Product FAQ

How much serum is in Opti-MEM I Medium?

Answer

There is no serum in Opti-MEM I Medium. The term "reduced serum" means that one can reduce the amount of serum that is used with classical cell culture media. When working with Opti-MEM I medium, most cells that are routinely cultured in serum-supplemented medium may be transferred directly into Opti-MEM I medium with a minimum of 50% reduction in serum.

Answer Id: E2993

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Product FAQ

Where can I find the osmolality for my lot of Opti-MEM I Reduced Serum Medium?

Answer

We do provide osmolality information on the certificate of analysis. All lots of Opti-MEM I Reduced Serum Medium will meet the osmolality specification of ≥280 to ≤320 mOsm/kg.

Answer Id: E17409

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Product FAQ

I ran out of OptiPRO SFM. Can I use use Opti-MEM I Reduced Serum Medium to make my DNA-ExpiFectamine 293 transfection complexes?

Answer

We do not recommend using OptiPRO SFM to make DNA-ExpiFectamine 293 transfection complexes. If you require an animal origin-free system, you may use FreeStyle 293 Expression Medium to make the complexes, but keep in mind that there will be a 10-20% drop in final protein yields.

Answer Id: E9263

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Product FAQ

Can I aliquot and freeze Opti-MEM I Reduced Serum Medium to conserve it longer? If so, for how long can it be stored?

Answer

Sorry, we don't have data to show that frozen Opti-MEM I Reduced Serum Medium has a longer shelf life. Actually, we recommend storing this medium at 2-8 degrees C in the dark. We do not recommend freezing the medium because it may precipitate.

Answer Id: E17375

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Manual / Product Insert

OPTI-MEM I Reduced Serum Material

Version: MAN0007448 1.01 (16 July 14)

Product FAQ

I noticed that Opti-MEM I Reduced Serum Medium does not contain any serum. Why do you then refer to it as “Reduced Serum Medium”?

Answer

Opti-MEM I Reduced Serum Medium is a serum-free medium. The reason why it is called “reduced-serum medium” is because you only need to supplement this medium with 1-5% FBS instead of the standard 10% FBS.

Opti-MEM I Reduced Serum Medium can be used as is (without serum) for transfection of cells. However, for long-term cell culture, you need to add serum to the medium to keep cells healthy.

Answer Id: E17374

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Product FAQ

Is there any other difference between Opti-MEM I Reduced Serum Medium and Opti-MEM I Reduced Serum Medium, no phenol red besides the presence/absence of phenol red?

Answer

The formulation of Opti-MEM I Reduced Serum Medium and that of Opti-MEM I Reduced Serum Medium, no phenol red is the same except for the presence of phenol red in the former medium.

Answer Id: E17451

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Product FAQ

Can I use OptiPRO SFM instead of Opti-MEM I Reduced Serum Medium to make my DNA-ExpiFectamine 293 transfection complexes?

Answer

We do not recommend using OptiPRO SFM to make DNA-ExpiFectamine 293 transfection complexes. If you require an animal origin-free system, you may use FreeStyle 293 Expression Medium to make the complexes, but keep in mind that there will be a 10-20% drop in final protein yields.

Answer Id: E9228

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Manual / Product Insert

Product Sheet: CTS Opti-MEM I

Product FAQ

Is the Expi293 Expression System completely animal-origin free (AOF)?

Answer

All the components of the system are animal-origin free except for the Opti-MEM I Reduced Serum Medium that is serum-free but not animal-origin free. Please see the Application Note for using the Expi293 Expression System under animal origin-free conditions:
http://www.thermofisher.com/content/dam/LifeTech/migration/files/proteins-expression-isolation-analysis/pdfs.par.5943.file.dat/expi-293-animal-origin-free-co25751.pdf

Answer Id: E9223

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Product FAQ

Can siRNA and plasmid be co-transfected into cells?

Answer

Yes, although you must use specific transfection methods and reagents to optimize the reaction. The procedure below is recommended to cotransfect your plasmid DNA and an RNAi molecule into mammalian cells using Lipofectamine 2000. A more detailed version can be found on our website by searching "RNAi Transfection Protocols".

1. One day before transfection, plate cells in the appropriate amount of growth medium without antibiotics such that they will be 80-90% confluent at the time of transfection.
2. For each transfection sample, prepare DNA-RNAi molecule-Lipofectamine 2000 complexes as follows:
a. Dilute the DNA and RNAi molecule in the appropriate amount of Opti-MEM I Medium without serum. Mix gently.
b. Mix Lipofectamine 2000 gently before use, then dilute the appropriate amount in Opti-MEM I Medium without serum. Mix gently and incubate for 5 minutes at room temperature.
c. After the 5 minute incubation, combine the diluted DNA and RNAi molecule with the diluted Lipofectamine 2000. Mix gently and incubate for 20 minutes at room temperature to allow complex formation to occur. The solution may appear cloudy, but this will not impede the transfection.
3. Add the DNA-RNAi molecule-Lipofectamine 2000 complexes to each well containing cells and medium. Mix gently by rocking the plate back and forth.
4. Incubate the cells at 37°C in a CO2 incubator until you are ready to harvest cells and assay for your target gene. Removal of complexes or media change is not required; however, growth medium may be replaced after 4-6 hours without loss of transfection activity.

