title: Immunological detection of a novel advanced glycation end-product.
abstract: BACKGROUND
  The advanced stage of the Maillard reaction that leads to the formation of advanced glycation end-products (AGEs) plays an important role in the pathogenesis of angiopathy in diabetic patients and in the aging process. Recently, it has been proposed that the intermediates contributing to AGE formation include dicarbonyl intermediates such as glyoxal, methylglyoxal, and 3-deoxyglucosone (3-DG). In the present study, we developed a novel, non-carboxymethyllysine (CML) anti-AGE antibody that recognizes serum proteins and peptides modified by 3-DG in vivo.
  
  
  METHODS
  AGE-modified serum albumins were prepared by incubation of rabbit serum albumin with 3-DG or D-glucose. After immunization of rabbits, anti-AGE antisera were subjected to affinity chromatography on a Sepharose 4B column coupled with CML-BSA, or AGE-BSA created by incubation with 3-DG (AGE-6) or D-glucose (AGE-1). The AGE-Ab-6 and AGE-Ab-1 thus obtained was used to investigate AGEs in serum from diabetic patients on hemodialysis.
  
  
  RESULTS
  Characterization of the novel AGE-Ab-6 obtained by immunoaffinity chromatography was performed with a competitive ELISA and immunoblot analysis. This antibody specifically cross-reacted with proteins modified by 3-DG. AGE-6 was detected in diabetic serum as three peaks with apparent molecular weights of 200, 1.15, and 0.85 kD, while AGE-1 was detected as four peaks with apparent molecular weights of 200, 65, 1.15, and 0.85 kD.
  
  
  CONCLUSIONS
  This study provides new data on the pathways of AGE formation from 3-DG and methods for the immunochemical detection of AGEs. We also provide immunochemical evidence for the existence of six distinct AGEs in vivo among the AGE-modified proteins and peptides in the serum of diabetic patients on hemodialysis.
authors.affiliation: Department of Biochemistry, Faculty of Pharmaceutical Science, Hokuriku University, Kanazawa, Japan. m-takeuchi@hokuriku-u.ac.jp
authors.forename: M
authors.initials: M
authors.lastname: Takeuchi
authors.forename: Y
authors.initials: Y
authors.lastname: Yanase
authors.forename: N
authors.initials: N
authors.lastname: Matsuura
authors.forename: S
authors.initials: S
authors.lastname: Yamagishi Si
authors.forename: Y
authors.initials: Y
authors.lastname: Kameda
authors.forename: R
authors.initials: R
authors.lastname: Bucala
authors.forename: Z
authors.initials: Z
authors.lastname: Makita
mesh_terms: D000818:Animals; D003847:Deoxyglucose / Q000031:analogs & derivatives* / Q000737:chemistry; D017127:Glycation End Products, Advanced / Q000032:analysis* / Q000737:chemistry / Q000276:immunology; D006801:Humans; D007118:Immunoassay; D011499:Protein Processing, Post-Translational; D011817:Rabbits
chemical_list: D017127:Glycation End Products, Advanced; D003847:Deoxyglucose; C016350:3-deoxyglucosone