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In my Nucleofection® Experiment, how critical is the time point for analysis post transfection?

It is very critical. Waiting too long for analysis post transfection can lead to loss of peak expression. The optimal time for analysis is related to two factors: stability of protein being expressed and transfection method. For a short-lived protein...

Do we sell the Nucleofector® Solution (or any other component of the Nucleofector® Kit) separately?

We don't sell the individual components of the kits separately because our product line is a kit concept line to ensure the use of certified products with quality controlled lot numbers that guarantee optimal results.

Following Nucleofection®, I cannot obtain stable clones from single cells. What could be the reason?

Some cell lines need certain cell densities to proliferate. For those cells you can try using conditioned medium for setting up limiting dilution or culturing single cells together with feeder cells.

How to maximize cell viability in a Nucleofection® experiment?

Pay careful attention to centrifugation speeds prior to Nucleofection®; be sure not to centrifuge the cells at higher than 90xg. Any trypsinization should use the minimal amount of reagent and time necessary in order to minimize stress to the cells,...

What antibiotic concentration should I use for selecting stable transfectants?

We highly recommend to perform a titration for G418 in a range of 0,1 mg/ml to 1,5 mg/ml; in serum-free culture expand range down to 20 µg/ml).Choose the concentration which is 0,1 or 0,2 mg/ml above the one which shows complete cell death as...

What is the function of retinoic acid in the BEGM® media? 

In vivo, bronchial epithelial cells differentiate along an abnormal squamous pathway under conditions of retinoid deficiency. The squamous phenotype is characterized by the induction of specific markers such as keratin 13, and the enzymes...

What are mycoplasmas?

Mycoplasmas belong to the family Mollicutes, which includes Acholeplasma, Ureaplasma and other species.However the term "Mycoplasma" is most often used as a "cover-all".More than 180 species have been identified of which 20 distinct Mycoplasma...

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

You should use the amount of DNA recommended in the optimized protocol for transient transfection. If obtaining a very high number of clones is important for your experiments, you can try increasing the amount of DNA. For further tips see our...

How often do mycoplasma contaminations occur in cell cultures?

A conservative estimate states that between 15-35% of all continuous cell cultures are contaminated with mycoplasma (Drexler HG, Uphof CC; 2002), some estimates are even higher (up to 80 % in some countries (Koshimizu K, Kotani H; 1981).

Can I use SDS as a detergent for a positive control if I do cytotoxicity assay with ToxiLight?

No. SDS has a negative influence on Adenylate Kinase (AK) activity. SDS is a cationic detergent, it is a very powerful protein denaturing agent, you will be aware of its use in SDS-PAGE for those reasons. ToxiLight™ works by measuring the activity of...
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