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How much DNA can I use when transfecting hepatocytes with the standard Nucleofector™?

We have used up to 6 µg of DNA per 100 µl reaction with no deleterious effect.

Can I order any of the Nucleofector™ Kit components separately?

No. The Nucleofector™ Kits are only available as full kits.

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.

Who is the distributor for my country?

Please click on the link provided to search for your distributor.

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 Nucleofection™....

Why do you have different Optimized Protocols for the Nucleofection™ of unstimulated and stimulated Human T cells?

We found that the stimulation of T cells not only changes the cells' function but also significantly alters the requirements for successful Nucleofection™. That's why Lonza developed two Optimized Protocols for transfection giving different...

Is it possible to use frozen primary CD34+ hematopoietic stem cells for Nucleofection™?

Yes. For cryopreserved PBMCs or enriched CD34 populations, we recommend culturing the thawed cells 1-2 hours in culture medium before Nucleofection™. Any further enrichment procedure after thawing is not recommended. Viability of frozen nucleofected...

What are the critical steps for successful Nucleofection™ of monocytic cell lines like THP-1, HL60 and U937?

To achieve optimal results, we strongly recommend following the Optimized Protocols in regard to cell culture conditions (medium, splitting cycle, density before Nucleofection),DNA amount and purity. Please also be sure not to exceed 90xg when...

Do you recommend a particular supplier for siRNA duplexes for use in Nucleofection™ ?

For siRNA applications using the Nucleofector technology we recommend siRNA duplexes from Dharmacon, part of Thermo Fischer Scientific.

Can I use the protocol for MCF7 cells for other breast cancer cell lines?

No. Often times closely related cell types will require completely different conditions for Nucleofection™. Therefore, you may want to consider performing a full optimization by using the Cell Line Optimization Kit. Please click on the related link...

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