Madison, WI is home for Promega, and while it is not a huge city, Madison is home to many biotech companies, fed mostly by the local, world-class University of Wisconsin-Madison. Many scientists and scientist families work, live and play near one another here. It is not uncommon for two scientists from different companies to talk to one another and discover that their respective companies have products or processes that could benefit both companies.
Case in point: Scientists at Promega have a good working relationship with Cellular Dynamics International (CDI), a biotech firm that specializes in differentiated iPSC-derived cells. We want to demonstrate that our assays work in iPSC cells and CDI wants to demonstrate the range of assays that can be performed with their iPSC-derived cells.
Differentiated iPSC cells are as close to primary cells as you can get, and primary cells are notoriously difficult to transfect due to their slow rate of growth and increased propensity for death. CDI reported great success with ViaFect™ Reagent and generously shared their data with us (see image). Continue reading “ViaFect™ Reagent for Transfection of iPSC-derived Cell Lines”
Not every lab has a tried and true transfection protocol that can be used by all lab members. Few researchers will use the same cell type and same construct to generate data. Many times, a scientist may need to transfect different constructs or even different molecules (e.g., short-interfering RNA [siRNA]) into the same cell line, or test a single construct in different cultured cell lines. One construct could be easily transfected into several different cell lines or a transfection protocol may work for several different constructs. However, some cells like primary cells can be difficult to transfect and some nucleic acids will need to be optimized for successful transfection. Here are some tips that may help you improve your transfection success.
Transfect healthy, actively dividing cells at a consistent cell density. Cells should be at a low passage number and 50–80% confluent when transfected. Using the same cell density reduces variability for replicates. Keep cells Mycoplasma-free to ensure optimal growth.
Transfect using high-quality DNA. Transfection-quality DNA is free from protein, RNA and chemical contamination with an A260/A280 ratio of 1.7–1.9. Prepare purified DNA in sterile water or TE buffer at a final concentration of 0.2–1mg/ml.
Continue reading “Improving the Success of Your Transfection”