Puromycin 2HCl (10 mg/mL Solution), Sterile, IC-1609C

Product Introduction

This product is a ready-to-use, sterile-filtered 10 mg/mL solution of puromycin dihydrochloride, a broad-spectrum aminonucleoside antibiotic produced by Streptomyces alboniger. Puromycin functions as a structural analog of the 3''''-terminal aminoacyl-adenosine moiety of charged tRNA, causing premature chain termination during translation by incorporation into the growing polypeptide chain in both prokaryotic and eukaryotic systems. This mechanism of action makes puromycin a potent and rapid inducer of cell death in non-resistant cells. In molecular biology and cell engineering, puromycin is the most widely used selection antibiotic for the establishment and maintenance of stable mammalian cell lines expressing the puromycin N-acetyltransferase (pac) resistance gene. Cells transfected with the pac gene detoxify puromycin by acetylation, enabling selective survival and propagation of successfully transfected populations. This sterile, ready-to-use solution is supplied at a standard concentration of 10 mg/mL in sterile deionized water, eliminating the need for powder weighing, dissolution, and filter sterilization, thereby reducing contamination risk and ensuring reproducible selection conditions.

Product Features

1. Ready-to-Use Sterile Solution: Supplied as a sterile-filtered 10 mg/mL solution requiring no preparation, dilution, or sterilization before use.

2. Rapid and Potent Selection: Puromycin induces rapid cell death in sensitive cells, typically achieving complete selection within 2-7 days depending on cell type.

3. Broad-Spectrum Activity: Effective against both prokaryotic and eukaryotic cells, providing reliable selection pressure for stable cell line generation.

4. Standard Working Concentration: The 10 mg/mL stock allows convenient dilution to typical working concentrations of 1-10 µg/mL for most mammalian cell lines.

5. Consistent Quality: Each lot is tested for sterility, concentration accuracy, and selection efficacy to ensure reproducible performance.

Specifications

Size: Available in 500 µL, 1 mL, and 5 mL

Concentration: 10 mg/mL in sterile deionized water

Form: Sterile-filtered solution

Antibiotic Class: Aminonucleoside antibiotic

Selection Marker: Puromycin N-acetyltransferase (pac gene)

Typical Working Concentration: 1-10 µg/mL for mammalian cells (varies by cell type)

Storage and Stability

Storage Conditions: Store at -20¡ãC, protected from light. For frequent use, aliquot and store at -20¡ãC. Avoid repeated freeze-thaw cycles. Working aliquots may be stored at 4¡ãC for up to 2 weeks.

Shelf Life: The product is stable for 24 months from the date of manufacture when stored as directed.

Protocol (For Reference Only)

Important: Puromycin is toxic. Handle with appropriate safety precautions. Determine the optimal selection concentration by performing a kill curve for each cell line before initiating stable cell line generation.

1. Kill Curve Determination: Seed cells in a 24-well plate at a density that yields 50-70% confluency after 24 hours. Add puromycin at concentrations ranging from 0.5-20 µg/mL (e.g., 0, 0.5, 1, 2, 5, 10, 15, 20 µg/mL) in complete culture medium. Monitor cell viability daily and replace medium with fresh puromycin-containing medium every 2-3 days. Identify the minimum concentration that kills 100% of non-resistant cells within 3-7 days. This concentration should be used for selection of stable cell lines.

2. Stable Cell Line Selection: 24-48 hours after transfection with a pac-containing vector, replace the culture medium with complete medium containing puromycin at the predetermined optimal concentration. Maintain selection pressure by replacing puromycin-containing medium every 2-3 days. Non-resistant cells will die within 2-7 days. Continue selection until resistant colonies are established.

3. Maintenance of Stable Cell Lines: For established stable cell lines, maintain puromycin at half the selection concentration (typically 0.5-2 µg/mL) in culture medium to prevent loss of transgene expression. Continuous selection pressure is recommended for long-term maintenance.

4. Bacterial Selection: For E. coli selection, use puromycin at concentrations of 50-125 µg/mL in LB medium. Note that puromycin is less commonly used for bacterial selection compared to ampicillin or kanamycin; verify compatibility with your specific vector and host strain.

5. Dose-Response and Time-Course Assays: For metabolic labeling or translation inhibition studies, treat cells with puromycin at 1-10 µg/mL for defined time periods (typically 10-60 minutes). Assess translation inhibition by puromycin incorporation assays using anti-puromycin antibodies or metabolic labeling techniques.

Precautions

1. Puromycin is toxic; wear appropriate protective clothing, gloves, and eye protection. Handle in a biosafety cabinet and dispose of waste according to institutional guidelines for hazardous compounds.

2. Avoid repeated freeze-thaw cycles; aliquot the solution into single-use volumes upon receipt.

3. The optimal working concentration must be determined empirically for each cell line. Sensitivity varies significantly between cell types.

4. Puromycin selection may require 2-7 days for complete killing of non-resistant cells depending on cell type, seeding density, and antibiotic concentration.

5. For research use only. Not for use in diagnostic or therapeutic procedures.

FAQ (Simplified)

Q1: What concentration of puromycin should I use for my cell line?

A1: The optimal puromycin concentration varies by cell type. Perform a kill curve experiment testing concentrations from 0.5-20 µg/mL to determine the minimum concentration that kills 100% of non-transfected cells within 3-7 days. Common working concentrations for mammalian cells range from 1-10 µg/mL.

Q2: How long does puromycin selection take?

A2: Complete selection typically requires 2-7 days depending on cell type, antibiotic concentration, and transfection efficiency. Replace puromycin-containing medium every 2-3 days to maintain selection pressure.

Q3: Can I maintain stable cell lines without continuous puromycin selection?

A3: While stable integration can persist without selection, continuous low-level puromycin (typically half the selection concentration) is recommended to prevent gradual loss of transgene expression due to epigenetic silencing or outgrowth of non-expressing cells.

Q4: Is this solution sterile?

A4: Yes. This product is supplied as a sterile-filtered solution, allowing direct addition to cell culture medium without further sterilization.

Disclaimer

1. For Research Use Only. Not for use in diagnostic or therapeutic procedures.

2. Due to the variable nature of biological research, determination of the optimal puromycin concentration by kill curve analysis is recommended for each cell line.

3. This warranty is limited to the replacement of the product. The manufacturer assumes no liability for incidental or consequential damages, including loss of samples or data.

4. Wear appropriate protective clothing and gloves when handling this product. Puromycin is toxic; review the Safety Data Sheet before handling.

Ordering Information

Catalog Number: IC-1609C

Product Name: Puromycin 2HCl (10 mg/mL Solution), Sterile

Size: 500 µL / 1 mL / 5 mL

Price:

500 µL: CNY ¥200.00 / USD $20.00 / EUR €24.00 / JPY ¥3600.00

1 mL: CNY ¥300.00 / USD $30.00 / EUR €36.00 / JPY ¥5400.00

5 mL: CNY ¥800.00 / USD $80.00 / EUR €96.00 / JPY ¥14400.00

                                                                                                                                                                                                                    2023 Version