Answer Id: E4531

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Product FAQ

What should I use to dilute my RNAi duplexes and Lipofectamine RNAiMAX before complexing?

Answer

We recommend using Opti-MEM I Reduced Serum Medium (Cat. No. 31985062).

Answer Id: E9038

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Product FAQ

How can I improve transfection efficiency with cationic lipids?

Answer

1. Select the cationic lipid reagent that is likely to result in highest transfection efficiency for your cell type. Please refer to the Transfection Reagent Selection Guide (https://www.thermofisher.com/us/en/home/life-science/cell-culture/transfection/transfection-reagent-application-table.html) to make the right choice.
2. Optimize the cationic lipid reagent and DNA amounts. The most important parameter after the condition of the cells is the ratio of lipid to DNA.
3. Do not use serum during complex formation. Serum may contain components that could interfere with complex formation. We recommend using Opti-MEM I Reduced-Serum Medium for optimal complex formation. However, serum-free DMEM or serum-free RPMI 1640 Medium can be used, keeping in mind that the efficiency of complex formation may not be as high as with Opti-MEM I Reduced-Serum Medium.
4. Do not use antibiotics, EDTA, citrate, phosphate, chondroitin sulfate, hyaluronic acid, dextran sulfate, or other sulfated proteoglycans in the medium used to prepare the DNA-cationic lipid reagent complexes.
5. Cell density should be from 50% to 80% confluency at the time of transfection (for Lipofectamine 2000, we recommend >90% confluency). Cells should be in the mid-log growth phase. For better consistency of results between transfection experiments, it would be best to accurately count your cells with a hemocytometer or with the Countess II FL Automated Cell Counter (Cat. No. AMQAF1000).
6. Make sure the promoter-enhancer of the transfected DNA is compatible with the target cell type.
7. Do not use cationic lipid reagent that has been frozen or stored in a section of the refrigerator where the temperature is below 4 degrees C.
8. Include a positive control for the transfection assay (for example, Cat. No. A14146 for plasmid DNA transfection and Cat. No. 14750100 for siRNA transfection).

Also, please take a look at the tips outlined here (https://www.thermofisher.com/us/en/home/life-science/cell-culture/transfection/transfection-support/troubleshooting-transfection-experiments.html).

Answer Id: E3994

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Product FAQ

What points should I consider to achieve optimal transfection with a lipid-based transfection reagent?

Answer

Here are some points to consider:

1. Select the lipid reagent that is likely to result in highest transfection efficiency for your cell type, payload, and application. Please refer to the Transfection Reagent Selection Guide (https://www.thermofisher.com/us/en/home/life-science/cell-culture/transfection/transfection-reagent-application-table.html) to choose the best reagent.
2. Optimize both lipid reagent and DNA quantities. The most important parameter after the condition of the cells is the ratio of lipid to DNA.
3. Do not use serum during complex formation. Serum may contain components that could interfere with complex formation. We recommend using Opti-MEM I Reduced-Serum Medium for optimal complex formation. However, serum-free DMEM or serum-free RPMI 1640 Medium can be used, but the efficiency of complex formation may not be as high as with Opti-MEM I Reduced-Serum Medium.
4. Do not use antibiotics, EDTA, citrate, phosphate, chondroitin sulfate, hyaluronic acid, dextran sulfate, or other sulfated proteoglycans in the medium used to prepare the DNA-cationic lipid reagent complexes.
5. Cell density should be between 50% to 80% confluency at the time of transfection (please refer to specific reagent manual for details). Cells should be in the mid-log growth phase. For better consistency and reproducibility of results between transfection experiments, accurately count your cells with either a hemocytometer or the Countess II FL Automated Cell Counter (Cat. No. AMQAF1000).
6. Confirm that the promoter and/or enhancer (any gene regulatory sequences) of the transfected DNA is compatible with the target cell type.
7. Do not use a cationic lipid reagent that has been frozen or stored at temperatures below 4 degrees C.
8. Include a positive control for the transfection assay (for example, Cat. No. A14146 for plasmid DNA transfection and Cat. No. 14750100 for siRNA transfection).

For additional tips ,please take a look at the tips outlined here (https://www.thermofisher.com/us/en/home/life-science/cell-culture/transfection/transfection-support/troubleshooting-transfection-experiments.html).

Answer Id: E8989

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Product FAQ

Are all components of the LV-MAX Production System animal origin-free?

Answer

Yes, all components of the LV-MAX Production System are animal origin-free. However, if Opti-MEM I Reduced Serum Medium is used as the DNA-LV-MAX Transfection Reagent complexation medium, it will no longer be animal origin-free.

Answer Id: E15230

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