(1019-D) Automated technique to examine the cell viability of 3D cancer spheroids using the the OT-2
Tuesday, February 6, 2024
2:00 PM – 3:00 PM EST
Location: Exhibit Halls AB
Abstract: Cancer spheroids exhibit a greater resemblance to tumors in terms of their form, complexity, phenotype, response to chemotherapeutics, and diverse characteristics, as compared to cancer cells cultivated as monolayers. The utilization of prospective compounds to study dose-response curves in cancer spheroids is a well-established methodology in drug development for therapeutic purposes. The present study has implemented a technique for assessing the cellular viability of three-dimensional cancer spheroids subsequent to therapeutic interventions. This evaluation approach employs a resazurin-based reagent known as PrestoBlue HS. The process, starting with the seeding of cells for the production of spheroids, and including the automated administration of drugs to the spheroids, has been carried out using the Opentrons OT-2. The utilization of lung carcinoma cell line A549, hepatocellular carcinoma cell line HepG2, and pancreatic cancer cell line PANC1 has been employed in the examination of 3D spheroid formation and dosage response experiments involving Gambogic acid (GA) and Doxorubicin (DR), respectively. The spheroids were cultivated on Nunclon Sphera plates with low attachment properties in order to promote the development of well-formed and compact spheroids. Subsequently, the spheroids are subjected to treatment with gambogic acid in the case of A549 cells, while doxorubicin is administered in relation to HepG2 or PANC1 cells. Dose-response research was conducted to assess the EC50 values for a particular cancer spheroid by trying a variety of drug concentrations. After an incubation period of either 96 hours or 72 hours for the individual spheroids, a microplate readout is performed to investigate the cell viability of the spheroids. This is achieved by utilizing PrestoBlue HS reagent, which exhibits fluorescence excitation/emission maxima at 560 nm/590 nm. We conducted a non-linear regression analysis to determine the variable slope of the logarithm of the inhibitor in relation to the response. This study was performed using GraphPad Prism 10 software in order to compute the EC50.The EC50 value of gambogic acid for A549 cells is determined to be 7.83971 µM during a 72-hour treatment period. Similarly, the EC50 value of doxorubicin for PANC1 cells is found to be 2.38781µM, while for HepG2 cells, the EC50 value is measured to be 2.299678 µM after a 72-hour treatment with doxorubicin. The liquid handler OT-2 is capable of achieving the highest level of efficiency in terms of precision, accuracy, and consistency of spheroid size across replicates. In summary, the OT-2 exhibits a notable level of reliability and reproducibility, along with a commendable efficiency in automating the 3D cell culture technique. Furthermore, its adaptability for high throughput screening of numerous compounds/drugs expands the potential applications of the OT-2.