May 22, 2023
Automation, live-cell imaging, and endpoint cell viability for 96-well plate drug screens
Peer-reviewed method
- Rolando DZ Lyles1,2,3,
- M Julia Martinez1,4,
- Benjamin Sherman1,4,
- Kerry Burnstein1,4,3
- 1University of Miami Miller School of Medicine;
- 2Sheila and David Fuente Graduate Program in Cancer Biology;
- 3Sylvester Comprehensive Cancer Center;
- 4Department of Molecular & Cellular Pharmacology
Collection Citation: Rolando DZ Lyles, M Julia Martinez, Benjamin Sherman, Kerry Burnstein 2023. Automation, live-cell imaging, and endpoint cell viability for 96-well plate drug screens. protocols.io https://dx.doi.org/10.17504/protocols.io.x54v9dojzg3e/v1
Manuscript citation:
Lyles RDZ, Martinez MJ, Sherman B, Schürer S, Burnstein KL (2023) Automation, live-cell imaging, and endpoint cell viability for prostate cancer drug screens. PLoS ONE 18(10): e0287126. https://doi.org/10.1371/journal.pone.0287126
License: This is an open access collection distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this collection and it's working
Created: March 01, 2023
Last Modified: June 02, 2023
Collection Integer ID: 77846
Keywords: live-cell imaging, endpoint cell viability, 96-well plate drug screens
Abstract
To streamline the identification of potentially active cancer therapeutics, here we describe a highly adaptable semi-automated approach to screen compounds simultaneously across a panel of cell lines. These protocols leverage automation to enhance robustness, reproducibility, and throughput while integrating the IncuCyte ZOOM live-cell imaging platform and the CellTiter-Glo endpoint viability assay to assess drug efficacy. The integration of both evaluative methods strategically bypasses the shortcomings of each approach individually allowing for more thorough and detailed analysis within a single drug screen. The expected output from protocol utilization includes traditional dose-response curves, IC50, area-under the curve (AUC), GR50, area-over the curve (AOC) as well as live-cell imaging to identify cell specific morphological changes, cytostatic, or cytotoxic effects of 72-hour drug treatments. This adaptable protocol can be employed across cancer model systems and represents a reproducible procedure to optimize 96-well plate cell growth conditions compatible with the integrated drug screen and simultaneously assess drug efficacy across multiple cell lines in future cancer research studies.
Attachments
S1.pdf
1.3MB
Materials
For this protocol you will need:
CellTiter-glo (Promega, Cat# G7572) Luminescence Viability Assay or comparable endpoint cell viability assay
Luminometer compatible with the Promega CellTiter-Glo assay. This protocol features the Promega GloMax (Promega, Cat# GM3500) Explorer Multimode Microplate Reader.
A liquid handler system. This protocol features the Opentrons OT-2 Robot liquid handler system.
OT-2 Required attachments:
- P300 Single-Channel GEN2 Pipette
- P300 8-Channel GEN2 Pipette
- Opentrons 96 Tip Racks 300 μL
- 12-Channel Reservoirs for Automation (USA Scientific, Cat# 1061-8150)
White opaque welled, clear bottom 96-well tissue culture compatible plates. This protocol features Greiner Bio-one Cell culture microplate, 96 well, PS, F-Bottom, μClear (Sigma Aldrich,M0437-32EA)
Sartorius IncuCyte ZOOM Live-cell analysis system or any comparable live cell imaging platform.
- Compatible IncuCyte ZOOM Live cell analysis software.
Sterile tissue culture hood
Standard 8-channel p200 multi-channel pipette
White Opaque Lab Tape
Standard 25 mL reagent reservoirs (VWR, Cat# 89094-662)
Standard tissue culture reagents
Standard 96-well plate (clear)
Standard 24-well plate (clear)
Attachments
S1.pdf
1.3MB
