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What kind of cuvettes can one use for the transformation of bacteria using the Nucleofector® I/II/2b?

We recommend using cuvettes with the 0.1 cm gap width.

Why does Lonza recommend using non-ventilated caps when culturing insect cells?

The reason is that insect cells are often incubated in very simple incubators that are not humidified (and without CO2 at 24°C to 28°C). Insect media typically do not need CO2 for buffering pH, so gas exchange is not required. To avoid evaporation of...

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...

What is the ratio of Supplement to Nucleofector® Solution?

The ratio is 1:4,5 (500 µL of supplement is added to 2.25 mL of Nucleofector® Solution).For a single 100 µL reaction, use 18 µL of supplement plus 82 µL of solution to make the 100 µL.

Does the GFP sequence in the pmaxGFP™ control plasmid has a stop codon?

Yes. The coding sequence ends with a stop codon (TGA/UGA).

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...

Can I apply more than one program (transfection condition) per plate with the 96-well Shuttle® System?

With the 96-well Shuttle® system, it is possible to apply up to 96 different programs (transfection conditions) per 96-well Nucleocuvette® Plate. Therefore, optimization of Nucleofection® Conditions for a particular cell line can be...

What cell number do I need per well when using the 96-well Shuttle® Device or the 4D-Nucleocuvette® Stripes?

The cell number is very much dependent on the cell type you use. For high throughput Nucleofection®, we usually start with one fifth of the optimal amount recommended for the standard Nucleofection® in the 100µl cuvette. Currently the absolute...

What is the amount of DNA needed per well when using the 96-well Shuttle® Device?

The amount depends on the cell type and on the DNA construct. As a rule of thumb 400 ng of DNA per well should be applied. However, for some cells or constructs even 100 ng may be sufficient.

What is the amount of siRNA needed per Nucleofection® Reaction?

When performing siRNA-mediated knockdown experiments it is advisable to conduct a dose-response (concentration) analysis to determine the minimum siRNA concentration necessary for sufficient target knockdown on the mRNA, protein, or functional level....

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