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Is there a special recommendation on Human T cell enrichment before Nucleofection?

For Nucleofection™ it is preferable to use Human T cell populations enriched by magnetic separation. For example, MACS™ Microbeads (Miltenyi) or by a rosetting method. For magnetic separation we recommend negative selection or detaching beads after...

Is it possible to use different substrates (e.g. siRNA duplexes) with the 96-well Shuttle Device?

Yes. Transfection can be done with any substrate AND under exactly the same conditions. For example, the same conditions (96-well Shuttle™ Solution and program) are used for siRNA transfection or shRNA vectors. This greatly facilitates the...

Are there any recommendations to avoid excessive mortality during T cell Nucleofection?

Try to keep the DNA amount for Nucleofection™ quite low (e.g. 1 µg plasmid DNA per 100 µl reaction) since higher DNA amounts might cause increasing toxicity and mortality. It is also important to use highly purified DNA, e.g. by using Endofree™...

What is the advantage of the 96-well Shuttle Device over common lipofection, especially in a high throughput framework?

Several primary cells of high research relevance can simply not be transfected at relevant efficiencies and viabilities using lipofection. The same holds true for many suspension cell lines which are often used for protein production. These difficult...

How can I increase cell viability of Dendritic cells after Nucleofection?

DNA amount and quality are very critical for Nucleofection™ of Dendritic Cells (recommended 0.5-1µg; maximal 2 µg). LPS (lipopolysaccharides) have a strong negative effect on cell viability. Please make sure that all components you use for dendritic...

What are the contents of a 96-well Shuttle Kit?

As the system will most likely be used for high-throughput approaches we provide plates which are pre-equipped with 96-well Nucleocuvette™ modules (2x8). In addition to that, the 96-well Shuttle solution, the supplement, and the pmaxGFP™ are also...

I want to stimulate Human T cells post Nucleofection. Are there any precautions to keep in mind?

After Nucleofection™, wait at least four hours before stimulation. Stimulating immediately after Nucleofection™ may lead to increased cell mortality and poor activation of cells. For more details on the stimulation of Human T cells post...

What is the advantage of the Nucleofector Technology over common electroporation (systems)?

High cell viability and high transfection efficiency with the Nucleofection™ Technology are the most prominent features. Nucleofection™, i.e., the transfer of DNA into the nucleus and not only the cytosol, is essential when working with non-dividing...

Is it possible to get neomycin resistant clones after Nucleofection with pmaxGFP?

The pmaxGFP control DNA is not suitable for making stable clones because it lacks a selectable marker for mammalian cells. The kanamycin resistance expressed by this plasmid is only suitable for bacterial selection.

Where can I find the expiration dates for the Nucleofector Kit and the Nucleofector Solution?

The expiration dates are located on the outside label of the kit box and on the label of the solution box.
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