Prostaglandin E2 (PGE2) is derived from PGH2 which in turn is synthesized from arachidonic acid through the cyclooxygenase pathway. Many cell types, such as epithelial cells, fibroblasts, and macrophages produce PGE2. PGE2 possesses vasoactivity, modulates immune funtions, regulates sleep-awake cycles and exhibits many other activities.
Principle Of The Assay:
This is an monoclonal ELISA (Enzyme-Linked ImmunoSorbent Assay) for the quantitative analysis of Prostaglandin E2 levels in biological fluid. This test kit operates on the basis of competition between the enzyme conjugate and the PGE2 in the sample for a limited number of binding sites on the antibody coated plate. The sample or standard solution is first added to the microplate. Next, the diluted enzyme conjugate is added and the mixture is shaken and incubated at room temperature for one hour. During the incubation, competition for binding sites is taking place. The plate is then washed removing all the unbound material. The bound enzyme conjugate is detected by the addition of substrate which generates an optimal color after 30 minutes. Quantitative test results may be obtained by measuring and comparing the absorbance reading of the wells of the samples against the standards with a microplate reader at 650nm. The extent of color development is inversely proportional to the amount of PGE2 in the sample or standard. For example, the absence of PGE2 in the sample will result in a bright blue color, whereas the presence of PGE2 will result in decreased or no color development.
P9053-30A: EIA Buffer: 30ml. Provided to dilute enzyme conjugate and PGE2 standards.
P9053-30B: Wash Buffer (10X): 20ml. Dilute 10 fold with deionized water. Diluted wash buffer is used to wash all unbound enzyme conjugate, samples and standards from the wells after the one hour incubation.
P9053-30C: Substrate: 20ml. Stabilized 3,3', 5,5' Tetramethylbenzidine (TMB) plus Hydrogen Peroxide (H2O2) in a single bottle. It is used to develop the color in the wells after they have been washed. Light sensitive. Keep substrate refrigerated.
P9053-30D: Extraction Buffer (5X): 30ml. Dilute 5 fold with deionized water. This buffer is used for diluting extracted and non-extracted samples.
P9053-30E: Prostaglandin E2 Enzyme Conjugate: Two vials of lyophilized PGE2 HRP conjugate. Reconstitution with 75ul of deionized water results in a 50:1 concentrate. Blue capped vials.
P9053-30F: Prostaglandin E2 Standard: 100ul. PGE2 standard provided at the concentration of 1ug/ml. Glass vial. Solution is viscous, take precaution when pipetting.
P9053-30G: Prostaglandin E2 Antibody Coated Plate: A 96 well Costar microplate precoated with monoclonal mouse antibody against PGE2 The plate is ready for use as is. Do not wash.
Storage and Stability:
Store all components at 4 degrees C except P9053-30E. Store lyophilized P9053-30E at -20 degrees C. Store reconstituted P9053-30E at 4 degrees C.