Background: Advanced glycation end-products (AGEs) are formed during cascade reactions between reducing sugars or reactive aldehydes and proteins, lipids or DNA molecules. They constitute a group of various stable compounds. Advanced glycation end-products are considered potential biomarkers of metabolic disorders. However, so far only a few methods to determine the level of individual AGEs have been developed.
Objectives: The aim of the study was to compare the efficiency of the slot-dot blot method and direct enzyme-linked immunosorbent assay (ELISA) in detecting non-standard epitopes of methylglyoxal (MGO)-modified proteins (AGE4) found in diabetes serum in trace amounts, and to assess AGE4 in diabetes and associated metabolic abnormalities.
Material and methods: The presence of AGE4 was detected using 2 methods: direct ELISA and the slot-dot blot method - a newly developed immunoassay based on monoclonal, commercially available antibody detection of non-standard AGE epitopes. AGE4 quantification, expressed as median AGE4 in arbitrary units (AU) and AGE4 positivity (the percent of samples with detectable AGE4) was related to diabetes-associated metabolic abnormalities, complications and treatment.
Results: Slot-dot blot was significantly more efficient than ELISA in detecting non-standard AGE4 epitopes. AGE4 positivity was less frequent in patients with microangiopathy and in those with polyneuropathy. In patients with abnormal glucose metabolism, metformin treatment was associated with higher AGE4. AGE4 positivity was significantly lower in gliptin-treated patients. Multivariate analysis showed that polyneuropathy and obesity were independently associated with AGE4 positivity, with odds ratios (ORs) of 0.21 and 3.02, respectively. Moreover, logistic regression showed that AGE4 positivity and HbA1c are independent predictors of polyneuropathy. Considering both indicators allows correct classification of 70.4% of cases with a general accuracy of 76%.
Conclusions: The slot dot-blot method detects compounds found in serum in trace amounts. Accumulation of AGE4 was associated with glucose metabolism abnormalities. A tendency toward AGE4 positivity was less frequent in patients with microangiopathy and in non-treated and gliptin-treated diabetes patients.
Keywords: ELISA; advanced glycation end-products (AGEs); diabetes; methylglyoxal; slot dot-blot.