Predictive Residual Response assessment using HALO®
Predicting Cellular Residual Response
and Change in Sensitivity of Primitive
Stem Cells after Drug Administration In
Vitro using HALO®
The HALO® Predictive Residual Response Contract Service and Assay Kit uses the primitive high proliferative potential stem and progenitor cell (HPP-SP) population to detect the proportion of cells remaining after an initial insult and any change in sensitivity (IC50) of the cells to the agent.
The HPP-SP cell population is, under normal steady state conditions, quiescent; that is, very few cells are in cell cycle. The HALO® Predictive Residual Response (PRR) protocol is a 2-stage culture system that takes between 10 and 14 days to complete depending on which HALO® Platform is used.
During the first stage or primary culture, the HPP-SP cell population is “primed” out of quiescence and into cell cycle using a combination of growth factors and cytokines in the presence of the test compound. Sufficient replicate wells are setup to allow for multiple parameters to be tested if required. After an initial 5-7 day incubation period, a proportion of the replicate wells are used to determine the proliferative status of the cells by luminescence. The cells in the remaining wells are then removed and pooled. The cells are counted and phenotypic analysis can be performed on the cells if sufficient are available. The remaining cells are then re-plated into a second, 96-well plate culture in the absence of the agent, but in the presence of a combination of growth factors and cytokines that results in full expansion of the "primed" cells. After the secondary re-plating, a proportion of the replicate wells are used to determine the proliferative status of the expanded cells. Any remaining wells can be used to remove the cells and perform phenotypic analysis to determine the percentages and types of cells that have been produced. If sufficient cells are available, gene expression analysis can also be performed.
The dose response curves obtained after primary and secondary culture using “priming” and “full stimulation” conditions provides information on how the HPP-SP population responded to the agent. The proportion of cells remaining and whether these cells exhibit an increased sensitivity to the agent would imply damage to the remaining cells at lower agent doses if multiple doses were administered.
The HALO® Predictive Residual Response Kit can be used as a separate kit, but can also be combined with the HALO® 7-Population Predictive Hemotoxicity Kit.
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