Interfacial binding of human gastric lipase to lipid monolayers, measured with an ELISA

Biochemistry
M AoubalaR Verger

Abstract

Two sandwich enzyme linked immunosorbent assays (ELISA) were developed for evaluating the surface excess at the lipid/water interface of the human gastric lipase (HGL) and two anti-HGL monoclonal antibodies (mAbs). These assays were adapted to the monomolecular film technique used previously for measuring lipase kinetics. HGL and the two anti-HGL mAbs (4-3 and 218-13) were biotinylated without any significant loss of their biological activities occurring. They were further detected by ELISA using either anti-HGL or anti-mouse IgG polyclonal antibodies as specific captors before being revealed using a streptavidin--peroxidase conjugate as tracer. The detection limit was 25 and 85 pg in the case of HGL and mAb, respectively. By combining the above sandwich ELISA technique with the monomolecular film technique, it was possible for the first time to measure the enzymatic activity of HGL on 1,2-didecanoyl-sn-glycerol (dicaprin) monolayers as well as to determine the corresponding interfacial excess of the enzyme. The HGL turnover number increased steadily with the lipid packing. The specific activities determined on dicaprin films spread at 35 mN.m-1 were found to be in the range of the values measured under optimal bulk assay condi...Continue Reading

Citations

Aug 31, 2013·Analytical and Bioanalytical Chemistry·Susann BarigK Peter Stahmann
Dec 16, 1998·Chemistry and Physics of Lipids·J A FlipsenH M Verheij
Aug 28, 2003·Comparative Biochemistry and Physiology. Part B, Biochemistry & Molecular Biology·N MiledS Canaan
Mar 1, 1996·The Proceedings of the Nutrition Society·R VergerA De Caro
Jan 17, 2002·Bioscience, Biotechnology, and Biochemistry·A KobayashiF Mizutani
Sep 29, 2005·Analytical Biochemistry·W E MomsenH L Brockman
Jul 11, 2013·Colloids and Surfaces. B, Biointerfaces·Anaïs BénaroucheJean-François Cavalier
Jul 19, 2002·Biophysical Journal·Fu WangSen-fang Sui
Feb 19, 1998·The American Journal of Gastroenterology·M WøjdemannO Olsen
Sep 20, 2000·Accounts of Chemical Research·J F CavalierR Verger

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