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515 results sorted by
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D
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There is a publication reporting on the successful reinjection of modified hepatocytes into mice.Chen et al. were able to monitor the expression over several days. This group worked with a preliminary test solution for mouse hepatocyte as the...
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®
s
t
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l
?
Yes, transfected mouse hepatocytes show albumin synthesis rates comparable to those of untransfected cells. Moreover the morphology and polarization of rat hepatocytes is not affected by Nucleofection®.
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N
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n
™
?
Specific blood cell populations can be enriched/purified for Nucleofection™ by MACS™ selection Kits (Miltenyi Biotec GmbH), RosetteSep™ Kits (StemCell Technologies Inc.) or Dynal™ beads (Invitrogen Corporation).
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?
One reason could be that your gene product is toxic to the cells. We recommend testing the construct for expression of the gene of interest by transient Nucleofection® of your cells. If the proportion of expressing cells drops between 4 and 48 hours...
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?
Depending on individual cell type and doubling rate, selection of stable transfectants will take between 7 and 28 days. Expansion and characterization of single cell clones will take several weeks in addition.
D
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N
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®
?
We recommend using negative selection (depletion) because your purified cells are "untouched", not influenced, and strictly not activated.Positive selection may cause increasing amounts of dead cells and could also lead to activation of the cells due...
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C
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6
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?
Yes. To check the quality of your DNA we strongly recommend determining the A260:A280 ratio. It should be ideally 1.8 - 2.0 for a good DNA preparation, but least 1.6.
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b
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x
p
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d
?
This is the normal pattern you should expect to see. Only a small proportion (1/10,000 to 1/100) of all transfected cells will integrate the transfected DNA into their genome and become stable transfectants. The remaining cells lose the transfected...
C
a
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D
N
A
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?
Yes. You should verify the integrity of your DNA on an agarose gel to see if it is degraded. Compare undigested plasmid to plasmid DNA digested with a suitable single cut restriction enzyme to linearize. Supercoiled plasmid will run faster than...
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l
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i
n
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s
?
The most commonly used marker is the neomycin phosphotransferase (neo) gene that confers resistance to G418 to eukaryotic cells. Other markers are Puromycin, Hygromycin, Zeocin, or the HPRT gene that can be used in HPRT-deficient cells.
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