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Cisbio's HTRF cortisol assay performed on the PHERAstar FS

Jean-Luc Tardieu, Francois Degorce Cisbio 03/2007

This data demonstrates that with increasing cortisol concentration the anti-cortisol cryptate is displaced proportionally resulting in a decreasing signal curve.


Table 1:  EC50 values of cortisol standard curve after different incubation times.

 

InhibitorIC50 µM
Carbenoxolone1.03
Glycyrrhetinic acid1.17

 

 

Conclusion


The cortisol assay allows fast and efficient determination of cortisol in complex samples such as serum and whole cells. Screening for both active 11 beta-hydroxysteroid dehydrogenase type 1 and its inhibitors  is also simple and effective using this homogeneous assay.


The PHERAstar FS in combination with the optimized HTRF® optical module is the ideal tool to run HTRF® assays. The PHERAstar FS optical design provides  outstanding sensitivity and accuracy in fluorescence and luminescence assays; moreover, the dual simultaneous measurement minimizes the read time for assays.

Cortisol is a corticosteroid hormone present in many metabolic processes, inducing key enzymes of carbohydrate, fat and protein metabolism. Cortisol acts anti-inflammatory and as an immunosuppressor. One way to create thehormone is by reduction of cortisone by 11 beta-hydroxysteroid dehydrogenase type 1 (11 beta- HSD1).

 

The cortisol assay is a monoclonal antibody based competitive assay running in two steps. After the dehydrogenase reaction is finished (stimulation), anti-cortisol cryptate (donor) and d2 labeled cortisol (acceptor) are added to the reaction. The anti-cortisol cryptate and the d2 labeled cortisol will bind to each other leading to a high HTRF® signal. Cortisol built during the enzymatic reaction will compete with d2-labeled cortisol for the binding to the cryptate conjugate, resulting in signal loss (detection).

 

The PHERAstar® FS microplate reader in combination with the optimized HTRF® optical module provides outstanding sensitivity and accuracy. Moreover, the dual simultaneous measurement minimizes the read time for assays.

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