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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 Cell Line...

What is the best way to establish the Nucleofector Technology in my lab?

We strongly recommend establishing the Nucleofector™ Technology with the positive control vector pmaxGFP™ provided in our kits. pmaxGFP encodes the green fluorescent protein (GFP) from Copepod Pontellina sp. Just like eGFP expressing cells, maxGFP™...

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...

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...

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...

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™...

Do you recommend positive selection or depletion for the purification of Human monocytes for Nucleofection?

We only recommend the depletion method. For Example, the RosetteSep™ Isolation Kit for human monocytes [Stem Cell Technologies, Cat. No 15028]. The advantage of depletion is that the monocytes are left untouched by antibodies during the process.
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