Hygromycin B (50 mg/mL) - FAQs

Which of your antibiotics (Geneticin, Zeocin, Hygromycin B, Blasticidin, and Puromycin) can be used together for stable selection in mammalian cells?

All of our antibiotics (Geneticin, Zeocin, Hygromycin B, Blasticidin, and Puromycin) can be used together for making multiple stable cell lines. However, kill curves will need to be performed for each combination of antibiotics since sensitivity to a given antibiotic tends to increase when combined with other antibiotics.

What are the recommended concentrations of antibiotics to use for selection in prokaryotes and eukaryotes?

For best results, optimal concentrations for selection should be determined empirically in each unique experiment through dose response curves. However, to get a general idea of concentrations that have worked for individual cell types, please click on the following url: http://www.thermofisher.com/us/en/home/life-science/cell-culture/transfection/selection.html or type in “Selection Antibiotics” into our main search on www.thermofisher.com.

What is the range of specific activity of Hygromycin B?

The specific activity range of Hygromycin B is from 386-482 micrograms/mg.

Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.

What solvent is used to prepare your Hygromycin B solution? How should it be stored?

Hygromycin B is supplied as a 50 mg/mL solution in phosphate buffered saline. It is manufactured aseptically and sterile filtered, and is suitable for use in tissue culture applications. We recommend storing it at 2-8 degrees C.

What is the mode of action on the following antibiotics: Blasticidin, Geneticin (G418), Hygromycin, and Zeocin?

Blasticidin: Nucleoside Inhibits protein synthesis in prokaryotic and eukaryotic cells by interfering with peptidyl transfer reaction of protein synthesis, causing early termination of translation.

Geneticin (G418): Aminoglycoside Blocks protein synthesis in mammalian cells by interfering with ribosomal function.

Hygromycin: Aminocyclitol Inhibits protein synthesis by disrupting translocation and promoting mistranslation.

Zeocin: Intercalates with DNA and cleaves it.

What is the shelf life of Hygromycin B (50 mg/mL) (Cat. No. 10687010)?

The shelf life of Hygromycin B is 2 years from the date of manufacture, provided the product is subjected to normal and proper usage and storage. The date of manufacture can be found on the Certificate of Analysis of each lot number.

Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.

How can I decontaminate my cultures?

When an irreplaceable culture becomes contaminated, researchers may attempt to eliminate or control the contamination.

1. Determine if the contamination is bacteria, fungus, mycoplasma, or yeast. Read more here to view characteristics of each contaminant.
2. Isolate the contaminated culture from other cell lines.
3. Clean incubators and laminar flow hoods with a laboratory disinfectant, and check HEPA filters.
4. Antibiotics and antimycotics at high concentrations can be toxic to some cell lines. Therefore, perform a dose-response test to determine the level at which an antibiotic or antimycotic becomes toxic. This is particularly important when using an antimycotic such as Gibco Fungizone reagent or an antibiotic such as tylosin.

The following is a suggested procedure for determining toxicity levels and decontaminating cultures:

1. Dissociate, count, and dilute the cells in antibiotic-free media. Dilute the cells to the concentration used for regular cell passage.
2. Dispense the cell suspension into a multiwell culture plate or several small flasks. Add the antibiotic of choice to each well in a range of concentrations. For example, we suggest the following concentrations for Gibco Fungizone reagent: 0.25, 0.50, 1.0, 2.0, 4.0, and 8.0 µg/mL.
3. Observe the cells daily for signs of toxicity such as sloughing, appearance of vacuoles, decrease in confluency, and rounding.
4. When the toxic antibiotic level has been determined, culture the cells for two to three passages using the antibiotic at a concentration one- to two-fold lower than the toxic concentration.
5. Culture the cells for one passage in antibiotic-free media.
6. Repeat step 4.
7. Culture the cells in antibiotic-free medium for four to six passages to determine if the contamination has been eliminated.

Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.

What antibiotics do you offer to help control or eliminate cell culture contamination?

Please view the following page to browse the cell culture antibiotics we offer (https://www.thermofisher.com/us/en/home/life-science/cell-culture/mammalian-cell-culture/antibiotics.html).

Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.