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515 results sorted by
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The recommended cell number will vary depending on which Optimized protocol is being used. In general, using less than 2x10exp5cells per reaction causes a major increase in cell mortality. For some cell lines we have tried cell numbers up to...
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The mycoplasma specific metabolism detected by the MycoAlert® Assay is present in all members of the mollicute family (Mycoplasma, Acholeplasma, Entomoplasma and Spiroplasma) except Ureaplasma (which are no usual suspects in cell culture). 95% of all...
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We determine cell viability after Nucleofection® in two ways: 1) FACS determination of viable/dead cells by PROPIDIUM IODIDE STAINING. We normally analyze transfection efficiency in living cells by FACS: We first exclude cellular debris by...
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We routinely use plasmids of 4-7 kb in our laboratories and plasmids up to approximately 20kb should not be a problem. Using plasmids larger than this will most likely result in lower transfection efficiency. Some preliminary results we have also...
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NO - the lysis reagent in the MycoAlert® Assay is not strong enough to lyse bacteria / yeast. Also, the enzymes utilized are not very common in bacteria or yeast – but are universal among the mollicute family. Bacterial contamination is...
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The quality and the concentration of DNA used for Nucleofection® in general plays a central role for the efficiency of gene transfer. We strongly recommend using endotoxin free prepared DNA. Endotoxin free Kits are available from several...
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There is evidence to suggest that mycoplasma can survive in the absence of cells, some may even proliferate, but as a rule they are much more viable if cells are present.
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P
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?
For optimal assay conditions, reconstituted reagent and substrate should be used fresh within 2 hours after reconstitution.During this time they can be kept at room temperature.For the MycoAlert® Kit: Unused, reconstituted components can be aliquoted...
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A
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D
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The standard protocol calls for taking 2 ml of culture media, spinning down any cells and removing 100 µl of supernatant as sample.Smaller initial aliquots of media may be taken to start with if necessary.
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P
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?
The duration time of a program is dependent on the number of the program. The duration is also dependent on how long the Nucleofector® System has been turned on as well as the number of Nucleofections you have done in the last couple of...
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