Mammalian Lumio™ Gateway® Vectors, with Dual Lumio™ Red and Green In-Cell Detection Kit - FAQs

Can fluorescent protein-expressing cells be fixed?

Yes, fluorescent protein-expressing cells can be fixed using 4% paraformaldehyde in PBS for 10 min followed by one quick PBS rinse and 3 x 5 min washes with 1 mL PBS.

Are the fluorescent proteins offered by Thermo Fisher Scientific (EmGFP, YFP, CFP, BFP and Cycle 3 GFP) humanized?

Yes, all of the fluorescent proteins offered by (EmGFP, YFP, CFP, BFP and Cycle 3 GFP) have been humanized for optimal mammalian expression.

Can I use E. coli containing an F' plasmid, such as Top 10F', to propagate a vector with the ccdB gene?

While the F' plasmid does contain the ccdA gene that can inhibit or reduce the toxicity of the ccdB gene product, the ccdA expression level is likely to be too low, or inhibition may not be complete, and the bacteria would still be exposed to the ccdB gene product and thus not grow. Therefore, bacterial strains containing the F' plasmid are not recommended as hosts for propagation of ccdB containing vectors.

For propagation of Gateway vectors containing ccdB, we recommend the One Shot ccdB Survival 2 T1R Competent Cells (A10460), which were specifically designed for that purpose. However, please note that these cells are not validated for propagation of other ccdB-containing vectors like the older pZErO plasmids, and in most cases they are not expected to work due to very high levels of ccdB protein expressed in those vectors.