Prostaglandin E2 Assay
Prostaglandin E2 (PGE2) is a primary product of arachidonic metabolism and is synthesized via the cyclooxygenase (COX) and prostaglandin synthase pathways. PGE2 production is a commonly used method for the detection of COX-1 and COX-2 modulation and prostaglandin synthases. The HTRF® PGE2 assay is a highly sensitive method for quantifying PGE2 either in cell supernatants or directly in the presence of whole cells. Our reagents have simplified protocols requiring only two reagent additions. These protocols are easily miniaturized and amenable to automation and HTS.
- Cell-based assay format
- Simple and rapid assay protocol
- Large assay working range
- High Z' factor
- Miniaturization to <10 µL
- No cross reactivity with PGH2 and PGF2
- PGE2 activity monitoring
- Measurement of COX and prostaglandin synthases modulation
Assay Principle:
The PGE2 assay is a competitive immunoassay that uses Eu3+-Cryptate labeled anti-PGE2. Free PGE2 from the sample competes with d2 labeled PGE2 for binding to the Cryptate conjugated anti-PGE2 antibody. The assay can be run in two easy steps and requires only a single incubation following cell stimulation. Assays can also be run with purified enzymes.
Product Specifications:
| Detection limit: | 10 pg/mL |
| Dynamic range: | 8 to 1000 pg/mL |
| EC50: | 70 pg/mL |
| S/B: | 31 |
| Z': | 0.9 for 20 µl assay volumes |
The PGE2 assay has a high level of assay flexibility. It can be performed using cell supernatants or directly on stimulated cells (see figure 1 below). The incubation time and temperature following addition of the HTRF detection reagents has little effect on the assay results providing another level of assay flexibility.
Assay Performance:
One benefit of the HTRF PGE2 assay is the ability to quantify PGE2 in the presence of cells without sacrificing assay performance. As seen in Figure 1, the results comparing direct cell-based versus supernatant transfer did not differ significantly. Monocytes were stimulated to produce PGE2 in the presence or absence of indomethacin, a known inhibitor of PGE2 production. The IC50 of indomethacin determined using HTRF technology (1.0 ± 0.4 nM), was in agreement with previously published data (Palomer A. et al. 3 ± 2 nM ; Chan C.C. et al. 7 ± 1 nM).
Figure 1: U937 cells were cultured in the presence of 10 nM phorbol 12-myristate 13-acetate (PMA) for 48 hours. Cells were then dispensed at a density of 1,105 cells/well/100 µl in the presence of 10 µg/ml lipopolysaccharides (LPS) and indomethacin at various concentrations. PGE2 detection reagents were either added directly to stimulated cells or 50 µl of supernatant transferred to a new plate and detection reagents added. Plates were incubated at RT for 4 hours after reagent additions and emissions read at 620 and 665nm.
Ordering Info:
PGE2 Assay Reagents:
| Description | Quantity* | Cat no. | Product Insert | MSDS | |
| PGE2 | 1,000 tests | 62P2APEB | - | ||
| PGE2 bulk | 20,000 tests** | 62P2APEC | |||
| *based on 20 µl assay volume ** upon request |
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Companion Products:
| Description | Cat no. |
| PGE2 calibrator | 62PG2CDA |
| PGE2 control | 62PG2TDA |
| Diluent for standard curve prep. | 62DL2DDD |
For more information or To Place an Order:
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