Human Haptoglobin / Zonulin ELISA Kit

Catalog No.: BEK1261

Size: 96T

Range: 156 pg/ml-10,000 pg/ml

Sensitivity < 10 pg/ml

Storage and Expiration: Store at 2-8℃ for 6 months, or at -20℃ for 12 months.

Application: For quantitative detection of Haptoglobin in human serum, plasma, body fluids, tissue homogenate or cell culture supernatants.

Introduction

Haptoglobin (Hp) is an abundant and conserved plasma glycoprotein, which binds acellular adult hemoglobin (Hb) dimers with high affinity and facilitates their rapid clearance from circulation after hemolysis. Humans possess three main phenotypes of Hp, designated Hp 1-1, Hp 2-1, and Hp 2-2. These variants exhibit diverse structural configurations and have been reported to be functionally nonequivalent. Hp fractions examined are equivalent to one another with respect to Hb binding and associated stability and redox properties and that this result should be taken into account in the design of phenotype-specific Hp therapeutics aimed at countering Hb-mediated vascular disease. Hp scavenges free hemoglobin following malaria-induced hemolysis.

Principle of the Assay  

This kit was based on sandwich enzyme-linked immune-sorbent assay technology. Anti-Haptoglobin polyclonal antibody was pre-coated onto 96-well plates. And the biotin conjugated anti-Haptoglobin polyclonal antibody was used as detection antibodies. The standards, test samples and biotin conjugated detection antibody were added to the wells subsequently, and wash with wash buffer. Avidin-Biotin-Peroxidase Complex was added and unbound conjugates were washed away with wash buffer. TMB substrates were used to visualize HRP enzymatic reaction. TMB was catalyzed by HRP to produce a blue color product that changed into yellow after adding acidic stop solution. The density of yellow is proportional to the Haptoglobin amount of sample captured in plate. Read the O.D. absorbance at 450nm in a microplate reader, and then the concentration of Haptoglobin can be calculated.

Kit components

1. One 96-well plate pre-coated with anti-Human Haptoglobin antibody
2. Lyophilized Human Haptoglobin standards: 2 tubes (10 ng / tube)
3. Sample / Standard diluent buffer: 30 ml
4. Biotin conjugated anti-Human Haptoglobin antibody (Concentrated): 230 μl. Dilution: 1:50
5. Antibody diluent buffer: 12 ml
6. Avidin-Biotin-Peroxidase Complex (ABC) (Concentrated): 130 μl. Dilution: 1:100
7. ABC diluent buffer: 12 ml
8. TMB substrate: 10 ml
9. Stop solution: 10 ml
10. Wash buffer(25X): 30 ml

Note: Reconstitute standards and test samples with Kit Component 3.

Material Required But Not Provided

1. 37℃ incubator
2. Microplate reader (wavelength: 450nm)
3. Precise pipette and disposable pipette tips
4. Automated plate washer
5. ELISA shaker
6. 1.5ml of Eppendorf tubes
7. Plate cover
8. Absorbent filter papers
9. Plastic or glass container with volume of above 1L