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Investigating Novel Therapeutic Targets for Triple Negative Breast Cancer using an Automated Kinetic Proliferation Assay下載
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February 05, 2018
Authors: Xavier Amouretti and Joe Clayton, BioTek Instruments, Inc., Winooski, VT USA; Cindy Fonseca, Carolyn Marquis, Lisa Wood and Jason Stumpff, Molecular Physiology and Biophysics, University of Vermont, Burlington, VT USA
Measuring cell proliferation is a vital aspect of cancer research and therapeutic drug development. Most current methods for evaluating cell proliferation rely on end point assays, often using indirect measurements to approximate the size of a cell population based on a single predetermined time point. Imaging-based kinetic proliferation assays offer considerable advantages over these conventional approaches by capturing a detailed profile of cell population growth over time using direct cell counts or percent confluence values.
Although fluorescent label-based methods are available to determine either cell counts or percent confluence, they present substantial constraints concerning kinetic applications. The use of intercalating nuclear stains limits the maximum duration of experiments due to cytotoxic effects; while creating stable cell lines expressing a fluorescent marker is a costly and time-consuming task.
Given these limitations, label-free techniques for measuring cell population size and determining proliferation rates are preferable over methods that require fluorescent labels. BioTek’s kinetic label-free proliferation assay simultaneously determines both direct cell counts and percent confluence values using high contrast brightfield imaging. The fully automated imaging system provides complete environmental control for up to eight microplates, enabling long-term proliferation experiments for diverse applications.
Here we describe a study in which this novel technique was used to evaluate a promising class of drug targets for specifically limiting the growth of triple negative breast cancer (TNBC) cells. Five different kinesin motor proteins involved in regulating mitotic spindle integrity were inhibited in three TNBC subtypes (basal-like 1, basal-like 2, and mesenchymal). The versatility of this assay enabled accurate evaluation of cell proliferation rates for each cell type across a range of cell densities and conditions. Our findings indicate that inhibition of the kinesin Kif18A significantly slows TNBC proliferation but not normal cells, supporting the development of Kif18A-targeted therapeutics for TNBC.