What is the difference between Lipofectamine® and Lipofectamine® PLUS™ reagent?

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Answer

Lipofectamine® and the PLUS™ Reagent are utilized for transfection of a wide range of cell types. In the past, Invitrogen™ sold Lipofectamine® with the PLUS™ reagent. This is no longer the case. Generally speaking, one would not use the PLUS™ Reagent by itself. It is used in conjunction with some other reagent, such as Lipofectamine®, Lipofectamine® LTX or in some cases, with Lipofectin®. Lipofectamine® LTX is offered in combination with the PLUS™ reagent in catalog numbers A12621 and 15338100. We do not recommend using the PLUS™ Reagent with Lipofectamine® 2000 as it does not generally help. .

Answer Id: E3035

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ae8c64bd66d35b9eca16b1d46b810464_FAQ

Are there any guidelines for choosing a lipid transfection reagent?

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Answer

It is best to optimize for your cells and application. Here are some basic guidelines:

- Lipofectamine® LTX and PLUS™ Reagent: Minimal optimization, excellent efficiency with adherent eukaryotic cells DNA, difficult cell lines
- Lipofectamine® Reagent: Adherent eukaryotic cells, DNA, oligonucleotides
- Lipofectin® Reagent: Transfecting DNA in eukaryotic cell
- Cellfectin® II: Insect cells
- DMRIE-C Reagent: DNA, RNA, suspension cells
- Oligofectamine™: Oligonucleotides
- Lipofectamine® RNAiMAX: siRNA, pre-miR, miRNA, anti-miR

NOTE: Please also visit our online Transfection Selection Tool to get specific recommendation for your cell line

Answer Id: E3079

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a44e61a1d926f9486ec0d15596251cba_FAQ

What kind of tubes can I use to form DNA:lipid complexes?

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Answer

We recommend using polypropylene tubes.

Answer Id: E3124

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aab9bac642dd798e1a1bf44d4408dc76_FAQ

I have optimized my conditions for a particular cell line but I seem to be getting inconsistent results with my transfection. What factors contribute to this inconsistency?

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Answer

Cells from a different passage number may behave differently. Also, if cells were sitting at confluence prior to plating for transfection, they may not transfect efficiently. To minimize such inconsistencies, passage the cells while they are still growing exponentially. Actively dividing cells transfect better.

Answer Id: E3125

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1c1f5027dd54726203418309d0a5000b_FAQ

Can I use the same amount of any lipid reagent for transfection of my particular cell line?

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Answer

No. The amount of lipid for each lipid reagent should be optimized for each cell line. Each lipid reagent has different composition/formulation, which will have different impact on each cell line. Therefore please optimize whenever you have a new lipid for each cell line.

Answer Id: E3126

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46c1c8d405f800444c08e5ab73e5feb3_FAQ

Does the method of generating lipid-DNA complex affect transfection efficiency?

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Answer

YES. Please follow the recommended procedure for each one of the reagents, found in the product manual.

Answer Id: E3127

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b85873045a0a02ad90bfd8aea5853fb9_FAQ

What types of molecules can be transfected with cationic lipid reagents?

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Answer

Lipid reagents can be used to transfect DNA, RNA, oligonucleotides and siRNA. The DNA can be plasmids, cosmids, or even YAC clones up to 600 kb. Lipofectin® and Lipofectamine® 2000 Reagent have also been used to transfect cells with proteins (Beta galactosidase and T3 polymerase).

Answer Id: E3128

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25a97d644624328d60fc3a57921caf82_FAQ

Can antibiotics be used in media during transfection?

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Answer

We discourage using any antibiotics during transfection (e.g. Geneticin®, Hygromycin, Gentamycin, Penicillin, etc). There can be higher cell death when antibiotics are present during transfection. Even though some such as penicillin and streptomycin are not toxic to eukaryotic cells in a healthy culture, during transfection the cell permeability increases so that much higher levels of antibiotics get into cells. For stable transfections, wait at least 24-48 h after transfection before adding selective antibiotics.

Answer Id: E3129

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b696afc7c66bb4ee77c91bbe14d9bc49_FAQ

Is it necessary to use serum-free media during lipid transfection?

