Commonly know as G418 or G-418, Geneticin reagent is an aminoglycoside related to Gentamicin, and is commonly used as a selective agent for eukaryotic cells. As an analog of neomycin sulfate, it interferes with the function of 80S ribosomes and protein synthesis in eukaryotic cells. 

The dominant-acting resistance genes (neor) used on common mammalian expression vectors contain elements derived from either transposons Tn601 (903) or Tn5. When grown in medium containing Geneticin selection agent, stable colonies of mammalian cells expressing these resistance markers can be generated in 10 to 14 days.

What's in your G-418?

To be effective, selection agents need to be reliable, highly pure, and consistent in nature.  Geneticin selective antibiotic, considered the gold standard for nearly two decades, is the only G-418 reagent that delivers these qualities to your mammalian cell cultures.

Compared to other manufacturers’ G-418 products, Geneticin antibiotic exhibits higher purity, so you can use less and produce greater selection pressure with minimal toxicity from contaminants.

When evaluating G-418 products, consider more than just potency. Look at purity and working range. Taken together, these three characteristics provide a true evaluation of the effectiveness of your G-418.

Purity.  The purity of Geneticin® selection agent is consistently greater than 90%, as determined by HPLC — far higher than any other supplier. This means:

  • 15-30% lower concentrations of Geneticin® agent can be used, as compared to other G-418 products, to achieve comparable selection results.
  • Surviving clonal colonies may arise faster, and cells may appear healthier when using Geneticin® instead of lower purity G-418 products.
Potency.  Potency as a descriptive term is often used synonymously with purity, but potency as reported in QC data is actually not related to purity.  The potency value is a measure of bacterial growth inhibition.

  • Gentamicins that commonly contaminate G-418 preparations from other suppliers will add to potency in bacterial assays yet have no effect on mammalian cell selectivity.
  • Other bacterial contaminants in impure G-418 may be toxic to mammalian cells, consequently lowering the ED 50 value.

Therefore, comparing potency values between different suppliers is not a relevant specification for mammalian cell selection.

ED50 Assay. This is a true measure of eukaryotic growth selectivity as determined on reference NIH3T3 cells. Higher purity generally translates to higher ED50 values.

  •  Toxic impurities lower the ED50 value, which means a more narrow working range of antibiotic selection.
  •  A consistent ED50 from a supplier assures performance reproducibility from lot-to-lot in mammalian cell culture applications.


This means when purchasing a new lot of Geneticin®, you do not need to re-optimize for ideal antibiotic concentration (assuming that no other media alterations). The wide ranges of ED50 displayed in different lots from alternative G-418 suppliers may require you to re-optimize for each lot. Lot-to-lot consistency with Geneticin® is sufficiently high that you will not notice any difference in selection performance between lots.

Purity.  The purity of Geneticin® selection agent is consistently greater than 90%, as determined by HPLC — far higher than any other supplier. This means:

  • 15-30% lower concentrations of Geneticin® agent can be used, as compared to other G-418 products, to achieve comparable selection results.
  • Surviving clonal colonies may arise faster, and cells may appear healthier when using Geneticin® instead of lower purity G-418 products.
Potency.  Potency as a descriptive term is often used synonymously with purity, but potency as reported in QC data is actually not related to purity.  The potency value is a measure of bacterial growth inhibition.

  • Gentamicins that commonly contaminate G-418 preparations from other suppliers will add to potency in bacterial assays yet have no effect on mammalian cell selectivity.
  • Other bacterial contaminants in impure G-418 may be toxic to mammalian cells, consequently lowering the ED 50 value.

Therefore, comparing potency values between different suppliers is not a relevant specification for mammalian cell selection.

ED50 Assay. This is a true measure of eukaryotic growth selectivity as determined on reference NIH3T3 cells. Higher purity generally translates to higher ED50 values.

  •  Toxic impurities lower the ED50 value, which means a more narrow working range of antibiotic selection.
  •  A consistent ED50 from a supplier assures performance reproducibility from lot-to-lot in mammalian cell culture applications.


This means when purchasing a new lot of Geneticin®, you do not need to re-optimize for ideal antibiotic concentration (assuming that no other media alterations). The wide ranges of ED50 displayed in different lots from alternative G-418 suppliers may require you to re-optimize for each lot. Lot-to-lot consistency with Geneticin® is sufficiently high that you will not notice any difference in selection performance between lots.

Table 1. Comparison of Geneticin specifications with alternative G-418 suppliers.

Specification
Invitrogen
Supplier A
Supplier B
The Geneticin Advantage
Purity (HPLC)
>90-93%
66-75%    65-82% Significantly higher purity
Potency claimed (µg/mg)
718-735
712-724
673-735
More stringent potency assay
Potency upon retest (µg/mg)
718-735
640-659
621-677
Consistently higher actual potency
ED50 Assay (µg/ml)
2450-2700
1350-3100
600-2350
Reliable selection lot-to-lot


Table 1. Multiple lots of G-418 from two different suppliers were analyzed according to Invitrogen raw material QC protocols for Geneticin. It was observed that results obtained with G-418 from Supplier A and Supplier B displayed wide variations on all measured parameters, including an inconsistency between the actual potency and the value reported on the supplied product literature.  Discrepancies between re-tested potency values and that claimed by other suppliers may be due to differences in G-418 reference lots and assay procedures.

Figure 1. Geneticin displays greater purity than G-418 from alternative suppliers.



Figure 1. Comparison of HPLC data of Geneticin with selected lots of G-418 from Supplier A and Supplier B. Samples were prepared and analyzed according to the procedure published in Clinical Chem. (1978), Vol. 24, No.11, p1940.