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Hygromycin B EvoPure®

Working concentrations and Hygromycin Selection Protocols

Hygromycin B working concentrations, for the purpose of selection, vary with cell type, media, growth conditions
and cell metabolic rate. Recommended hygromycin B concentrations for the selection of resistant cells is
25-1000 μg/mL. Commonly used concentrations for selection are 200-500 μg/mL for mammalian cells,
100-300 μg/mL for plant & bacterial cells and 300-1000 μg/mL for fungi. Your optimum hygromycin
concentration should be tested experimentally only once for EvoPure®.

Most cells growing aerobically are killed by hygromycin B in the concentration range of 10
to 1000 μg/mL. EvoPure® hygromycin B is less pH dependent in comparison with other
commercially available hygromycin B batches.

Due to the higher purity of EvoPure™ agents, suggested Hygromycin EvoPure™ selection concentrations
are lower than normal Hygromycin. For more rapid and accurate selections and to avoid tedious
re-optimizations, even higher concentrations of Hygromycin can be used. The following are
some examples of working concentrations:

Species Tissue Cell Line Plasmid/ Vector Culture/Media Selection Concentration (μg/mL)
Normal EvoPure® EvoPure®(rapid selection)
Rat Glioma C6 pCEP4 DMEM, 10% FBS 200 170 300
N/A N/A PhoeNX LXSH DMEM, 10% FBS 150 130 250
Human embryo Kidney HEK293 pUHD10-3, pUHC13-6, pUHD15-1, pUHG17-1, pUHD172-1 DMEM, 10% FBS 50 40 70
Procyclic trypanosomes N/A Trypanosoma brucei 427-60 pI-ble-KO SDM-79 40 30 80
Mouse Embryo J1 ES pBS524 N/A 200 170 300
Human Kidney 293 pCR2.1-TOPO, pCMVSPORTbGal DMEM, 9% FBS 200 170 300
Drosophila N/A S2 pAc, pCoHygro SF900II 500 430 700
Leishmania donovani N/A L. donovani promastigotes pX63-NEO, pX63-HYG DME-L 50 40 75
Homo sapiens (human) Uterus MES-SA/DX5 pSMDR-HYG,pSCONT-HYG McCoy 5A medium,1.5
mM L-glutamine,10% FCS,37℃,95%humidity,5%CO2
250 210 350
Homo sapiens (human) Burkitt's lymphoma BJAB EBV-based vector RPMI1640,10% FBS,1% non-essential amino acids,1 mM sodium pyruvate,2 mM
L-glutamine,37°C, 5% CO2
250 210 350
Homo sapiens (human) Burkitt's lymphoma Raji EBV-based vector RPMI1640,10% FBS,1% non-essential amino acids,1 mM sodium pyruvate,2 mM
L-glutamine,37°C, 5% CO2
300 260 500
Homo sapiens (human) Breast cancer T47D pVgRXR MEM,10% FBS,292 g/l L-glutamine,2.2 g/l sodium
bicarbonate, 6 ng/ml insulin,37°C, 5% CO2
200 170 300
Rat   Rat-1 pHyg DMEM,10% FBS 200 170  
Homo sapiens (human) Bone hFOB pAR-HYG DMEM,10%(v/v) csFBS 100 80 250
Homo sapiens (human) Dermal microvascular endothelial cell line HMEC-1 pCEP-4 MCDB-131,10% FBS,2 mM L-glutamine,1 mg/ml hydrocortisone,37°C, 5% CO2 200 170 300