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How long does it take to obtain stable transfectants?

Depending on individual cell type and doubling rate, selection of stable transfectants will take between 7 and 28 days. Expansion and characterization of single cell clones will take several weeks in addition.

What is the best way to centrifuge Human Hepatocytes prior to Nucleofection™?

In order to avoid cell compaction in the pellet and difficulties in cell resuspension, we recommend using round bottom 2 ml vials or 50 ml BD Falcon™ tubes for all centrifugation steps prior to Nucleofection™.

I have a very high transfection efficiency but most of my cells die during selection, is this to be expected?

This is the normal pattern you should expect to see. Only a small proportion (1/10,000 to 1/100) of all transfected cells will integrate the transfected DNA into their genome and become stable transfectants. The remaining cells lose the transfected...

When can I start induction studies on Human Hepatocytes transfected with the Nucleofector™?

We recommend allowing the transfected hepatocytes to recover in Plating medium for 72 hours post-Nucleofection™. For induction studies (e.g. CYP3A4), the Plating medium is replaced with maintenance medium (William's E without FCS and EGF) and...

What species of rats and mice are your Clonetics™ embryonic neuronal cells isolated from?

The rats are Sprague-Dawley and the mice are CD1 or C57.

What is the lowest amount of Human Hepatocytes that can be used with the 96-well Shuttle™ System or the 4D-Nucleofector™ X-Unit?

Human hepatocytes do not like to be cultured and transfected at low densities. Therefore, using less than 100,000 cells per Nucleofection™ is not recommended.

Do the Clonetics™ rat/mouse embryonic neuronal cells proliferate?

No, the neurons do not proliferate.

What kind of effects could a mycoplasma contamination have on my cells?

"Mycoplasma contamination can: > Inhibit cell metabolism > Cause cell death > Induce morphological alterations > Alter DNA, RNA, and...

What passage number are the Clonetics™ mouse/rat embryonic neuronal cells cryopreserved at?

The cells are cryopreserved at passage 0.

How can I detect mycoplasma contamination of my cells?

We recommend using the 20-minute MycoAlert™ Mycoplasma Detection Kit from Lonza. The simple four-step MycoAlert™ Assay is a selective biochemical test that exploits the activity of mycoplasmal enzymes. The presence of these enzymes provides a rapid...

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