Japan Institute for the Control of Aging: JaICA Oxidative Stress
Rev.081024
Anti-methylglyoxal antibody
(Anti MG monoclonal antibody, specific for argpyrimidine, clone #3C)
FOR RESEARCH USE ONLY
About methylglyoxal (MG)
Methylglyoxal (MG), an endogenous metabolite that increases in diabetes and is a common intermediate in the Maillard reaction (glycation), reacts with proteins and forms advanced glycation end products (AGE). MG reacts with arginine residue in protein and forms numerous numbers of adducts, such as argpyrimidine. This antibody is specific for argpyrimidine: N-(5-hydroxy-4,6-dimethylpyrimidine-2-yl)-L-ornithine.
Specifications
Clone #: 3C
Antigen: MG-modified keyhole-lympet hemocyanine
Specificity: Specific for MG-modified protein (especially Argpyrimidine)
Subclass: Mouse IgG2a(kappa)
Form: Frozen (10mM PBS containing 0.1% NaN3 and 0.5% BSA, 100 µg/mL IgG. Protein A purified.
Application: Immunohistochemistryˇ¨It is recommended that the antibody be tried at 0.5-1.0 µg/mL on paraformaldehyde fixed tissue.
 
Product name Code Content
Anti Methylglyoxal (MG) monoclonal antibody MMG-030n 30 µg IgG

Manufactured by NOF corporation, Japan.

FOR RESEARCH USE ONLY. Not for diagnostic, medical or other use.
 
 
 
Immunohistochemistry
Procedure
1) Paraffin sections were deparaffinized and rehydrated.
2) Sections were quenched for several minutes with 3% hydrogen peroxide, rinsed in PBS, pretreated for 30 min with 3% nonimmune animal serum in PBS.
3) Sections were incubated overnight at 4°C with a antibody at a dilution of 1 µg/mL.
4) Antibody binding was visualized by the avidin-biotin-immuno peroxidase complex method using the Vectastain ABC kit (Vector, Burlingame, CA) according to the manufacturer's instructions.
5) 3,3-Diaminobenzidine tetrahydrochloride was used as the chromogen.
6) Hematoxylin or Eosin was used as the counter stain.
Technical Note
1) Sections from which the primary antibodies were omitted served as negative reaction controls.
2) For frozen materials, samples were fixed in 10% formalin, immersed in 30% sucrose in PBS, embedded at OCT (Sakura, Tokyo, Japan), and stored at 80°C.
3) For paraffin embedded materials, samples were fixed in 10% formalin, dehydrated, embedded in paraffin, and stored at room temperature.
4) Microwave treatment improves the target staining, whereas it declines non-specific background.
5) Protease K treatment declines the target staining.
Positive control
  Aortic wall in the renal tissue of diabetic nephropathy or aortic wall of atherosclerosis.
References
1) Protein modification by a Maillard reaction intermediate methylglyoxal. Immunochemical detection of fluorescent 5-methylimidazolone derivatives in vivo. Uchida K, Khor OT, Oya T, Osawa T, Yasuda Y, Miyata T. FEBS Lett. 410(2-3), p313-318. (1997)
2) Methylglyoxal Modification of Protein -Chemical and Immunochemical Characterization of Methylglyoxal-Arginine Adduct. T.Oya, N.Hattori, Y.Mizuno, S.Miyata, S.Maeda, T.Osawa and K.Uchida. J.Biological Chemistry, 274, 18492-18502 (1999)
3) Advanced glycation and lipidoxidation of the peritoneal membrane: respective roles of serum and peritoneal fluid reactive carbonyl compounds. Miyata T, Horie K, Ueda Y, Fujita Y, Izuhara Y, Hirano H, Uchida K, Saito A, van Ypersele de Strihou C, Kurokawa K. Kidney Int. 58(1), p425-435.(2000)
4) Argpyrimidine, a blue fluorophore in human lens proteins: high levels in brunescent cataractous lenses. Padayatti PS, Ng AS, Uchida K, Glomb MA, Nagaraj RH. Invest Ophthalmol Vis Sci. 42(6), p1299-1304. (2001)
5) High concentrations of glucose induce synthesis of argpyrimidine in retinal endothelial cells. Padayatti PS, Jiang C, Glomb MA, Uchida K, Nagaraj RH. Curr Eye Res. 23(2), p106-115. (2001)
6) Curcumin Inhibits ROS Formation and Apoptosis in Methylglyoxal- Treated Human Hepatoma G2 Cells. WEN-HSIUNG CHAN, HSIN-JUNG WU and YAN-DER HSUUW. Ann. N.Y. Acad. Sci. 1042: 372-378 (2005)
FOR RESEARCH USE ONLY. Not for diagnostic, medical or other use.