Haptoglobin, Human BioAssay(TM) ELISA Kit (HP)

Haptoglobin (abbreviated as Hp) is a protein in the blood plasma that binds free hemoglobin released from erythrocytes with high affinity and removes it from the circulation to prevent kidney injury, and iron loss following hemolysis. Also, haptoglobin prevents bacteria from using the iron present in hemoglobin to grow, regulates the activity of many cell types of the immune system, and is an extracellular chaperone. Haptoglobin, as an antioxidant, has antibacterial activity and plays a role in modulating many aspects of the acute phase response. In addition, haptoglobin acts as a potent immunosuppressor of lymphocyte function and modulates the helper T-cell type 1 and type 2 (Th1/Th2) balance within the body. Haptoglobin plays a role in stimulation of angiogenesis and has highly potent cholesterol crystallization-promoting activity, also. Changes in the measured concentrations of haptoglobin in serum may help to assess the disease status of patients with inflammations, infections, malignancy etc. (increases) as well as in haemolytic conditions (decreases). Haptoglobin in its simplest form consists of two a- and two b-chains, connected by disulfide bridges. The chains originate from a common precursor protein which is proteolytically cleaved during protein synthesis. Haptoglobin exists in two allelic forms in the human population, so called Hp1 and Hp2; the latter one having arisen due to the partial duplication of Hp1 gene. Three phenotypes of haptoglobin therefore are found in humans: Hp1-1, Hp2-1, and Hp2-2. Hp of different phenotypes have been shown to bind hemoglobin with different affinities. Hp 1-1 is biologically the most effective in binding free hemoglobin and suppressing inflammatory responses associated with free hemoglobin. Hp 2-2 is biologically the least active, and Hp 2-1 is moderately active. The possible association of allelic polymorphism of haptoglobin with various pathologic conditions such as cardiovascular disease, autoimmune disorders, malignancy has been studied. Hp 2-2 phenotype presents characteristics of being an independent risk factor for cardiovascular disease (CVD) and more significant in diabetic CVD. In chronic kidney disease (CKD), due to various processes that take place simultaneously, the combined effect is different in the elderly and the young. The functional differences between the Haptoglobin phenotypes are most probably due to their antioxidative property and to macrophage activation with differential release of cytokines.\n\nIntended Use:\nHuman Haptoglobin ELISA kit is to be used for the in vitro quantitative determination of human Haptoglobin in human serum, human plasma, cell lysate and buffered solution. The assay will recognize both native and recombinant human Haptoglobin. This kit has been configured for research use only and is not to be used in diagnostic procedures.\n\nSensitivity:\nThe minimal detectable dose of human Haptoglobin was calculated to be 0.78ng/ml, by subtracting two standard deviations from the mean of 10 zero standard replicates (ELISA buffer, S0) and intersecting this value with the standard curve obtained in the same calculation.\n\nSpecificity:\nThe following substances have been tested and found to have no cross-reactivity: human Albumin, IgG, Transferrin, Fibrinogen, a-2 marcroglobulin, Complement C3, Antitrypsin.\n\nTest Principle:\nThe design of this assay is based on a sandwich Enzyme-Linked Immunosorbent Assay (ELISA). The microtiter plate provided in this kit has been pre-coated with a monoclonal antibody specific to human Haptoglobin. Samples are pippetted into these wells. Nonbound Haptoglobin and other components of the sample should be removed by washing, then biotin- conjugated monoclonal antibody specific to Haptoglobin added. In order to quantitatively determine the amount of Haptoglobin present in the sample, Avidin conjugated to Horseradish Peroxidase (HRP) should be added to each microplate well. The final step, a TMB-substrate solution added to each well. Finally, a sulfuric acid solution is added and the resulting yellow colored product is measured at 450nm. Since the increases in absorbency is directly proportional to the amount of captured Haptoglobin.\n\nKit Components:\nMicroplate: 1x96 wells\nIncubation buffer: 1x30ml\nWashing buffer, (10x): 1x100ml\nStandard protein: 1 glass vial lyophilized\nStandard/sample dilution buffer: 1x25ml\nSecond antibody: 1 glass vial lyophilized\nAV-HRP, (100x): 1x150ul\nSecondary antibody/AV-HRP dilution buffer: 1x25ml\nSubstrate (TMB): 1x20ml\nStop solution: 1x20ml\n\nStorage and Stability:\nStore components at 4 degrees C. Stable for at least 6 months. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.

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SPECIFICATIONS

Catalog Number

H1820-21

Size

96Tests

References

1. Sadrzadeh SM, Bozorgmehr J. Am J Clin Pathol. 2004; vol.121: pp.S97-104. 2. Wassell J. Clin Lab. 2000; vol.46(11-12): pp.547-52. 3. Dobryszycka W. Eur J Clin Chem Clin Biochem. 1997; vol.35(9): pp.647-54.