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How much DNA should I use in my Nucleofection® Reaction to obtain stable clones?

...Guideline for Generation of Stable Cell Lines"....

Missing: ~~linesGuideline~~

At which time point after Nucleofection® should I start selection of stable integration?

Antibiotics for selection should be added 24-48 h after Nucleofection®.

Missing: ~~cell~~ ~~linesGuideline~~

How often should I change the medium during selection of stable clones after Nucleofection®?

The culture medium should be changed every 2-3 days during the selection process.This eliminates potentially toxic substances produced by dying cells and secondly, it keeps the concentration of the antibiotic at a constant level.

Missing: ~~linesGuideline~~

Do you have a protocol for stimulation of mouse B cells?

Yes, if you want to transfect mouse B cells, we recommend to stimulate the cells before. The information is also available in the mouse B cell Nucleofection Protocol. Our R&D has used following combination : RPMI1640 supplemented...

Missing: ~~linesGuideline~~

Does the Mouse T cell protocol work for stimulated mouse T cells?

Yes. Stimulate the cells with anti CD3 and anti CD28 antibody for 24 hours. Use the optimized protocol. Often times, this will result in higher transfection efficiency (60-70% is possible). Alternatively, you can stimulate with ConA and IL-2 for 2-3...

Missing: ~~stable~~ ~~linesGuideline~~

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

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

Missing: ~~stable~~ ~~linesGuideline~~

Where are Lonza’s PrSC cells isolated from? What cells will be in the PrSC population?

Lonza’s Prostate Stromal Cells (PrSC) are isolated from the stroma of the prostate. Lonza’s Prostate Stromal Cells (PrSC) are a mixed population of cells, however, the majority of the cells in this population are fibroblastic in origin...

Missing: ~~stable~~ ~~linesGuideline~~

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...

Missing: ~~stable~~ ~~linesGuideline~~

Do you have any tips for the culture of primary cells or cell line prior to Nucleofection?

Yes. Please see our Techincal Reference Guide which contains many helpful tips.You can find additional information in the Optimized Nucleofection Protocol for your cell type of interest.

Missing: ~~stable~~ ~~linesGuideline~~

Can Lonza’s PrEC cells be co-cultured with Lonza’s PrSC cells?

While Lonza has never attempted to co-culture Prostate Epithelial Cells (PrEC) and Prostate Stromal Cells (PrSC) together in-house, the recommendation of Lonza Research & Development would be to plate and culture the PrEC and PrSC cells together...

Missing: ~~stable~~ ~~linesGuideline~~