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Using Improved Camera Technology to Increase Digital Microscopy ThroughputDownload
Related Products: Cytation 5
September 23, 2019
As many as a third of all putative drugs fail from toxicity issues which contributes to the high cost of pharmaceutical R&D, particularly if toxicity is proven in clinical trials. The extent of cytotoxicity can be assessed by either the measurement of cellular proliferation or cytotoxicity. While quantitative absorbance or fluorescence based assays were originally employed, the use of high content screening with live cells is becoming more popular due to the wealth of information garnished from this approach. The use of higher magnification objectives often limits the field of view that can be analyzed. In order to provide adequate statistical cellular analysis, multiple overlapping images are captured and electronically stitched together prior to analysis. This process adds considerable time to the image capture routine, resulting in decreased throughput. In order to overcome this bottleneck, BioTek has incorporated a wide field of view (WFOV) camera that captures the entire well of a 384-well microplate in a single image using a 4x objective.
To demonstrate the value of this camera system, we have evaluated the effects of oridonin, methotrexate, tamoxifen, 5’-azocytidine, and mevinolin on HT1080 cells using the fluorogenic reagent CellTox™ Green to quantitate cytotoxicity. In addition, several other cytotoxic compounds were used to demonstrate apoptosis, necrosis and cell proliferation. Using pSIVA-IANBD, apoptotic cells were quantitated after exposure to staurosporine, camptothecin, cyclohexamide, and nocodazole. Necrosis was measured concurrently with propidium iodide, which will only stain cells with compromised cellular membranes, while proliferation was measured using label-free high contrast brightfield cell counting.
Many of these compounds are known inhibitors of cell growth through a number of different mechanisms. For example, oridonin has been shown to induce potent growth inhibition though cell cycle arrest at the G2/M phase, while mevinolin causes cell cycle arrest in G1 though the inhibition of steroid synthesis and nocodazole causes cycle arrest in G2 through microtubule destabilization. Camptothecin inhibits DNA replication by preventing ligation and staurosporine is a protein kinase inhibitor, known to induce apoptosis in numerous cell lines.