Category: Assay Development and Screening
High-content imaging assays are the preferred compound phenotypic screening method because they provide a rich amount of cellular information for compound screening. Antibody and fluorescence dye staining is commonly used in high-content assays, but the multiple cell wash steps required to remove background signal are a major bottleneck for their high-throughput application. Although automated cell washers may improve the assay throughput, artifacts created by cell detachment and well-to-well variability are an inevitable disadvantage for screening. Here, we have applied the TTP LabTech Mirrorball technology to perform high-throughput imaging assays using antibodies and fluorescence dyes without washing in 96-, 384-well plates. Our assay holds several key advantages in that it is scalable from 96- to 384-well plate format, can be used in live or fixed cells, requires no washing or exchanging solutions while executing the protocol, and can be multiplexed with other fluorescent dyes. Using the Mirrorball fluorescence plate cytometer, we have performed a test screening of 1480 compounds in 384-well plate format and will report on the hit confirmation as well as follow-up results. This no-wash, add-and-read Mirrorball assay enables easy and efficient high-content screening of small molecules.
Kirill Gorshkov– Postdoctoral Fellow, NIH/NCATS, Rockville, MD
Dr. Kirill Gorshkov received his Ph.D. in 2016 from the Johns Hopkins School of Medicine Department of Pharmacology in the Laboratory of Dr. Jin Zhang. His thesis work, for which he was awarded a National Science Foundation Graduate Research Fellowship and Scheinberg Travel Award, uncovered the molecular mechanisms of cAMP/PKA signaling involved in axon outgrowth by utilizing genetically encoded FRET-based biosensors in primary embryonic rat hippocampal neurons. He has a strong background in cell signaling, neurobiology, and cell-based imaging techniques which he uses in his postdoctoral work at the NIH National Center for Advancing Translational Sciences Therapeutics for Rare and Neglected Diseases. Dr. Gorshkov chose to build upon his basic science research foundation by joining a translational research laboratory headed by Dr. Wei Zheng where he is becoming an expert in high-content imaging for drug discovery, high-throughput screening of small molecules, and lysosomal storage diseases. Dr. Gorshkov received his B.S. in Neurobiology at the University of Texas at Austin where he studied the effects of ethanol on cell signaling pathways in PC-12 cells under the guidance of Dr. Armando Salinas and Dr. Richard Morrisett. He grew up in Houston, Texas and attended Westside High School in West Houston where he began his life science research in his senior year studying how copper electrodes attached to the locust ganglion affect their longevity.