5-HIAA, Human (5-hydroxy-3-indole Acetic Acid) BioAssay™ ELISA Kit
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The primary carcinoid tumor is usually derived from the enterochromaffin cells of the midgut and is located most frequently in the terminal ileum. Carcinoid tumors generally secrete various amounts of indoles. The carcinoid syndrome is generally characterized by an increased urinary excretion of 5-hydroxy-3-indole acetic acid (5-HIAA), the end product of serotonin (5-HT) metabolism. Traditionally, 5-hydroxy-3-indole acetic acid is assayed by diazotization with nitrosonaphtol to form a purple color. However, it is well documented that many other substances present in the urine interfere with this reaction to give false-positive results. Attempts were made to overcome this problem by a combination of ion exchange chromatography and fluorometry. These methods, however, lack sensitivity and are time consuming. Recently, high performance liquid chromatographic analyses of 5-HIAA with fluorometry in the ultraviolet region of the spectrum or electrochemical detection have been described. Both methods require solvent extraction because of the numerous interfering compounds present in urine. The 5-HIAA enzyme immunoassay is a new and simple method for the quantification of this important marker of carcinoid syndrome in small urine samples.
Enzyme immunoassay for the quantitative determination of 5-HIAA in human urine.
The assay procedure follows the basic principle of competitive ELISA whereby there is competition between a biotinylated and a non-biotinylated antigen for a fixed number of antibody binding sites. The amount of biotinylated antigen bound to the antibody is inversely proportional to the analyte concentration of the sample. When the system is in equilibrium, the free biotinylated antigen is removed by a washing step and the antibody bound biotinylated antigen is determined by use of anti-biotin alkaline phosphatase as marker and p-nitrophenyl phosphate as substrate. Quantification of unknowns is achieved by comparing the enzymatic activity of unknowns with a response curve prepared by using known standards.
Microtiter Plate 1x96 wells
5-HIAA Pab 1x5ml
5-HIAA (Biotin) 1x5ml
Biotin Polyclonal antibody (AP) (100X)1x0.2ml
Standard 0mg/L 1x500ul
Standard 0.4mg/L 1x500ul
Standard 1.0mg/L 1x500ul
Standard 2.75mg/L 1x500ul
Standard 7.5mg/L 1x500ul
Standard 20mg/L 1x500ul
Standard 55mg/L 1x500ul
Control 1 normal, 1x1vial
Control 2 pathological , 1x1vial
Methylation Reagent 1x2ml
HCl Solution, 1x1ml, 0.1M HCl.
Assay Buffer, Concentrate, 10X 1x50ml
Dilution Reagent 1x4ml
Wash Buffer, Concentrate, 20x, 1x50ml
PNPP Substrate, 9x1Tablets
Substrate Buffer 1x27ml
Stop Solution 1x5ml
Storage and Stability:
Store all components at 4°C. Stable for at least 6 months. For maximum recovery of product, centrifuge the original vial prior to removing the cap.