GPCR

High Performance GPCR Assays in Less Time

 

Reduce assay development time to days by eliminating the need for stable cell lines. Easily overexpress GPCRs of interest or fusion proteins, artificially couple receptors to specific G alpha subunits or knock out signaling components in your cells of choice. Cryopreserve cells post transfection to create an assay ready cell bank without sacrificing performance.

  • Eliminates the need for stable cell lines
  • >90% transfection efficiency and >90% cell viability for most cell types
  • Use physiologically relevant cell types including primary or stem cells
  • Computer-controlled for reduced day-to-day and user-to-user assay variability
  • Rapid assay development
  • Fully scalable, able to support assay development through large screening campaigns

APPLICATIONS INCLUDE:

Functional, cell-based GPCR assays

Second messenger assays including cAMP, IP3, & FLIPR®

Binding assays

High throughput screening

High content screening

 

 

Develop GPCR Assays in a Day

MaxCyte Transfection systems use a single buffer for all cell types and come pre-loaded with cell-type specific electroporation protocols. All you need to do to develop your assay is perform a DNA titration experiment. By altering the concentration of the target expression plasmid used for transfection, researchers can easily control target expression levels and fine tune assay sensitivity.

A). HEK 293H cells were transfected with increasing concentrations of M1 muscarinic receptor plasmid DNA (0.5 to 2.0 mg/1×106 cells) and treated with 0.1µM carbachol 24 hours post electroporation. FLIPR® assays were run using the Fluo-8 Calcium Assay Kit. The magnitude of calcium flux (relative light units) directly correlated with M1 [DNA].  B). Untransfected or transfected cells (2 µg/1×106 cells) were treated with increasing concentrations of carbachol and FLIPR assays performed (n=8).  Transfected cells demonstrated a dose-dependent response upon carbachol activation. The EC50 value of transfected cells (0.12µM) is considerably lower than untransfected cells (7.6µM), indicating a clear distinction between M1 muscarinic receptor-specific and endogenous background responses to carbachol.  Data courtesy of Charles River Laboratories (formerly ChanTest Corporation).

Transfect in Bulk for Assay Ready Cell Banks or Screening Campaigns

High content and high throughput campaigns require a large number of cells to perform a single screen. Other transfection technologies require multiple small-scale transfections, re-optimization of transfection protocols, and/or bulk usage of costly transfection agents. MaxCyte Flow Electroporation™ technology easily scales from assay development to full library screening and creation of assay ready cell banks.  Migration from small-scale to bulk transfection is seamless, requiring no further optimization.

HEK 293F suspension cells were transfected with a GPCR expression plasmid using either small- or large-scale electroporation. Increasing numbers of cells/well were plated in 96-well plates and GPCR activity was assayed using a cAMP ELISA at 18 hours post electroporation. Small- and large-scale transfected cells produced nearly identical results at all cell concentrations.

Reduce Your Reliance on Stable Cell Lines Using Reproducible Transient Transfection

By using MaxCyte-driven transient GPCR expression, researchers can reduce the time and/cost of securing stable cell lines, move to screen faster, and perform more reliable assays by avoiding the drift in GPCR expression levels of stable cell lines

CHO K1 cells were transfected via two independent large-scale electroporation runs with DNA encoding a β2 adrenergic receptor:eGFP fusion protein.  Transiently transfected cells and stable CHO cells expressing the non-modified β2A receptor were stimulated with isoproterenol and functional responses assessed using a commercially available cAMP kit. Results from transiently transfected cells were nearly identical to those of a reference stable cell line over expressing non-GFP linked β2 adrenergic receptor.

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