Growth Hormone Receptor (GHR)
|Product Family||Product Number||Product Description||Technical Manual|
|IB14001-32||Human GHR Reporter Assay System, 3 x 32 assays in 96-well format||Technical Manual|
|IB14001||Human GHR Reporter Assay System, 1 x 96-well format assays||Technical Manual|
|IB14002||Human GHR Reporter Assay System, 1 x 384-well format assays|
Human Growth Hormone Receptor Assay Kit
This Human Growth Hormone Receptor (GHR) assay kit is an all-inclusive firefly luciferase reporter assay system that includes in addition the the GHR Reporter Cells, two optimized media for use in recovering the cryopreserved cells and for diluting test samples. Also included is the reference agonist GH, Luciferase Detection Reagents, and a cell culture-ready assay plate.
Reporter Cells are transiently transfected and prepared as frozen stocks using INDIGO's proprietary CryoMite™ process. This cryo-preservation method allows for the immediate dispensing of healthy, division-competent reporter cells into assay plates. There is no need for cumbersome intermediate treatment steps such as spin-and-rinse of cells, viability determinations, or cell titer adjustments prior to assay setup.
INDIGO’s Growth Hormone Receptor assay kits feature a luciferase detection reagent specially formulated to provide stable light emission between 5 and 90+ minutes after initiating the luciferase reaction. Incorporating a 5-minute reaction-rest period ensures that light emission profiles attain maximal stability, thereby allowing assay plates to be processed in batch. By doing so, the signal output from all sample wells, from one plate to the next, may be directly compared within an experimental set.
Growth Hormone Receptor Assay Kits are offered in different assay formats to accommodate researchers' needs: 3x 32, 1x 96, and 1x 384 assay formats for screening small numbers of test compounds, as well as custom bulk reagents for HTS applications.
Bulk assay reagents can be custom manufactured to accommodate any scale of HTS. Please inquire.
Human Growth Hormone Receptor Assay Services
The primary application of INDIGO's cell-based GHR assays are to quantitatively assess the bioactivity of a test compound as an agonist or antagonist of the receptor. Service assays include a positive control reference compound and 'vehicle' control for every experiment. A formal study report and all data files are provided to the client upon completion of the study. To receive a quote for your proposed study, complete & submit the online "Request a Quote" form or contact an INDIGO Customer Service Representative to discuss your desired study parameters.
Human Growth Hormone Receptor Assay Background
The Growth Hormone Receptor (GHR) is a single-pass transmembrane receptor that functions as a homo-dimer. Growth Hormone (GH, aka Somatotropin) activates GHR to initiate signal transduction by a variety of pathways, including Ras/ERK, PI3K/Akt, and JAK2/STAT1,3,5,6. The activation of these various pathways may culminate in the activation of cytosolic targets, or in the activation of specific transcription factors and the induction of their respective target genes.
JAK2 dependent phosphorylation and activation of the transcription factor STAT5 is a prominent outcome of GHR activation, and it is the signaling pathway exploited by the reporter cells in this assay. Specifically, INDIGO's Reporter Cells contain the luciferase reporter gene functionally linked to an engineered minimal promoter sequence with upstream tandem STAT5 genetic response element (GRE) sequences. Growth hormone activates the GHR in a dose-dependent manner, thereby triggering the JAK2/STAT signal transduction cascade. Activated STAT5 binds to its consensus GREs to initiate the formation of a complete transcription complex that drives expression of the Luc reporter gene. Therefore, quantifying changes in luciferase activity from peptide-, drug-, or antibody-treated reporter cells relative to that of "untreated" cells provides a sensitive, dose-dependent surrogate measure of changes in the activity of GHR.
GH/GHR play critical roles in the physiological processes of early bone and muscle development, and the regulation of lipid and carbohydrate metabolism. Also important is the role of GH-dependent activation of GHR in modulating the production and secretion of insulin-like growth factor-1 (IGF-1) within the liver, known as the GH-IGF1 axis. Dysregulation of GH production, either up or down, has significant physiological consequences. The development of body mass and size, insulin sensitivity, oncogenesis, and the onset of obesity, liver disease, and age-related diseases are greatly impacted by a chronic imbalance of GH production and/or function.
Not surprisingly, the development of novel GH-like peptide therapeutics, as well as small molecule and antibody modulators of Growth Hormone Receptor activity, continues to command much interest. Accordingly, the primary application of this assay is to screen test materials for any functional activity, either agonist or antagonist, that they may exert against the human GHR.
This assay utilizes proprietary human cells that provide constitutive expression of the Human Growth Hormone Receptor, isoform 1 (GHR).
Synonyms: Growth Hormone Receptor, GHR, Human Growth Hormone Receptor isoform 1
Human Growth Hormone Receptor Assay Data
GHR Activation assay. Dose-response analyses were performed according to the protocol provided in the assay Technical Manual. 200 µl / well of GHR Reporter Cell suspension was dispensed into the 96-well assay plate, which was then incubated for 4 hours. The concentrated stock of the peptide GH (provided) was further diluted using CSM to produce treatment media at the desired assay concentrations. The pre-culture media were discarded from the assay wells and 200 µl per well of the prepared treatment media were dispensed (n = 3/conc.), including ‘untreated’ control wells. ‘Mock’ reporter cells, which contain the STAT5-Luc reporter gene, but lack expression of GHR, were similarly treated with GH. Following a 22 hr incubation period treatment media were discarded, Luciferase Detection Reagent was added, and luminescence intensity per well was quantified. Values of average relative light units (RLU) and corresponding values of standard deviation (SD), percent coefficient of variation (%CV), Fold-Activation and Z’ were determined for each treatment concentration. Non-linear regression analyses of Fold Activation vs. Log10[ng/mL] and EC50 determination was performed using GraphPad Prism software.