Puromycin 10 mg/mL Cell Culture Reagent

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• Puromycin for Mammalian Cell SelectionPuromycin is an aminonucleoside antibiotic from Streptomyces alboniger that mimics the aminoacyl end of tRNA, enters the ribosomal A site and causes premature release of a truncated peptide chain, killing cells that cannot inactivate it. In mammalian cell culture it is used as a selection agent at 0.5-10 ug/mL, most commonly 1-2 ug/mL, and it kills non-resistant cells faster than any other common selection antibiotic -- typically within 2-5 days. Resistance is conferred by the pac gene encoding puromycin N-acetyltransferase, which acetylates the drug and renders it inactive. Because the effective concentration depends strongly on cell line, cell density and serum content, a kill curve must be run for each new cell line and each new lot of puromycin.
• Penicillin-Streptomycin and Antibiotic-Antimycotic in Cell CulturePenicillin-streptomycin, universally shortened to pen-strep, is supplied as a 100X sterile solution containing 10,000 units/mL penicillin G and 10,000 ug/mL streptomycin, diluted 1:100 into medium to give a working concentration of 100 U/mL penicillin and 100 ug/mL streptomycin. Penicillin blocks bacterial cell wall synthesis and covers mainly Gram-positive organisms; streptomycin binds the bacterial 30S ribosomal subunit and covers mainly Gram-negatives, so the pair gives broad antibacterial coverage. Antibiotic-antimycotic (anti-anti) is the same combination plus 25 ug/mL amphotericin B at 100X, giving 0.25 ug/mL in use for antifungal coverage. Neither controls mycoplasma, and major cell banks including ATCC recommend against routine antibiotic use because it masks low-level contamination rather than preventing it.
• Selection Antibiotics for Mammalian Cell LinesSelection antibiotics are cytotoxic agents used to isolate mammalian cells that carry a co-transfected resistance gene, by killing every cell that does not. The five standard agents are G418/geneticin (100–2000 µg/mL, resistance gene neo), hygromycin B (50–1000 µg/mL, hph), blasticidin S (1–20 µg/mL, bsd or bsr), zeocin (50–1000 µg/mL, Sh ble) and puromycin (0.5–10 µg/mL, pac). The correct concentration is not the one on the datasheet but the one determined empirically for your cell line by a kill curve — the lowest concentration that kills 100% of untransfected cells within the agent's normal selection window, which is 2–4 days for puromycin and up to 14 days for G418.