scholarly journals A MORE SENSITIVE AND SPECIFIC HISTOCHEMICAL PEROXIDASE STAIN FOR THE LOCALIZATION OF CELLULAR ANTIGEN BY THE ENZYME-ANTIBODY CONJUGATE METHOD

1974 ◽  
Vol 22 (12) ◽  
pp. 1135-1140 ◽  
Author(s):  
E. EARL WEIR ◽  
THOMAS G. PRETLOW ◽  
ANNETTE PITTS ◽  
EDWIN E. WILLIAMS
Keyword(s):  
Hydrogen Peroxide ◽  
Horseradish Peroxidase ◽  
Sensitivity And Specificity ◽  
Ph Optimum ◽  
Concentration Time ◽  
Antibody Conjugate ◽  
Cellular Antigen

In an attempt to increase the sensitivity and specificity of immunohistochemical procedures using peroxidase-labeled antibody, we have modified the technique. This modification takes advantage of the pH optimum of horseradish peroxidase. The best buffer salts, hydrogen peroxide concentration, time of staining, fixation after incubation with labeled antibody and counterstains were determined empirically. Compounds similar to the substrate are known to undergo photooxidation, and specificity was enhanced by keeping the solutions in the dark.

scholarly journals Oxidation of 2-nitropropane by horseradish peroxidase. Involvement of hydrogen peroxide and of superoxide in the reaction mechanism

1978 ◽  
Vol 175 (2) ◽  
pp. 601-606 ◽  
Author(s):  
Johan De Rycker ◽  
Barry Halliwell
Keyword(s):  
Hydrogen Peroxide ◽  
Superoxide Dismutase ◽  
Reaction Mechanism ◽  
Horseradish Peroxidase ◽  
Aqueous Solutions ◽  
Oxidation Reaction ◽  
Superoxide Ion ◽  
Radical Species ◽  
Ph Optimum ◽  
Nitro Aromatics

Incubation of aqueous solutions of 2-nitropropane in air causes a slow oxidation reaction that generates H2O2. Purified horseradish peroxidase catalyses the oxidation of such preincubated 2-nitropropane solutions according to the equation: [Formula: see text] The pH optimum is 4.5 and Km for 2-nitropropane is 16mm. Other nitroalkanes or nitro-aromatics tested are not oxidized at significant rates by peroxidase. H2O2 or 2,4-dichlorophenol increases the rate of 2-nitropropane oxidation by peroxidase. Catalase inhibits the reaction completely. Superoxide dismutase or mannitol, a scavenger of the hydroxyl radical, OH., each inhibits partially. Aniline and guaiacol are also powerful inhibitors of 2-nitropropane oxidation. It is suggested that peroxidase uses the traces of H2O2 generated during preincubation of 2-nitropropane to catalyse oxidation of this substrate into a radical species that can reduce O2 to the superoxide ion, O2−..O2−., or OH. derived from it, then appears to react with more nitropropane, generating further radicals and H2O2 to continue the oxidation. Inhibition by aniline and guaiacol seems to be due to a competition for H2O2.

scholarly journals Monitoring the heme iron state in horseradish peroxidase to detect ultratrace amounts of hydrogen peroxide in alcohols

RSC Advances ◽  
2021 ◽  
Vol 11 (17) ◽  
pp. 9901-9910
Author(s):  
Raheleh Ravanfar ◽  
Alireza Abbaspourrad
Keyword(s):  
Hydrogen Peroxide ◽  
Horseradish Peroxidase ◽  
Oxidative Damage ◽  
Heme Iron ◽  
Aqueous Systems ◽  
Iron State ◽  
Low Concentrations

Despite the importance of hydrogen peroxide (H2O2) in initiating oxidative damage and its connection to various diseases, the detection of low concentrations of H2O2 (<10 μM) is still limited using current methods, particularly in non-aqueous systems.

scholarly journals The Reaction of Ferrous Horseradish Peroxidase with Hydrogen Peroxide

