The monoclonal antibody CML26 recognizes human CML (carboxymethyl-lysine). CML is known to be formed from the oxidation of both carbohydrates and lipids. This makes CML- a biomarker of general oxidative stress. Carboxymethyl-lysine (CML) is a well-characterized glycoxidation product that accumulates in tissues with age, and its rate of accumulation is accelerated in diabetes. Glycoxidation products are a subset of advanced glycation endproducts (AGEs) that are formed by the nonenzymatic glycation and subsequent irreversible oxidation of proteins. Oxidative stress and protein modification have been implicated in the pathogenesis of the chronic complications of diabetes, including nephropathy and atherosclerosis. The accumulation of CML in long-lived tissue such as skin collagen reflects oxidative stress over an extended period of the life-span, and has been shown to be greater in patients with diabetic complications than those without complications.
Frozen sections, Immuno assays, Immuno fluorescence, Paraffin sections, Western blot
P: fixation in 4% formalin; cardiac tissue sections (4 mm) deparaffinised for 10 min in xylene at room temperature, dehydrated by decreasing ethanol. Sections stained with haematoxylin and eosin. Blocking endogenous peroxidase activity with 0.3% hydrogen peroxide in methanol for 30 min. No heating to prevent artificial induction of CML. (Ref 1)
IF: After fixation in 2% phosphate-buffered glutaraldehyde solution the heart tissue was post-fixed in 1% osmium tetroxide. The tissue was dehydrated through a graded series of ethanol. 0.53.0- mm-thick sections were cut with a glass knife.(Ref 1)
For immunohistochemistry, dilutions to be used depend on detection system applied. It is recommended that users test the reagent and determine their own optimal dilutions. The typical starting working dilution is 1:50. For functional studies, in vitro dilutions have to be optimized in user’s experimental setting.