No, due to their unique immune characteristics, primary T cells are sensitive to exogenous double-stranded DNA by the intracellular cGAS-STING signaling pathway, which not only causes cell cycle arrest, but also leads to cell death through the endogenous apoptotic pathway, and the cell viability and positivity rate will be significantly reduced under conventional transfection conditions. See Reference: Enhancement of the viability of T cells electroporated with DNA via osmotic dampening of the DNA-sensing cGAS–STING pathway. Nature Biomedical Engineering.
Suspension cells are cultured in suspension, allowing for a higher cell density per well compared to adherent cells. Both types of cells can be transfected during cell passaging, so adherent cells do not need to be plated before use. In addition, with numbers of experiments tested, transfect cell suspension mixture in an EP tube help maximize the transfection efficiency.
① The centrifugation protocols described in this manual are standardized at room temperature, 300g, for 5 minutes;
② After transfection, add 1 mL of complete medium and mix gently to terminate the transfection process and mitigate the potential impact of the transfection reagent on subsequent cell centrifugation (Note: The slight viscosity of the transfection reagent is typical and does not indicate an issue);
③ After centrifugation, the cell pellet may not be readily visible, especially when working with low cell numbers. This is a common occurrence and does not necessarily indicate a procedural issue.
① Optimize Cell Condition: Prior to transfection, ensure cells viability exceed 90%. This can be assessed using standard viability assays such as trypan blue exclusion or propidium iodide staining.
② Adjust Reagent Concentrations: Incrementally increase the amounts of transfection reagent and nucleic acid to identify the optimal ratio that maximizes transfection efficiency without compromising cell viability.
③ Optimize Incubation Time: Experiment with varying incubation times to determine the ideal duration for your specific cell type. Both prolonged and shortened incubation periods can influence transfection efficiency and cell viability, so it is essential to find a balance that yields the best results.
① Optimize Cell Condition: Prior to transfection, ensure cells viability exceed 90%. This can be assessed using standard viability assays such as trypan blue exclusion or propidium iodide staining.
② Adjust Reagent Concentrations: Incrementally increase the amounts of transfection reagent and nucleic acid to identify the optimal ratio that maximizes transfection efficiency without compromising cell viability.
③ Optimize Incubation Time: Experiment with varying incubation times to determine the ideal duration for your specific cell type. Both prolonged and shortened incubation periods can influence transfection efficiency and cell viability, so it is essential to find a balance that yields the best results.