CELL-BASED ASSAYS use live cells as the test systems to detect the
effect of test molecules. The live cells can be either primary cells isolated from
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organism or immortal cell lines grown in artificial laboratory conditions. In general,
cell-based assays are more complex and are less defined when compared with
biochemical assays. However, cell-based assays possess many unique properties that
are not shared with biochemical assays. Cell-based assays are particularly useful
with less defined assay targets or in the situation that the target protein is known
but unknown cofactors are required for the proper function of the target protein. In
cell-based assays, the target protein is in its native cellular environment with
appropriate cofactors at physiological concentrations for its proper function. Even
when assaying a known target, cell-based assay eliminates the need to develop
expression and purification procedures for the target, which can save a lot of time and
effort in assay development. Cell-based assays can distinguish between agonist and
antagonist and can study the synergic effect of two or more test compounds. Thus,
cell-based assays are indispensable parts of bioassays and they complement
biochemical assays very well.
After cells are exposed to test molecules, many intracellular changes and cells’
responses may occur if the test molecules have effects on the cells. Cell-based assay
development is a process to evaluate many of the intracellular changes and the cell
response after the cells are exposed to test molecules and then define the final
readout for the assay. The assay signals may be continuously monitored with live
cells or be detected only at the end of the assay after the cells are fixed or lysed.
The fixed cells can be examined with flow cytometry or imaging-based methods,
which will be discussed in Chapter 12. The exposed cellular components from the
lysed cells can be assayed biochemically. One or a few of the detectable signals may be
selected as the final readout based on the relevance to the biological questions, the
nature of the signal, the reliability and ease to measure the signal, and many o [Show Less]