1970 ◽  
Vol 245 (9) ◽  
pp. 2409-2413
Author(s):  
Robert W. Noble ◽  
Quentin H. Gibson
Keyword(s):  
Hydrogen Peroxide ◽  
Horseradish Peroxidase

scholarly journals Fluorescent Derivatization of Aromatic Carboxylic Acids with Horseradish Peroxidase in the Presence of Excess Hydrogen Peroxide

2015 ◽  
Vol 31 (1) ◽  
pp. 37-44
Author(s):  
Junichi ODO ◽  
Masahiko INOGUCHI ◽  
Hiroyuki AOKI ◽  
Yuto SOGAWA ◽  
Masahiro NISHIMURA
Keyword(s):  
Hydrogen Peroxide ◽  
Horseradish Peroxidase ◽  
Carboxylic Acids ◽  
Aromatic Carboxylic Acids ◽  
Excess Hydrogen

scholarly journals Detection of a Tryptophan Radical as an Intermediate Species in the Reaction of Horseradish Peroxidase Mutant (Phe-221 → Trp) and Hydrogen Peroxide

1998 ◽  
Vol 273 (24) ◽  
pp. 14753-14760 ◽  
Author(s):  
Atsushi Morimoto ◽  
Motomasa Tanaka ◽  
Satoshi Takahashi ◽  
Koichiro Ishimori ◽  
Hiroshi Hori ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Horseradish Peroxidase ◽  
Intermediate Species

scholarly journals Factors affecting the sensitivity and specificity of the cytochemical reaction for the anti-horseradish peroxidase antibody in lymph tissue sections.

1980 ◽  
Vol 28 (7) ◽  
pp. 645-652 ◽  
Author(s):  
W Straus
Keyword(s):  
Horseradish Peroxidase ◽  
Lymph Nodes ◽  
Sensitivity And Specificity ◽  
Peroxidase Activity ◽  
Additional Advantage ◽  
Tissue Sections ◽  
Endogenous Peroxidase ◽  
Cytochemical Reaction ◽  
Cold Acetone ◽  
Antibody Reaction

Factors which increase the sensitivity and specificity of the cytochemical reaction for the antibody to horseradish peroxidase (HRP) in precursors of plasma cells and in lymphocytes were studied in sections of popliteal lymph nodes of rats. The lymph nodes were removed 3-5 days after a secondary injection of HRP into the footpads and were fixed for 5 hr in a 4% cold formaldehyde solution (Straus W: Histochemistry 53:273, 1977). Brief postfixation of the frozen sections with cold acetone improved the retention of the antigen at the sites of the antibody in the precursor cells, and it improved the quality of fixation without appreciably weakening the antigen-binding capacity of the antibody. The cytochemical reaction for the anti-HRP antibody was intensified by staining with diaminobenzidine (DAB) and H2O2 at pH 5-6, or by staining at pH 7.4 in the presence of imidazole. Imidazole partially inhibited endogenous peroxidase activity. Pretreatment with phenylhydrazine prevented nonspecific background adsorption of HRP. Phenylhydrazine had the additional advantage of inhibiting most of the endogenous peroxidase activity (Straus W: J Histochem Cytochem 20:949, 1972). The intensity of the antibody reaction in the proplasma cells developing in the medullary cords varied greatly depending on the stage of maturation from lymphocytes and blast cells. Many lymphocytes in the cortex of the lymph node showed a strong perinuclear antibody reaction when the tissue sections were postfixed with cold acetone, and the peroxidase complexed to the antibody was visualized by staining with DAB and H2O2 at pH 5-6. The antibody reaction also occurred at the surgace of many lymphocytes when the tissue sections, postfixed with cold acetone, were stained with DAB and H2O2 at pH 7.4 in the presence of imidazole. Other lymphocytes showed a strong surface, perinuclear, and cytoplasmic antibody reaction after staining at pH 5-6 as well as after staining at pH 7.4, while yet other lymphocytes remained unstained.

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