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f the same 96 well plate. Cells were grown for 24 hours, media was removed, and cells were washed once with 200 l of 1X PBS. 200 l of the Cells-to-Signal Lysis Buffer were added to the cells, and the plates were incubated for 5 min at room temperature. 3 l of lysate were used in a 10 l one-step RT-PCR using TaqMan Gene Expression Assay for GAPDH or VEG-F. The two amplification curves on the left are for GAPDH, and the curves on the right are for VEG-F. The amplification curves in red and dark green were generated using HeLa cell lysate. The amplification curves in purple and light green were generated using lysate prepared from MCF-7 cells.


SYBR Green Detection
Real-time detection of PCR products using SYBR Green is a commonly used method to quantitate gene expression by RT-PCR. Though SYBR Green detection requires more optimization than other real-time PCR detection
techniques, it is economical and can be carried out on virtually any real-time detection platform. The Cells-to-Signal Kit is compatible with SYBR Green detection used with two-step RT-PCR.


Long-term Stability
When working with multiple cell samples, it is not always feasible to simultaneously process them all. Likewise, cell lysates may be needed at a later stage for repeating prior experiments or for quantitating the expression of different genes from the same sample. Lysates made with the Cells-to-Signal Kit can be safely stored at -20C for two months or longer without RNA degradation (Figure 5) .

Figure 5. Stability of Cells-to-Signal Lysate
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