Cultivating Cell Lines for Assay Development & Drug Discovery

At Aptuit AG & Evotec Pharmaceuticals

Precise cell counting is critical for reproducibility in cellular assays. At Aptuit (Switzerland) AG, we are cultivating a variety of different cell lines which are used for assay development and high-throughput screening for hit identification in drug discovery projects.  

In that regard, reproducible cell counts are crucial to obtain comparable data across different batches and days in longer screening campaigns. In addition to cell number, assessment of cell viability is particularly important for functional assays with TR-FRET and FLIPR readouts performed at Aptuit. For these, and we rely heavily on the NucleoCounter® NC-200 for robust and reliable cell counting at moderate cost requiring little hands-on time. 

Results

Cell count parameters of the adherent cell lines CHO, 1321N1 and PC3 and the suspension cell line THP-1 were analyzed. Cell monolayers of adherent cells were detached using trypsin, resuspended in full growth medium and suspensions measured immediately. Suspension cells THP-1 were not further treated before measurement.  

All cell suspensions were loaded into a Via1-Cassette and analyzed on the NC-200 for count of viable cells (A), percent viability (B), cell diameter (C) and percent of aggregates (D).  

No cell aggregates were detected in the suspension cell line THP-1. Data in all graphs is the average of three measurements; error bars represent standard deviation from the mean. The percent coefficient of variation (CV) of the replicates for live cell count and viability is displayed in the poster. 

Conclusion

Counting cells using the NucleoCounter® NC-200™ is a fast and reliable method to determine cell numbers and viability with low running cost and little maintenance effort.

Reproducible cell counts with a low CV were obtained for all cell suspensions covering a wide range of cell concentrations and with fast throughput (<1 minute per measurement). No additives other than the Via1-Cassette™ were used for counting.

The standard protocol for the Cell Count an Viability Assay was used for the different adherent and suspension cells, even for the slightly clumpy PC3 cells. As cell aggregation never exceeded 20%, there was no need to switch to the Aggregated Cell Count Assay protocol. For THP-1 cells, a new protocol was defined by enlarging the counting gate to ensure all cells were counted correctly.

Authors

Wassim Eid, Livia Knörr and Laura Pfeiffer.

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