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A Sandwich ELISA for Measuring Benzo[a]pyrene-albumin Adducts in Human Plasma.


Anal Biochem. 2013 Jan 15;


Authors: Kei Chung M, Giovanni Regazzoni L, McClean M, Herrick R, Rappaport SM


Abstract

Exposures to polycyclic aromatic hydrocarbons (PAHs) have often been quantified via DNA or human-serum albumin (HSA) adducts of the carcinogenic metabolite, benzo[a]pyrene diolepoxide (BPDE). We previously reported a sandwich ELISA, using 8E11 as capture antibody and anti-HSA as detection antibody, that detected intact BPDE adducts in HSA isolated from plasma. After confirming that BPDE binds to HSA at His146 and Lys195, we modified the ELISA to measure intact BPDE-HSA directly in human plasma. To adjust for interferences due to nonspecifically bound HSA on well surfaces and to cross reactivity of the antibodies, the ELISA employs paired wells with and without addition of BPDE tetrols to deactivate 8E11. By performing assays in quadruplicate, a series of sample-specific adjustments and screening steps are used to reduce measurement errors that are a consequence of detecting low BPDE-HSA concentrations in the general population. ELISA measurements of BPDE-HSA in plasma from smoking and nonsmoking subjects (range: 0.335 - 0.941 ng BPDE-HSA/mg HSA; 4.59 - 47.2 fmol BPDE-HSA/mg HSA) and from highway workers with and without exposures to asphalt emissions (range: 0.346 - 13.8 ng BPDE-HSA/mg HSA; 5.68 - 228 fmol BPDE-HSA/mg HSA) detected differences in BPDE-HSA levels in the a priori- expected directions.

PMID: 23333225 [PubMed - as supplied by publisher]