Fig. 5
Size distribution of high molecular weight AGE-proteins from diabetic patients on hemodialysis.
High molecular weight protein fractions obtained from diabetic patients with end-stage renal disease on hemodialysis were subjected to gel filtration chromatography (Sephacryl S-200 column: 1.5 × 110 cm). The eluted fractions were monitored for non-CML AGE (glyceraldehyde, ◯; glycolaldehyde, ▲; methylglyoxal, ■; glyoxal, ◊) by ELISA (data from triplicate determinations), AGE fluorescence (Ex = 360 nm / Em = 440 nm, △) and absorbance at 280 nm (●). Elution of aldolase (160,000 Da), BSA (67,000 Da) chymotrypsinogen A (25,000 Da) and vitamin B12 (1,355 Da) (molecular weight markers) is indicated by arrows. (A) Chromatography monitored by AGE fluorescence and absorbance at 280 nm. (B) Chromatography with affinity-purified non-CML AGE antibodies obtained by incubation of AGE-RSA with glyceraldehyde and glycolaldehyde. (C) Chromatography with affinity-purified non-CML AGE antibodies obtained by incubation of AGE-RSA with methylglyoxal and glyoxal.