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Can I use larger cuvettes for my Nucleofection Reaction ? Can I use the cuvettes more than once?

No. The electrical parameters provided by the Nucleofector™ Device are optimized for the cuvettes contained in the Nucleofector™ Kits. The cuvettes are single-use only. Using the cuvettes more than once will result in higher cell death and lower...

If I use Miltenyi paramagnetic beads for positive selection of cells, i.e. beads remain on the cell surface, does that influence Nucleofection® results?

No, it seems that the beads bound to the cell surface do not have any influence on Nucleofection®. Larger magnetic beads (i.e. > 1µm) may disturb the Nucleofection® process and negatively influence cell viability and efficiency.

What is the proof that DNA enters the nucleus during Nucleofection?

There are two facts that prove that the DNA is directly transported into the nucleus: 1.) Using the Nucleofector™ Technology, transfection of non-dividing cells is possible. With conventional non-viral transfection methods cell division is a...

How can I remove the cells from the 100 uL Nucleocuvette® Vessel after Nucleofection®? Is there any alternative?

We recommend using the plastic single use pipettes provided in the kits. These pipettes prevent any damage to the cells and ensure a good cell recovery.

For hepatocytes, is plating density a concern post Nucleofection®?

Yes, the optimized protocols for the standard Nucleofector® recommend the use of 6 well plates. Plating into a 12-well plate will pose a problem as less cells will attach at high densities.

Four hours after Nucleofection®, I can see the hepatocytes have attached to the well, but the morphology does not look correct, should I be concerned?

There is no reason for alarm. The hepatocytes may not exhibit normal morphology a few hours after Nucleofection® but by 24 hours post Nucleofection®, morphology should be normal. Remember to perform a fluid change about 4 hours post...

What antibodies can be used against the GFP produced by pmaxGFP and pmaxFP-Green vectors? What antibodies can be used to detect denatured GFP? What antibodies can be used to detect the RFP expressed by pmaxFP-Red?

Most of the commercial antibodies for GFP do not detect GFP expressed from pmaxGFP or pmaxFP-Green vectors. The origin of the protein expressed by the pmaxGFP control plasmid and pmaxFP-Green is from the green fluorescent protein CopGFP, cloned from...

Is there a difference between, for example, the programs C-20 and C-020?

No, the programs C-20 and C-020 are identical regarding their electrical parameters. The letter minus a two-digit number is the nomenclature of the Nucleofector™ I. Since we have launched the Nucleofector™ II we changed it to a letter minus a three...

Is it more difficult to detect DsRed by FACS than GFP?

Yes, DsRed is much less bright than GFP and is more difficult to detect by FACS and microscopy because it bleaches quickly.

Why do you recommend 7AAD instead of PI staining for cell determination in mouse macrophages?

The results obtained using 7AAD staining are much clearer than those obtained using propidium iodide (PI).  Using flow cytrometry stained and unstained, transfected and non-transfected populations can be separated much more easily.
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