Volume 115, Issue 3, Pages 686-692 (September 1998) Detection of circulating antibodies to malondialdehyde-acetaldehyde adducts in ethanol- fed rats  Dongsheng Xu, Geoffrey M. Thiele, John L. Beckenhauer, Lynell W. Klassen, Michael F. Sorrell, Dean J. Tuma  Gastroenterology  Volume 115, Issue 3, Pages 686-692 (September 1998) DOI: 10.1016/S0016-5085(98)70148-9 Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 1 Structure of a major MAA-protein adduct. Gastroenterology 1998 115, 686-692DOI: (10.1016/S0016-5085(98)70148-9) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 2 Immunoreactivity of plasma from ethanol-fed and control rats to unmodified BSA and BSA-MAA as determined by Western blot analysis. Unmodified BSA (7.5 μg) and BSA-MAA (7.5 μg) were subjected to 10% SDS-PAGE, transferred to nitrocellulose membranes, and reacted with plasma (1/50 dilution) from ethanol-fed (E) and pair-fed controls (C) as described in Materials and Methods. Immunoblots of 2 representative pairs of animals of 12 pairs tested are shown. Gastroenterology 1998 115, 686-692DOI: (10.1016/S0016-5085(98)70148-9) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 3 Inhibition of immunoreactivity of plasma from ethanol-fed rats by hexyl-MAA. ELISA plates were coated with either (A) BSA-MAA or (B) HEL-MAA. Plasma from controls (○) or ethanol-fed rats (●) was preincubated with varying amounts of hexyl-MAA, and the percent inhibition of antibody binding was determined in a competitive ELISA as described in Materials and Methods. Gastroenterology 1998 115, 686-692DOI: (10.1016/S0016-5085(98)70148-9) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 4 Immunoreactivity of plasma from control and ethanol-fed rats with MAA-modified and unmodified rat liver cytosolic and microsomal proteins. ELISA plates were coated with either (A) microsomal or (B) cytosolic proteins with (MAA) or without (Native) MAA modification. Serial dilutions of rat plasma from control (□) or ethanol-fed (■) rats were added to the plates, and bound antibody was detected by ELISA as described in Materials and Methods. Results are expressed as nanograms per milliliter of Igs and represent means ± SEM for 12 animals. *P < 0.01, #P < 0.001 vs. controls. Gastroenterology 1998 115, 686-692DOI: (10.1016/S0016-5085(98)70148-9) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 5 Immunoreactivity of plasma from control and ethanol-fed rats with MAA-modified and unmodified rat liver plasma membrane. ELISA plates were coated with plasma membranes with (MAA) or without (Native) MAA modification. Serial dilutions of rat plasma from control (□) or ethanol-fed (■) rats were added to the plates, and bound antibody was detected by ELISA as described in Materials and Methods. For some experiments, plasma was preincubated with excess hexyl-MAA (20 nmol/well) before addition to the coated plates. Results are expressed as nanograms per milliliter of Igs and represent means ± SEM for 12 animals.*P < 0.01, #P < 0.001 vs. controls. Gastroenterology 1998 115, 686-692DOI: (10.1016/S0016-5085(98)70148-9) Copyright © 1998 American Gastroenterological Association Terms and Conditions