Retrospective detection of exposure to sulfur mustard: Improvements on an assay for liquid chromatography-tandem mass spectrometry analysis of albumin/sulfur mustard adducts

We here report on the further development of the method comprising the pronase digestion of albumin alkylated by sulfur mustard and the subsequent mass spectrometric analysis of an adducted tripeptide. This includes significant improvements in both the albumin isolation procedure and the automation of the microliquid chromatography-electrospray...

Comparison of immunoaffinity chromatography enrichment and nuclease P1 procedures for 32P-postlabelling analysis of PAH- DNA adducts

32P-postlabelling analysis for detecting DNA adducts formed by polycyclic aromatic compounds is one of the most widely used techniques for assessing genotoxicity associated with these compounds. In cases where the formation of adducts is extremely low, a crucial step in the analysis is an enrichment procedure for adducts prior to the...

Enzyme kinetics and substrate selectivities of rat glutathione S-transferase isoenzymes towards a series of new 2-substituted 1-chloro-4-nitrobenzenes

1. Four different rat glutathione S-transferase (GST) isoenzymes, belonging to three different classes, were examined for their GSH conjugating capacity towards 11 2-substituted 1-chloro-4-nitrobenzene derivatives. Significant differences were found in their enzyme kinetic parameters K(m), k(cat) and k(cat)/K(m). 2. Substrates with bulky...

Purification and characterization of eight glutathione S-transferase isoenzymes of hamster: comparison of subunit composition of enzymes from liver, kidney, testis, pancreas and trachea

Eight dimeric isoenzymes of glutathione S-transferase (GST) were purified from liver, kidney and testis of the Syrian golden hamster, using S-hexylglutathione affinity chromatography and chromatofocusing. The isoenzymes were characterized according to their substrate selectivity, physical properties and amino acid sequence analysis. Thus a...

One-step purification of tissue-type plasminogen activator using affintiy chromatography with a special monoclonal antibody under mild conditions

We have previously isolated a monoclonal antibody, designated as 1-3-1, specific for tissue-type plasminogen activator (t-PA). We have shown that t-PA dissociates from 1-3-1 in the presence of the lysine analogue 6-aminohexanoic acid (6-AHA). Here we describe a method for the one-step immunoaffinity purification of t-PA from conditioned melanoma...

A rapid and simple method for the separation of four molecular forms of human plasminogen

At least four molecular forms of plasminogen are known. Two of those forms have glutamic acid at their amino-terminal end, and are designated as glu-plasminogen. The other two have lysine, methionine and/or valine as amino-terminal amino acid and are collectively designated as lys-plasminogen. Two subforms (I and II) each of glu- and lys...

Evidence for the occurrence of a fast-acting inhibitor for tissue-type plasminogen activator in human plasma

Human plasma contains a fast-acting t-PA inhibitor, which is not identical with ??2-antiplasmin or ??2-macroglobulin. The concentration of this inhibitor in normal plasma is highly variable, much lower than that of known plasma protease inhibitors, and in the range of physiologically occurring plasma concentrations of t-PA (0-2 IU/ml). The...