Angela,
I have used a Waters Picotag system to quantify Glutamine in serum
using ice cold Methanol, added to give 80%(v/v) methanol, to precipitate
protein. The supernatant wasthen dried and used for analysis directly using
standard Picotag chemistry and an abbreviated gradient. Good luck.
Chris
"Angela c. Murphy" wrote:
> I have been asked if I can quantitate a single free amino acid from a
> yeast cell lysate. I think I can, with the help of my Beckman 6300,
> which uses sodium citrate buffers and post-column ninhydrin detection.
> I've asked that lipids be extracted with chloroform and that proteins also
> be removed. She's going to try to use ultrafiltration with a low
> molecular weight cutoff membrane, about 1,000. What else needs to be
> done so that free amino acids are just about the only things left and
> things that could mess up my column are removed? What concentrations of
> free amino acids are usually seen in yeast cell lysates - nanomoles/ml,
> micromoles/ml, millimoles/ml? Excuse my ignorance - I usually work with
> peptide and protein hydrolysates, this is my first time with a cell
> lysate.
> Thanks for any help.
> Angela C. Murphy
>
> *******************************************
> Angela C. Murphy, Chemist
> Lab. of Cell Biology, NHLBI, NIH
> 3 Center Drive, MSC 0301, Rm. B1-22
> 9000 Rockville Pike
> Bethesda, MD 20892-0301 USA
> tel.: (301) 496-2324
> fax: (301) 402-1519
> email: acmurphy@helix.nih.gov
> *******************************************
-- Chris Wood Senior Scientific Officer Department of Biochemistry University of Queensland Australia 4072 Phone 07 3365 2978
--------------2DFD8AF9515A7128F091FA56 Content-Type: text/x-vcard; charset=us-ascii; name="wood.vcf" Content-Transfer-Encoding: 7bit Content-Description: Card for Chris Wood Content-Disposition: attachment; filename="wood.vcf"
begin:vcard n:Wood;Chris tel;work:(07)3365 2978 x-mozilla-html:FALSE org:University of Queensland;Biochemistry version:2.1 email;internet:wood@florey.biosci.uq.edu.au title:Senior Scientific Officer adr;quoted-printable:;;Biochemistry Department =0D=0AUniversity of Queensland=0D=0A;Brisbane;Queensland;4072;Australia x-mozilla-cpt:;1 fn:Chris Wood end:vcard
--------------2DFD8AF9515A7128F091FA56--