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In vitro assays based on stable cell lines play a crucial role in modern drug discovery processes. When high expression of the target gene is important for assay development, GenScript's non-viral stable cell line service can deliver the stable cell pool or a single stable clone. GenScript also has the capability to test the function of the gene of interest in our in vitro assays.
GenScript offers a comprehensive range of cell line engineering, including both lentiviral and non-viral stable cell line services, and GenCRISPR™, CRISPR based knock-out or knock-in stable cell line services. We have a proven track record of success in stable cell line generation and assay development, offering clients a gene-to-assay solution for drug discovery.
In addition to offering a custom stable cell line service, GenScript also has a product catalog of over 170 GPCR & ion channel cell lines, ready to assay.
Workflow of Standard Non-viral Stable Cell Line Development
Workflow of Fast Stable Cell Line Development Process
Note: CHO-K1, CHO-K1/Gα15, CHO-K1/Gqi5, HEK293, 293T, 293EC18, 293EC18/CRE-Luc, 1321N1, RH7777, and U-2OS are available as starting cell lines. For special requirements, please feel free to contact us.
CellPower™ - lentiviral-based stable cell line service, ideal for difficult to transfect cell lines, and hard to express genes. Antibody and protein expression stable cell lines – Stable cell lines developed for bioproduction purposes GenCRISPR™ - CRISPR based knock out or knock in stable cell line service
Adenosine Receptor A1 Stable Cell Line Generation
A1 receptor is a Gαi-coupled GPCR which theoretically cannot be assayed by calcium mobilization assay. We co-expressed A1 receptor with Gα15, a promiscuous Gα protein, in CHO-K1 cells and selected a best clone as the primary clone and developed calcium mobilization assay.1. Biology of Adenosine Receptor A1
Activation of A1 receptor, a ubiquitous Gαi-coupled GPCR, elicits an inhibition of adenylate cyclase and causes decrease in cAMP concentration. Adenosine antagonists are widely used in neonatal medicine.2. Stable Clone Screening
We transfected CHO-K1 cells with the ADORA1 gene which encodes A1 receptor and got 96 primary clones by antibiotic selection. These 96 clones were further screened for the positive clones through calcium mobilization response (Fig. 1). Based on the long-lasting signal intensity profile over several months in culture, Clone 32 and Clone 7 were selected as the primary and backup clones for CHO-K1/ADORA1/Gα15 cell line, respectively (Fig. 2). All the further studies were performed on the primary clone, Clone 32.
▲ Figure 1
▲ Figure 23. 29-Passage Stability Test
We compared EC50 values of NECA, an ADORA1 agonist, at various passages of Clone 32. The variation of EC50 values of Clone 32 from passage 1 to 29 passages was within a small window (see below). Based on the pharmacology study, our results showed our Clone 32 has a high level of passage stability. The EC50 values that we have obtained matched very well with those reported in the literature (EC50 value: 20 ~ 173 nM1,2, Tab. 1).
▲ Table 1
1. Yolande Cordeaux et al. (2004) Coupling of the human A1 adenosine receptor to different heterotrimeric G proteins: evidence for agonist-specific G protein activation. Brit. J. Pharmacol., 143: 705-714.
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