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Answer

Not in all cases. What is essential is to form the lipid:nucleic acid complex in the absence of serum, because proteins can interfere with complex formation. Once the complexes are formed, they can be added to cells in serum containing medium. For optimal results, with Lipofectin® perform transfection in medium without serum.

Answer Id: E3131

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d128a808b58012e27949dbd6ad773aab_FAQ

What is the shelf life of the lipid transfection reagents?

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Answer

Stored at 4°C in a sealed container, the lipids are stable for 12 months. Do not freeze the lipids. At 4°C with long term storage, due to evaporation concentration of lipid may vary, please briefly spin the contents before use. Add less lipid if you start noticing toxicity.

Answer Id: E3132

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28e9e45a965960f6bf96fdfd9b2dadf5_FAQ

Can lipid reagents be used to cotransfect plasmids?

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Answer

Yes. The standard transfection protocol may be followed by keeping the total amount of DNA in the mixture constant. That is, if your protocol requires 1 ug plasmid, use 0.5 ug of each of two cotransfecting plasmids, or 0.25 ug of each of 4, etc. When performing cotransfections to introduce a selectable marker on a different plasmid, we recommend using a 3:1 to 10:1 molar excess of the plasmid of interest over the selectable plasmid to ensure that the plasmid of interest is present with the selectable plasmid.

Answer Id: E3133

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e511becc41e99f20519fe84d901b4e66_FAQ

How do cationic lipids compare with calcium phosphate in transfection efficiency?

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Answer

For many cell types, higher efficiencies are observed with cationic lipids than with calcium phosphate. Also, cationic lipid data are more reproducible from experiment to experiment. Calcium phosphate is inexpensive however, but pH variation as little as 0.2 can reduce transfection efficiency significantly.

Answer Id: E3137

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dec9a0f002f2e91166d3ae74df29452e_FAQ

How do I scale up or down my transfection reaction?

Product FAQ

Answer

The number of cells, DNA, lipid, and medium volumes should be scaled up proportionately to the surface area of the plate. For commonly used culture vessels, please refer to the information below regarding actual area and area relative to a 24-well plate well.

Vessel type, Area (cm2), Area Relative to 24-well
96-well, 0.3 cm2, 0.2
48-well, 0.7cm2, 0.4
24-well, 2 cm2, 1
12-well, 4 cm2, 2
6-well, 10 cm2, 5
35 mm, 10 cm2, 5
60 mm, 20 cm2, 10
100 mm, 60 cm2, 30
150 mm, 140 cm2, 70
T25, 25 cm2, 12.5
T75, 75 cm2, 37.5
T150, 150 cm2, 75
T162, 162 cm2, 81
T165, 165 cm2, 82.5
40-50 ml, 25 cm2, 12.5
250-300 ml, 75 cm2, 37.5
650-750 ml, 162-175 cm2, 81-87.5
900 ml, 225 cm2, 112.5

Answer Id: E3138

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e17c91a9c4d7125012544e0f649519ca_FAQ

How does the use of Lipofectamine® PLUS™ reagent improve transfection over other cationic lipid reagents?

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Answer

In general, the transfection activity peak with Lipofectamine® PLUS™ reagent is broader, thus essentially eliminating the need for lengthy optimization experiments. Because the protocol includes a pre-complexing step with PLUS™ reagent, only half the amount of Lipofectamine® reagent is required. The transfection time is short and the transfection can be done in the presence of serum. Difficult to transform cells (e.g. HT29, SKBR3 and MDCK) are transformed at a higher efficiency. See, FOCUS 19.3, 52.

Answer Id: E3163

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99dbc6156ee40713aff9cbd12bb5a069_FAQ

How do Lipofectamine® PLUS™ and Lipofectamine® compare in transfection efficiency?

Product FAQ

Answer

As a general rule, Lipofectamine® PLUS™ Reagent gives up to 5-times more peak activity with about half as much DNA than Lipofectamine® Reagent.

Answer Id: E3164

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c4c35b79bce367ca94451eef0f2f0c69_FAQ