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TransPass™ COS/293 Transfection Reagent |
 | | | | TransPass™ COS/293 Transfection Reagent | | | |
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Prices are in US dollars and valid only for US orders.
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Description:
The TransPass™ COS/293 Transfection Reagent is TransPass™ D2 Transfection Reagent, a non-lipid cationic polymer, tested for the transfection of plasmid DNA into COS and HEK293 cell lines in a serum-compatible protocol. Complex co-transfections of many plasmids can be efficiently transfected using the TransPass COS/293 Transfection Reagent.
Figure 1: COS-7 cells grown and transfected on a slide coverslip with eGFP and 2 µl TransPass COS/293 Transfection Reagent in 6-well plate format. Data shown is 24 hr post-transfection.
Figure 2: Titration of TransPass COS/293 Transfection Reagent in HEK293 cells using secreted Gaussia Luciferase (pCMV-GLuc Control Plasmid) as a reporter. Cells are transfected in 12-well plate format and GLuc activity was measured in 10 µl cell supernatant 24 hr post-transfection.
Background:
The introduction of recombinant DNA into cultured cells or "transfection", has become an essential tool for studying gene function and regulation. Initially, cells were transfected by methods such as Calcium Phosphate co-precipitation, electroporation or viral factors (1, 2). The introduction of cationic lipids or polymers as transfection agents has led to the development of reagents that efficiently deliver DNA through the cell membrane and into the nucleus (3).
Transfection Guidelines:
For consistent results, it is important to maintain healthy proliferating cells that are regularly passaged.
Use sterile plasmid DNA that is purified by CsCl gradient centrifugation or column chromatography.
The following parameters can be optimized in order to maximize the transfection efficiency for a particular cell line: the cell density at the time of transfection, amount of transfection reagent, amount of plasmid DNA and the culture incubation time before analysis. The specific amount of transfection reagent can be optimized for a particular cell line by performing a simple titration (Figure 2).
Reaction & Storage Conditions
Storage Temperature:
4°C
Notes
Usage notes:- In order to form the transfection complexes (step 2), serum-free medium is required.
- For optimal transfection of a certain cell density the amount of TransPass COS/293 Transfection Reagent can be titrated while keeping the amount of plasmid DNA constant. Use Table 1 as a guideline for amounts using different plate sizes. A convenient method for optimizing transfection conditions is to use pCMV-GLuc Control Plasmid (NEB #N8081), which contains the gene for the secreted Gaussia Luciferase. See Figure 2 for an example titration in HEK293 cells. In addition, multiple plasmids can be combined (step 2) and simultaneously transfected.
Quality Control for Current Lot
Quality control values for a specific lot can be found on the datacard which accompanies each product.
Quality Assurance Statement:
Each lot of transfection reagent is tested for efficient delivery of two different reporter plasmids in COS-7 and HEK293 cells.
References
- Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, (2nd ed.), Cold Spring Harbor: Cold Spring Harbor Laboratory .
- Ausubel, F. M. et al. (1987) Current Protocols in Molecular Biology, (2nd ed.), New York: Greene Publishing Associates and Wiley- .
- Felgner P.L. et al. (1987) Proc. Natl. Acad. Sci. USA, 84, 7413-7417.
Companion Products
Gaussia Luciferase Assay Kit
Luciferase Cell Lysis Buffer
pCMV-GLuc Control Plasmid
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