14,15 DHET ELISA Assay Kit

$330.00$2,620.00

The Eagle Biosciences 14,15 DHET ELISA Assay kit is intended for the quantitative determination of 14,15 DHET in biological samples by enzyme linked immunoassay (ELISA).  The Eagle Biosciences 14,15 DHET ELISA Assay kit is for research use only and not to be used in diagnostic procedures.

Clear
SKU: 14D39-K01 0 Categories: , ,

14,15 DHET ELISA Assay Kit

For Research Use Only

Size: 1×96 wells
Sensitivity: 0.01 ng/ml
Dynamic Range: 0.01 – 1000 ng/ml
Incubation Time: 2.5 hours
Sample Type: Serum, Plasma, Tissue, Biological Fluids
Sample Size: 100 µl

Product manufactured in the USA

Additional Information

Assay Background

This competitive 14,15 DHET ELISA Assay kit is for determination of 14,15-DHET levels in biological samples.  Specificity of the 14,15 DHET ELISA Assay kit has been published.  The 14,15-DHET level exhibited strong positive correlation with hypertension, brain injury and stroke in rodents.  14,15-DHET is a representative metabolite of soluble epoxide hydrolase-mediated metabolism of EETs, which are generated by arachidonic acid epoxygenase activity of cytochromes P450.  Human blood 14,15-DHET levels were measured using the 14,15-DHET ELISA kit.  Increased 14,15-DHET levels of human cells as detected by the 14,15-DHET ELISA were indicative of the neoplastic and metastatic phenotype of carcinoma cells.  This 14,15 DHET ELISA Assay kit can be used for the determination of 14,15-DHET in serum, plasma, cells, and tissues following proper isolation and purification.  Instructions are provided as to the proper isolation and purification in the following pages.  

This competitive ELISA kit, based on competition between 14,15-DHET epitope and 14,15-DHET-HRP conjugate for a limited number of binding sites available from the anti-14,15-DHET antibody, which is coated to the wells of the 96 well ELISA plate.  The conjugate concentration is held as a constant in each well, while the concentration of the 14,15-DHET is variable, based on the concentration of the sample or standard.  Thus the amount of the 14,15-DHET conjugate which is able to bind to each of the wells is inversely proportional to the concentration of 14,15-DHET in the standard or sample.  The amount of the conjugate which is bound to each well is then determined by the amount of color obtained when TMB is added.  The TMB reacts with the HRP available in the well.  With the addition of sulfuric acid, the blue colored product is converted into a yellow colored product, which can be read on a plate reader at 450 nm. 

Assay Procedure

  1. Load 200 microliters of Sample Dilution Buffer into the blank (BL) wells and 100 microliters of Sample Dilution Buffer into the maximum binding (BO) wells.
  2. Load 100 microliters of each of the standards into the appropriate wells.
  3. Load 100 microliters of each of the samples into the appropriate wells.
  4. Load 100 microliters of the diluted 14,15-DHET-HRP conjugate in the BO wells, the standard wells, and the sample wells.  Do NOT add HRP conjugate into the BL wells.
  5. Incubate the plate at room temperature for two hours.
  6. Wash the plate three times with 400 microliters of the diluted Wash Buffer per well.
  7. After the last of the three wash cycles pat the plate dry onto some paper toweling.
  8. Add 200 microliters of the TMB substrate to all of the wells (including BL wells).
  9. Incubate the plate at room temperature for 15-30 minutes.
  10. Add 50 micoliters of 2 N sulfuric acid to all of the wells.
  11. Read the plate at 450 nm.

Typical Standard Curve

Manual

Product Manual


Publications

References

1.    Cancer Res. 2005; 65:4707-15.
2.    Circulation. 2004; 110:2132
3.    Letters in Drug Design & Discovery. 2005; 2:239, etc.