Bioluminescent, Nonlytic, Real-Time Cell Viability Assay and Use in Inhibitor Screening

TY - JOUR

T1 - Bioluminescent, Nonlytic, Real-Time Cell Viability Assay and Use in Inhibitor Screening

AU - Duellman, Sarah J

AU - Zhou, Wenhui

AU - Meisenheimer, Poncho

AU - Vidugiris, Gediminas

AU - Cali, James J

AU - Gautam, Prson

AU - Wennerberg, Krister

AU - Vidugiriene, Jolanta

PY - 2015/10/1

Y1 - 2015/10/1

N2 - Real-time continuous monitoring of cellular processes offers distinct advantages over traditional endpoint assays. A comprehensive representation of the changes occurring in live cells over the entire length of an experiment provides information about the biological status of the cell and informs decisions about the timing of treatments or the use of other functional endpoint assays. We describe a homogeneous, nonlytic, bioluminescent assay that measures cell viability in real time. This time-dependent measurement allowed us to monitor cell health for 72 h from the same test samples, distinguish differential cell growth, and investigate drug mechanism of action by analyzing time- and dose-dependent drug effects. The real-time measurements also allowed us to detect cell death immediately (>75% signal decrease within 15 min of digitonin addition), analyze drug potency versus efficacy, and identify cytostatic versus toxic drug effects. We screened an oncology compound library (Z' = 0.7) and identified compounds with varying activity at different time points (1.6% of the library showed activity within 3 h, whereas 35.4% showed a response by 47 h). The assay compared well with orthogonal endpoint cell viability assays and additionally provided data at multiple time points and the opportunity to multiplex assays on the same cells. To test the advantage of time-dependent measurements to direct optimal timing of downstream applications, we used the real-time cell viability assay to determine the ideal time to measure caspase activity by monitoring the onset of cell death and multiplexing a luminescent caspase activation assay on the same test samples.

AB - Real-time continuous monitoring of cellular processes offers distinct advantages over traditional endpoint assays. A comprehensive representation of the changes occurring in live cells over the entire length of an experiment provides information about the biological status of the cell and informs decisions about the timing of treatments or the use of other functional endpoint assays. We describe a homogeneous, nonlytic, bioluminescent assay that measures cell viability in real time. This time-dependent measurement allowed us to monitor cell health for 72 h from the same test samples, distinguish differential cell growth, and investigate drug mechanism of action by analyzing time- and dose-dependent drug effects. The real-time measurements also allowed us to detect cell death immediately (>75% signal decrease within 15 min of digitonin addition), analyze drug potency versus efficacy, and identify cytostatic versus toxic drug effects. We screened an oncology compound library (Z' = 0.7) and identified compounds with varying activity at different time points (1.6% of the library showed activity within 3 h, whereas 35.4% showed a response by 47 h). The assay compared well with orthogonal endpoint cell viability assays and additionally provided data at multiple time points and the opportunity to multiplex assays on the same cells. To test the advantage of time-dependent measurements to direct optimal timing of downstream applications, we used the real-time cell viability assay to determine the ideal time to measure caspase activity by monitoring the onset of cell death and multiplexing a luminescent caspase activation assay on the same test samples.

KW - Antineoplastic Agents/analysis

KW - Apoptosis/drug effects

KW - Cell Survival/drug effects

KW - Computer Systems

KW - Dose-Response Relationship, Drug

KW - Drug Evaluation, Preclinical/methods

KW - Humans

KW - K562 Cells

KW - Luminescent Measurements/methods

KW - Small Molecule Libraries/analysis

U2 - 10.1089/adt.2015.669

DO - 10.1089/adt.2015.669

M3 - Journal article

C2 - 26383544

SN - 1540-658X

VL - 13

SP - 456

EP - 465

JO - Assay and Drug Development Technologies

JF - Assay and Drug Development Technologies

IS - 8

ER -