Development of a monoclonal antibody-based ELISA system for glyceraldehyde-derived advanced glycation end products.
We have previously found that glyceraldehyde-derived advanced glycation end products (glycer-AGEs) elicit oxidative stress generation and evoke inflammatory and thrombotic reactions through their higher binding affinity to RAGE (receptor for AGEs), thereby playing a role in vascular complications in diabetes. Furthermore, circulating levels of glycer-AGEs are elevated in diabetes. We characterized a monoclonal antibody (mAb) raised against glycer-AGEs and prepared its specific ELISA system in human serum. We developed here mAb reacted specifically with glycer-AGEs or glyceraldehyde-derived pyridinium, but not other structurally identified AGEs or AGE precursors. The mAb not only completely neutralized the deleterious effects of glycer-AGEs on endothelial cells, but also detected glycer-AGEs in the aorta of type 2 diabetic rats. Intra and inter-assay coefficient variations of the ELISA were 6 and 2.6%, respectively. ELISA linearity was shown intact within 5-fold dilution, and recovery ratio of added glycer-AGEs was 88-117%. Results of serum and plasma were comparable, and repeated freeze-thawing of samples did not affect the results (90.1-112.4%). Serum glycer-AGEs levels in 30 healthy subjects evaluated by the ELISA were strongly correlated with those by polyclonal Ab-based one (r=0.82). Our present study suggests the clinical utility of mAb for evaluating glycer-AGE levels in both tissue and serum.
['Matsui|Takanori|T|', 'Joo|Hoo Don|HD|', 'Lee|Jae Min|JM|', 'Ju|Sung Mi|SM|', 'Tao|Wang Hong|WH|', 'Higashimoto|Yuichiro|Y|', 'Fukami|Kei|K|', 'Yamagishi|Sho-ichi|S|']
D000368:Aged D000911:Antibodies, Monoclonal D002460:Cell Line D042783:Endothelial Cells D004797:Enzyme-Linked Immunosorbent Assay D005260:Female D017127:Glycation End Products, Advanced D005985:Glyceraldehyde D064368:Healthy Volunteers D006801:Humans D008297:Male D008875:Middle Aged
ELISA AGEs Diabetes Monoclonal antibody Endothelial cells