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Is your pmaxGFP control vector supplied with the Nucleofector® Kits suitable for stable expression?

No. This vector is only supplied in our Nucleofector® Kits as a positive control and cannot be used for selection because it does not contain a mammalian selection marker. The only resistance gene expressed by this vector is kanamycin which is...

Do you have any recommended methods for detaching human monocytes from the plate for assaying or before Nucleofection®?

We have had good results by incubating the cells in ice cold PBS for 10 minutes and then rinsing the plates. Alternatively, the cells can be detached without a medium change by gently pipetting the cell suspension up and down. Our experience has also...

How do you recommend that we isolate splenic lymphocytes from mouse spleens for Nucleofection®? Is erythrocyte lysis required?

For the preparation of mouse spleen cells we recommend cutting the spleen once and passing the tissue through a 100µM cell strainer or steel mesh using a plunger. The cells are flushed into a petri dish containing PBS. In order to remove fat, cell...

What is the basic principle of the Nucleofector® Technology?

The Nucleofector® Technology is a novel transfection method especially designed for the needs of primary cells and difficult-to-transfect cell lines. It is a non-viral method based on a unique combination of electrical parameters and cell-type...

Why is the Nucleofector®Technology ideal for primary cells and difficult-to-transfect cell lines?

There are several reasons to choose the Nucleofector® Technology for your gene transfer experiments. One is the direct transfer of DNA to the cell nucleus. This makes gene expression independent from cell division. Therefore, the technique is the...

What optimization is necessary to get the Nucleofector® Technology to work?

None. For each cell type in our product list we offer a cell-type specific solution and thoroughly optimized electrical parameters. These are already pre-programmed in the Nucleofector® Device. Additionally our optimized protocols give you...

What can I do when there is no optimized protocol available for my cell of interest?

Lonza is constantly developing new optimized kits and protocols for an increasing number of primary cells and cell lines. In order to obtain the latest information, check our website or contact our Scientific Support Team.We are offering Primary...

What are the requirements for direct Nucleofection® of mRNA for protein expression?

The mRNA should be capped and polyadenylated. The conditions of Nucleofection® will be the same as for DNA with the particular cell type, i.e. follow the same protocol and use the same program, except one will likely need to add a much higher...

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

For Nucleofectio® 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...

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™...
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