{Reference Type}: Journal Article
{Title}: Targeted LC-MS quantification of intact TAT-fusion therapeutics: a case study.
{Author}: Zhao X;Sun J;Sun L;Chen J;Cheng Y;Duan X;
{Journal}: Bioanalysis
{Volume}: 7
{Issue}: 8
{Year}: 2015
{Factor}: 2.695
{DOI}: 10.4155/bio.15.17
{Abstract}: <B>BACKGROUND: </B>While HIV-1 TAT peptide-conjugation shows great promise on improving intracellular delivery of biotherapeutics in vitro and in vivo, quantification of TAT-fusion therapeutics in biological matrices represents a daunting challenge.<BR><B>METHODS: </B>A sensitive MS approach for accurate quantification of intact TAT-fusion protein/polypeptide in plasma was developed. i) A semi-automated 96-well ion-exchange solid phase extraction was developed; ii) a rapid LC separation on C4 was devised; iii) a TAT-fusion analog was constructed as internal standard.<BR><B>RESULTS: </B>We reported that low percentage of supercharging reagents enabled a significant sensitivity improvement of MS for intact TAT-fusion protein/polypeptide analysis. We showed a proof of concept by successfully developing a sensitive LC/MRM-MS method for quantifying GAP161, a TAT-conjugating RasGAP mimics, in rat plasma.<BR><B>CONCLUSIONS: </B>This work represents the first quantification of TAT-fusion therapeutics in biological samples by an LC-MS based method.