Human Total Progesterone R/NR3C3 DuoSet IC ELISA Summary

Product Summary

This DuoSet IC ELISA contains the basic components required for the development of sandwich ELISAs to measure total Progesterone R / NR3C3 in cell lysates. An immobilized capture antibody specific for Progesterone R / NR3C3 binds both phosphorylated and unphosphorylated Progesterone R / NR3C3. After washing away unbound material, a biotinylated detection antibody is used to detect both phosphorylated and unphosphorylated protein, utilizing a standard Streptavidin-HRP format.

Preparation and Storage

Background: Progesterone R/NR3C3

The progesterone Receptor (PR; NR3C3) is a member of the steroid receptor superfamily which binds progesterone. The progesterone receptor is expressed in uterus, ovary, vagina, fallopian tubes and breast. Progesterone receptors play a central role in diverse reproductive events associated with the establishment and maintenance of pregnancy, alveolar development in the breast, and sexual behavior. Two isoforms of the progesterone receptor have been identified, progesterone receptor-A (PR-A) and progesterone receptor-B (PR-B), 94 and 114 kDa in size, respectively.

The B progesterone receptor (PR-B), or long form, is 933 amino acids (aa) in length. Progesterone receptor-A (PR-A) utilizes a different start site that shortens the N-terminus by 164 amino acids. The N-terminus in both progesterone receptor isoforms is rich in serine that is phosphorylated in response to hormone binding. In the absence of progesterone, few receptor molecules are phosphorylated at Serine 190 (S190). Progesterone increases this number two-fold, providing evidence for hormone stimulation. It has been shown that the two progesterone receptor isoforms can form homodimers as well as heterodimers.

Assay Procedure

Refer to the product datasheet for the complete assay procedure.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.

1、Prepare all reagents, standard dilutions, and samples as directed in the product insert.

2、Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

3、Add 50 μL of Assay Diluent to each well.

4、Add 50 μL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.

5、Aspirate each well and wash, repeating the process 4 times for a total of 5 washes.

6、Add 100 μL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.

7、Aspirate and wash 5 times.

8、Add 100 μL Substrate Solution to each well.

9、Add 100 μL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.