

What superb ideas from everyone! Thanks to all who have
given us suggestions
for improvements in our hydrolysis woes, and maybe even
saved us some big
bucks to boot.

The question I now have is this: If the Pico-Tag
workstation is no longer
going to be available, what alternatives are there for
convenient hydrolysis
methods, and how do they compare to the PicoTag system?
I recall some
discussion a long while back on microwave units, but
what about other methods
of hydrolysis? In reviewing the past ABRF AAA studies,
two addressed
hydrolysis: in 1989, the ABRF study showed that
hydrolysis was a highly
variable step in amino acid analysis, but it did not
focus on the relative
merits of different hydrolysis methods. The 1994 study
confirmed that good
hydrolysis is critical in obtaining accurate AA results,
regardless of the
derivatization method or instrument used. Of course,
hydrolysis has been an
internal part of the AAA Research Committee studies all
along, but as far as
I can tell, it has never been singled out for
comparative evaluation.

Might we consider a study on different hydrolysis
systems? Is this even a
do-able study? Do other ABRF'rs have an interest in this
issue? Or is
everyone happy with their current system, with no need
to replace the
instrument or method they use? How many out there do
high-throughput AAA
where instrument viability, performance degradation, and
future availability
of new equipment are causes for concern?

My overriding query: In the shrinking world of AAA
technology, as is evident
by the loss of instrumentation options for this
analytical method and the
lack of new technical developments, is it time to
re-evaluate what options
are left to those of us who continue to do AAA? I know
it isn't the most
glamorous analytical method anymore, but it is quite an
important quality
assurance test for biological products.

I'd like to hear your views (and I know I will!).

Thanks

Nadine Ritter
Abbott Diagnostics Division
Nadine.Ritter@add.ssw.abbott.com