Merck
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P8250

Sigma-Aldrich

Peroxidase from horseradish

Type II, essentially salt-free, lyophilized powder, 150-250 units/mg solid (using pyrogallol)

Synonym(s):
Donor:hydrogen-peroxide oxidoreductase, Horseradish peroxidase
CAS Number:
Enzyme Commission number:
EC Number:
MDL number:
eCl@ss:
32160410
NACRES:
NA.54

Quality Level

biological source

horseradish

type

Type II

form

essentially salt-free, lyophilized powder

specific activity

150-250 units/mg solid (using pyrogallol)

mol wt

~44 kDa

solubility

0.1 M phosphate buffer: soluble (pH 6.0)
H2O: soluble

absorbance ratio

RZ ≥1.8

application(s)

diagnostic assay manufacturing

storage temp.

2-8°C

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General description

Horseradish peroxidase is isolated from horseradish roots (Amoracia rusticana) and belongs to the ferroprotoporphyrin group of peroxidases. HRP is a single chain polypeptide containing four disulfide bridges. It is a glycoprotein containing 18% carbohydrate. The carbohydrate composition consists of galactose, arabinose, xylose, fucose, mannose, mannosamine, and galactosamine depending upon the specific isozyme. Its molecular weight (~44 kDa) includes the polypeptide chain (33,890 Daltons), hemin plus Ca2+ (~700 Daltons), and carbohydrate (~9,400 Daltons). At least seven isozymes of HRP exist. The isoelectric point for horseradish peroxidase isozymes ranges from 3.0 - 9.0.

Application

The enzyme has been used as a comparison during the peroxidase assay of extract from mature tall fescue leaf blades. It has also been used to measure H2O2 production.
Horseradish peroxidase (HRP) is isolated from horseradish roots (Amoracia rusticana). It is used in biochemistry applications such as western blots, ELISA and Immunohistochemistry. Horseradish peroxidase is used to amplify a weak signal and increase detectability of a target molecule, such as a protein. Horseradish peroxidase, product P8250, has been used to study nonoral antigens in inflamed gingiva and Ebola virus glycoprotein toxicity.

Packaging

5000, 25000, 50000, 100000, 200000 units in glass bottle

Biochem/physiol Actions

HRP readily combines with hydrogen peroxide (H2O2) and the resultant [HRP-H2O2] complex can oxidize a wide variety of hydrogen donors. The optimal pH is 6.0-6.5 and the enzyme is most stable in the pH range of 5.0-9.0. HRP can be conjugated to antibodies by several different methods including glutaraldehyde, periodate oxidation, through disulfide bonds, and also via amino and thiol directed cross-linkers. It is smaller and more stable than the enzyme labels β-galactosidase and alkaline phosphatase. Hence, it is the most desired label. Also, its glycosylation leads to lower non-specific binding. It is also used for the determination of glucose and peroxides in solution. Sodium azide, cyanide, L-cystine, dichromate, ethylenethiourea, hydroxylamine, sulfide, vanadate, p-aminobenzoic acid, and Cd2+, Co2+, Cu2+, Fe3+, Mn2+, Ni2+, Pb2+ ions are found to inhibit the enzyme activity.
When incubated with a substrate, horseradish peroxidase produces a coloured, fluorimetric, or luminescent derivative of the labeled molecule, allowing quantification. Horseradish peroxidase has been shown to slightly reduce the level of inhibition in a cydAB mutant.

Quality

Preliminary work shows it to contain at least five isoenzymes.

Unit Definition

One pyrogallol unit will form 1.0 mg purpurogallin from pyrogallol in 20 sec at pH 6.0 at 20 °C.

Analysis Note

The RZ (Reinheitszahl) is the absorbance ratio A403/A275 determined at 0.5-1.0 mg/ml in deionized water. It is a measure of hemin content, not enzymatic activity. Even preparations with high RZ may have low enzymatic activity.

Other Notes

Pictograms

Health hazard

Signal Word

Danger

Hazard Statements

Hazard Classifications

Resp. Sens. 1

Storage Class Code

11 - Combustible Solids

WGK

WGK 3

Flash Point(F)

Not applicable

Flash Point(C)

Not applicable

Personal Protective Equipment

dust mask type N95 (US), Eyeshields, Gloves

Certificate of Analysis

Enter Lot Number to search for Certificate of Analysis (COA).

Certificate of Origin

Enter Lot Number to search for Certificate of Origin (COO).

More Documents

Quotes and Ordering

W E Bullock et al.
The Journal of experimental medicine, 165(1), 195-210 (1987-01-01)
The principal host cell of H. capsulatum (Hc) is the M phi within which the pathogenic yeast phase of the fungus multiplies during active disease. The initial interaction between Hc yeasts and M phi therefore is a crucial step in
S M Mallison et al.
Infection and immunity, 56(4), 823-830 (1988-04-01)
In vitro experimentation indicates that periodontitis-associated bacteria contain potent polyclonal B-cell activators (PBA). We reasoned that if PBA were operative in vivo, plasma cells specific for nonoral antigens should be present in the inflamed gingival tissues, which are characterized by
J W Macadam et al.
Plant physiology, 99(3), 872-878 (1992-07-01)
Cessation of cell expansion has been associated with cell wall cross-linking reactions catalyzed by peroxidase. This study utilized two genotypes of tall fescue (Festuca arundinacea Schreb.) that differ in length of the leaf elongation zone to investigate the relationship between
Nancy J Sullivan et al.
Journal of virology, 79(1), 547-553 (2004-12-15)
Ebola virus infection causes a highly lethal hemorrhagic fever syndrome associated with profound immunosuppression through its ability to induce widespread inflammation and cellular damage. Though GP, the viral envelope glycoprotein, mediates many of these effects, the molecular events that underlie
Chuan Chiang-Ni et al.
Journal of microbiology, immunology, and infection = Wei mian yu gan ran za zhi, 49(6), 837-842 (2015-02-05)
The phenotypic heterogeneity of the human pathogen Streptococcus pyogenes [group A streptococcus (GAS)] is associated with bacterial virulence variation. During invasive GAS infection, mutations in the two-component regulatory system covR/covS leads to increases in hyaluronic acid capsule production, virulence genes

Protocols

Enzymatic Assay of Diamine Oxidase (E.C. No. 1.4.3.22)

To standardize a procedure for the enzymatic assay of Diamine Oxidase.

Enzymatic Assay of Alcohol Oxidase (EC 1.1.3.13)

This procedure may be used for all Alcohol Oxidase products. The continuous spectrophotometric rate determination (A405, Light path = 1 cm) is based on the following reactions.

Enzymatic Assay of Cholesterol Oxidase

This procedure applies to products that have a specification for the enzymatic activity of cholesterol oxidase. This assay is NOT to be used to assay Cholesterol Oxidase from Schizophyllum commune (Discontinued Product Number C7274) and from Brevibacterium sp. (Discontinued Product Number C8153).

Spectrophotometric Determination of Reinheitszahl (RZ) for Peroxidase (EC 1.11.1.7)

Reinheitszahl (RZ) is the ratio of absorbance due to hemin (A403, Soret region) to absorbance due to protein (A275).

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