Applications Frequently Asked Questions

What are the key applications of the CellKey™ technology?

Currently, the primary use of the CellKey™ System is the pharmacological evaluation of cell surface receptor function in drug discovery applications. Applications include receptor panning; signal pathway identification and deconvolution, and standard pharmacological-type assays.

Are measurements sensitive to cell density or confluency?

Yes. There is a cell density dependency to the response. MDS Analytical Technologies recommends using a confluent cell monolayer to ensure consistency across all wells.

Can the CellKey™ System measure receptor activation responses within non-adherent cells?

Yes. The CellKey™ System can evaluate receptor activation within suspension cells. Use more cells per well to ensure sufficient contact between the cells and the electrodes at the bottom of the plate.

How important is receptor over-expression for signal detection?

The CellKey™ System can detect endogenous receptor activity. You do not need to over-express the receptor of interest. However, as in other cell-based assays, the signal detection is dependent on how tightly the receptor is coupled to its downstream pathway.

Is the CellKey™ System sensitive to partial or weak agonists?

Yes. For example, in a screen for potent and selective D2L antagonists, the selective compound, CGP 20712A, was also identified as a partial agonist.

The CellKey™ System measures the integrated cellular response. Do you typically see non-specific responses?

The treatment of cells with compounds results in changes in cell volume, cell shape, and cell-cell interaction. These changes are measured by the CellKey™ System. However, many compounds, including vehicles such as DMSO, do not elicit a CDS response. To date, the CellKey™ responses measured are proven to be receptor-specific. Pre-treatment of the cells with receptor-specific antagonists consistently blocks the measured response to the compound.

What happens if a ligand hits multiple receptors?

Compound multiplexing studies have been performed. The studies indicate that a combination of any two ligands activating the GPCR subtypes produces a unique response.

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What is the DMSO tolerance of the technology?

Studies performed so far indicate that the cells and the assay are compatible with up to 1% DMSO. If the DMSO concentration must be high, you can minimize nonspecific effects by matching the concentration in the assay buffer.

How reproducible is the data from a CellKey™ assay?

Data with %CV under 10% and Z' factor above 0.5 and often in the range of 0.75 are routine.

How is the information obtained from the CellKey™ assay different from other more traditional technologies that measure GPCRs?

The information obtained from the CellKey™ assay is the integrated cellular response to receptor activation. More traditional technologies measure discrete intracellular events, such as Ca2+ flux or cAMP fluctuation.

How do the results obtained with the CellKey™ System compare to results from other second messenger assays such as Ca flux or cAMP?

CellKey™ assay results consistently show excellent correlation with results from other second messenger assays. EC₅₀ and IC₅₀ values are comparable. The same rank order of ligand potency is maintained.

What is the measurement update rate?

The system reads all 96 wells simultaneously with an update rate of two seconds.

How long is the typical CellKey™ assay?

Typical measurement times are five to fifteen minutes.

Do I have to use specific reagents for the CellKey™ System assay?

No. Cellular dielectric spectroscopy is a label-free technology. Specific reagents, such as fluorescent dyes, tags, or markers are not required.

How does cell growth on a CellKey™ custom plate compare to growth on regular cell culture microplates?

The CellKey™ Standard 96W plate is made of biocompatible materials and is tissue culture treated. Cell growth is identical to the growth on any standard cell culture microplate.

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What is the typical workflow for a CellKey™ assay?

The typical workflow is:

1.	Seed cells on the CellKey™ Standard 96-well plate.
2.	Store the plate overnight to let the cells grow and attach.
3.	Remove the assay growth media and then add the assay buffer.
4.	Place the plate in the CellKey™ System.

The CellKey™ System measures the impedance continuously while the onboard fluidics adds the test compounds.

At what temperature can I run assays?

The onboard environmental control enables assays to be run from room temperature + 5 ºC to 37 ºC. Assays are typically run at 28 ºC.

What cell types can I use with the CellKey™ System?

You can use adherent and non-adherent cell lines and primary cells with the CellKey™ System.

Can I use attachment coatings with the CellKey™ microtiter plate?

Yes. A wide range of attachment coatings have been successfully used to enhance adherence to the cell plate. Collagen and laminin appear to be the most widely applicable.

Do I need to aspirate the growth media from the wells before I run the assay?

Assays can easily be run in media. However you should exchange the media in which adherent cells are grown overnight with fresh media (or assay buffer) prior to running the assay. Exchanging the media standardizes the buffer conductivity across all wells and improves assay reproducibility.

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