What is 4-HHE ?
4-hydroxy-2-alkenal is one of the major lipid peroxidation products, and shows many biological effects such as high toxicity to cells. Among them, 4-hydroxy-2-hexenal (HHE) is an aldehyde formed during peroxidation of n-3 fatty acids such as docosahexaenoic acid. HHE is highly reactive aldehyde and reacts with histidine residue of protein to form Michael-addition type adducts. This antibody is specific for HHE-histidine Michael adduct (HHE-His) and enable to detect HHE-His in the tissue samples.
|Antigen:||HHE-modified keyhole-lympet hemocyanine|
|Specificity:||Specific for HHE-modified protein (especially HHE-HIs Michael adduct)|
|Subclass:||IgG1 kappa (clone HHE53)|
|Form:||Frozen in 10mM PBS containing 0.1 %NaN3 and 0.5% BSA.|
|Application:||Immunohistochemistry;It is recommended that the antibody be tried at 0.5-1.0 micro g/mL on paraformaldehyde fixed tissue|
|Storage:||Less than -20°C|
Specifity of anti 4-HHE antibody.
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 micro gram/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.
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.
Aortic wall in the renal tissue of diabetic nephropathy or aortic wall of atherosclerosis.