Diagnosis and Dosimetry of Exposure to Sulfur Mustard: Development of Standard Operating Procedures: Further Exploratory Research on Protein Adducts.
Abstract
In continuation of previous grants, a standard operating procedure is being developed for two methods for retrospective detection of exposure to sulfur mustard: an immunoslotblot assay and/or ELISA for DNA adducts in blood and skin and the modified Edman procedure for determination of adducts to the N-terminal valine in hemoglobin. In the first year of the new grant period, both procedures could already substantially be shortened, while their sensitivities were improved. Furthermore, exploratory research is performed aiming at the development of a fieldable immunochemical assay for sulfur mustard adducts with three proteins, i.e., hemoglobin, albumin, and keratin. A reliable synthetic route was developed for 14Csulfur mustard used in various experiments. In vitro exposure of human blood to 1 muM sulfur mustard was determined by LC-MS-MS analysis of the tryptic T5 peptide from albumin alkylated at cysteine-34. Treatment with base released 80% of the bound radioactivity as 14Cthiodiglycol from keratin isolated from 14Csulfur mustard exposed human callus, which suggests that most of the adducts formed are esters of glutamic and aspartic acid residues. This opens the way for mass spectrometric detection of sulfur mustard exposure of skin by GC-MS analysis of thiodiglycol after hydrolysis of the esters and derivatization. Three partial sequences of hemoglobin, the T5 fragment from albumin, and three partial sequences of keratin were synthesized as haptens for raising antibodies, which contain adducted histidine, cysteine, and glutamine or asparagine, respectively. Several clones have been obtained using the three peptides containing adducted histidine as haptens, in addition to the clone 3H6 raised against a partial hemoglobin sequence containing adducted cysteine. Antibodies of these clones show specificity not only for sulfur mustard treated hemoglobin but also for treated keratin.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jan 01, 1998
- Accession Number
- ADA351437
Entities
People
- Daan Noort
- Govert P. Van Der Schans
- Hendrik P. Bensehop
- Leo P. De Jong
Organizations
- Prins Maurits Laboratorium TNO