Bio-Rad has released an instructional video that shows best practices from pre-transfection cell counting with an automated cell counter to the analysis of gene expression using real-time PCR.
The video, entitled 'Using an Automated Cell Counter to Simplify Gene Expression Studies: siRNA Knockdown of IL-4 Dependent Gene Expression in Namalwa Cells', highlights how researchers can use the TC10 automated cell counter to quickly determine the number of live cells in a sample prior to transfection, using siRNA.
The Namalwa cell line, a human lymphoma cell line that is traditionally difficult to transfect, is used in this study.
Bio-Rad scientists use this procedure in their own laboratories to study expression of single genes and gene pathways.
The video begins with a researcher using the TC10 automated cell counter to determine the number of live Namalwa cells in suspension.
Cells are mixed with trypan blue and loaded onto a counting slide that is inserted into the TC10 cell counter.
The multi-plane auto-focus feature of the TC10 cell counter ensures an accurate count and provides a quick display of live and dead cell numbers as well as an image of the cells.
In addition, the TC10 automated cell counter can automatically calculate the dilution required for downstream experiments.
In the remainder of the video, siRNA is added to the cell suspension and the mixture is electroporated using Bio-Rad's Gene Pulser MXcell electroporation system.
After incubation, the cytokine IL-4 is added to the cells to induce transcription of specific genes.
Real-time PCR is completed on RNA extracted from the cells using the CFX96 real-time PCR system to measure gene expression.
