Question: We are planning to use FLICA using thawed human PBMCs. Do we need to grow the PBMCs in culture to the required cell concentration before doing the FLICA assay? Also, can we add nigericin and the staurosporine to aliquots of normal PBMCs to serve as controls for the assay or do we need these controls for each patient? Do we need to have a monocytic cell line to use as the controls?
Answer: Thank you for reaching out about our FLICA Caspase Inhibitors.
You only need enough cells to run the assay. They should not require culturing to be used, culturing is mainly for growing up large enough quantities of cells to test. Culturing itself can have an impact on cells, especially immune cells.
Ideally, you would be able to do nigericin and staurosporine treatment for each patient. As each patient may have slightly different composition of immune cells in their respective buffy coat.
With that said you could use normal PBMCs, nigericin and staurosporine would certainly affect them in the same way. I know patient samples are very precious, but use of normal PBMCs may have minor differences compared to patients. It is more of an immune system composition question than a functionality of the kit, the kit can only tell you if the active caspase(s) are present.
View our FLICA kits from our Cell Viability Assays product line. If you have questions you would like answered, please email help@immunochemistry.com.
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