Multiplexed screening of compounds targeting GPCRs

High Sensitivity Ca++ Mobilization Detection

Our proprietary patented screening system (US Patent 7,858,040) enables screening of compounds against a multitude of GPCRs in live cells.
The flow cytometry assay’s sensitivity is superior to that of a plate reader due to a principal differences in the readout: flow cytometry analyzes each cell in a population and calculates percentage of cells that are responding higher than a few standard deviations from the control population. Because of this ability, assays with fluorescent signal even barely above background can show the same percent of responding cells as an assay with a very strong fluorescent signal. For example, with flow cytometric analysis one can detect just 1% of the cells responding to a stimulus with S/N ratio greater than 6. The same assay in FLIPR or other plate reader requires greater than 10% of cells to respond for a statistically meaningful signal to be detected. 

This level of sensitivity afforded by monitoring of single cell responses allows for identifying ligands that signal through unusual for a particular GPCR pathway that is not typically associated with Calcium – ligand-directed signal trafficking. Another advantage of flow cytometry sensitivity with single cell analysis is that the heterogeneous cell populations with endogenously expressed receptors can easily be analyzed.

Multiplexing

Our system allows for a unique way of screening compounds against a mixture of cell populations carrying receptors of interest as well as naïve cells and cells with counter screen receptors in one well with the same compound sample. This tremendously reduces time and material consumption while fully characterizing active ligand selectivity profile.

Each cell line is labelled with one or more fluorescent tracking dyes. All labelled cell lines are mixed together and mixture is added to wells with sample compounds. Our analysis software deconvolutes and independently analyzes each cell population based on the tracking dye signature of each cell. Responses of up to 9 different cell lines can be analyzed simultaneously in a single well with compound utilizing just a single laser with two parameters for tracking and one laser for calcium signal. Utilizing additional lasers and tracking dyes we can further increase the number of concurrent cell populations.

 

GPCR Cell Library

Saryna has developed an extensive collection of GPCR-bearing cell lines. We can design, develop, and validate multiplexed compound profiling assays by utilizing our cell lines from the list below or procuring additional cells from other vendors or through assay transfer. A partial list of readily available receptors is shown below.

Adenylate cyclase activating polypeptide 1 (pituitary) receptor type I (ADCYAP1R1)
Adrenergic receptor alpha-1A (ADRA1A), transcript variant 1
Adrenergic receptor alpha-1B (ADRA1B)
Adrenergic receptor alpha-1D (ADRA1D)
Adrenergic receptor alpha-2A (ADRA2A)
Adrenergic receptor alpha-2B (ADRA2B)
Adrenergic receptor alpha-2C (ADRA2C)
Adrenergic receptor beta-2, surface (ADRB2)
Adrenergic receptor beta-3 (ADRB3)
Calcitonin receptor (CTR)
Calcitonin receptor-like receptor (CALCRL, CRLR)
Cannabinoid receptor 1 (CB1)
Cannabinoid receptor 1 (CB2)
Cholecystokinin A receptor (CCKAR)
Cholecystokinin B receptor (CCKBR)
Corticotrophin releasing factor receptor 1 (CRFR1, CRHR1)
Corticotrophin releasing factor receptor 2 (CRFR2, CRHR2)
Dopamine receptor 2A (DR2A)
Dopamine receptor 2L (DR2L)
Dopamine receptor 2S-B (DR2S-B)
Dopamine receptor 3 (DR3)
Dopamine receptor 4 (DR4)
Dopamine receptor 5 (DR5)
Galanin receptor 1 (GALR1),
Galanin receptor 3 (GALR3)
Gastric inhibitory polypeptide receptor (GIPR)
Glucagon receptor (GCGR)
Growth hormone secretagogue receptor 1a (GHSR1a)
Histamine receptor H1 (HRH1)
Histamine receptor H2 (HRH2)
Histamine receptor H3 (HRH3)
Histamine receptor H4 (HRH4)
Melanin-concentrating hormone receptor 1 (MCHR1, GPR24)
Melanin-concentrating hormone receptor 2 (MCHR2, GPR145)
Melanocortin receptor 3 (MC3R)
Melanocortin receptor 4 (MC4R)
Melatonin receptor 1 (MT1R, MTNR1A)
Melatonin receptor 2 (MT2R, MTNR1B)
Neuropeptide Y1 receptor (NPYR1)
Neuropeptide Y2 receptor (NPYR2)
Neuropeptide Y5 receptor (NPYR5)
Parathyroiod Hormone 1 receptor (PTH1R)
Serotonin receptor 1A (5HT-1A)
Serotonin receptor 1B (5HT-1B)
Serotonin receptor 1D (5HT-1D)
Serotonin receptor 1E (5HT-1E)
Serotonin receptor 1F (5HT-1F)
Serotonin receptor 2A (5HT-2A)
Serotonin receptor 2B (5HT-2B)
Serotonin receptor 2C (5HT-2C)
Serotonin receptor 4 (5HT-4)
Serotonin receptor 5 (5HT-5)
Serotonin receptor 6 (5HT-6)
Serotonin receptor 7A (5HT-7A)
Serotonin receptor 7B (5HT-7B)
Trace amine associated receptor 1 (TAAR1)
Trace amine associated receptor 5 (TAAR5)
Trace amine associated receptor 6 (TAAR6)
Trace amine associated receptor 8 (TAAR8)
Trace amine associated receptor 9 (TAAR9)
Vasoactive intestinal peptide receptor 1 (VIPR1)
Vasoactive intestinal peptide receptor 2 (VIPR2)
Saryna Technologies LLC
6990 Carroll Road, Suite A2
San Diego, CA 92121
Phone: 858-450-0993
Fax: 858-450-0992
Email: info@saryna.com