Cortisol Assay
Cortisol, also known as hydrocortisone, is the major glucocorticoid produced and secreted by the adrenal cortex. It is present in blood as free protein or bound to cortisol binding protein (CBG) and is involved in regulation of glucose metabolism, fat degradation, blood pressure levels and stress inhibition. Cortisol also diffuses into cells, where its level is highly regulated by 11β-hydroxysteroid dehydrogenase (type 1 or 2). Cisbio Bioassays's HTRF® cortisol assay allows for rapid and accurate cortisol measurement even in complex samples such as liver microsomes, whole cells and animal serum. This assay can also be used to assess 11β-HSD1 activity (native, recombinant or microsomal).
- Detection in complex samples such as serum and whole cells
- Low cross-reactivity with cortisone
- Signal to background ratio >25
- Highly sensitive assay with large dynamic range
- Adapted to 96, 384 and 1536-well formats
- Amenable to automation and HTS
- Cortisol measurement
- 11β-HSD1 screening
Assay Theory:
The cortisol assay is a monoclonal antibody-based competitive assay requiring a single 2 hour incubation following cell stimulation. The assay can also be run with purified 11β-HSD1 enzyme.
Assay Principle:
The basic assay is run in two steps:
Stimulation step:
cells (or microsomes) are stimulated with compounds
Detection step:
cortisol levels are measured by successive addition of d2-labeled cortisol and a
Cryptate-labeled anti-cortisol MAb. Fluorescence is read after a 2 hour incubation.
Product Specifications:
| Sample compatibility: | serum, whole cells, & liver microsomes |
| Detection limit: | 70 pg/mL (2 fmol cortisol/well) |
| Dynamic range: | 0.1 to 100 ng/mL |
| S/B: | >30 |
| Z' factors: | as high as 0.9 |
| Cross-reactivity: | Cortisone 0.9% Corticosterone 7.0% Prednisolone 5.7% Cortisone 0.9% Progesterone 2.7% Prednisone < 0.3% Testosterone < 0.002% DHEA < 0.02% Dexamethasone < 0.0007% |
Figure 1: A typical standard curve is shown. The assay was performed in a final 20µl assay volume in a low volume 384-well plate
Assay Performance:
Cisbio Bioassays's cortisol assay is based on our robust and patented HTRF technology. One benefit that arises from the robustness of HTRF is the ability to use samples such as serum, cells and microsome preparations. Figure 2 summarizes the results of a microsomal 11β-HSD1 assay performed using HTRF cortisol reagents in low-volume 384-well plates. Known inhibitors of 11β-HSD1, Carbenoxolone and Glycyrrhetinic acid, blocked the conversion of cortisone into cortisol. IC50 values were 0.33 and 0.31µM respectively.
Figure 2: Microsomal 11β-HSD1 (0.2mg/mL) was stimulated by NADPH (200µM) and cortisone (160nM) in a 384 low-volume plate. Various concentrations of the Glycyrrhetinic acid and Carbenoxolene inhibitors were added. A standard curve with known cortisol concentrations was run concomitantly. Plates were incubated for 2 hours at 37ºC and then cortisol-d2 conjugate and Eu3+ Cryptate labeled anti-cortisol antibody added.
Ordering Info:
Cortisol Assay Reagents:
| Description | Quantity* | Cat no. | Product Insert | ||
| Cortisol | 1,000 tests | 62CO2PEB | |||
| Cortisol bulk | 20,000 tests | 62CO2PEC | |||
| *based on 20µl assay volume | |||||
Companion Products:
| Description | Cat no. |
| Cortisol calibrator | 62CORCDA |
| Cortisol control | 62CORTDA |
| Diluent for standard curve prep. | 62DL1DDD |
For more information or To Place an Order:
U.S. and Canada
1-888-963-4567
Europe and other countries
+33(0)466 796 705
Japan
+81-3-5510-2932
