BrdU Cell Proliferation Assay

Protocol tips

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	- Cells are treated with BrdU and incubated overnight.

Protocol tips
- Cells are treated with BrdU and incubated overnight.

Publication protocol

HT22 cells were assessed for cell viability and proliferation. HT22 cells were either treated with insulin or transfected with PKCδII plasmid or PKCδII siRNA as described above. The treatments were performed in triplicate in a 48-well plate. The 5-bromo-2′-deoxyuridine (BrdU) cell proliferation assay kit was purchased from Millipore, catalogue no. 2750, and used as per manufacturer's instructions to quantitatively evaluate the number of actively proliferating cells. Briefly, 100 μl of BrdU was added per well of the 48-well plate and incubated overnight. BrdU incorporation was detected using peroxidase conjugate. The plate was read using a spectrophotometer microplate reader set at a dual wavelength of 450 nm/550 nm. The results were normalized against the blank and background readings. The cell proliferation kit was purchased from Chemicon International, catalogue no. 2210, and uses WST-1/ECS as per the manufacturer's instructions. The formazon dye produced by viable cells is quantified using a spectrophotometer set at a wavelength of 440 nm.

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Manufacturer protocol

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