GPT Selection Reagent, 500X Mycophenolic Acid; 100X Aminopterin

Description

• General description: Mycophenolic Acid is an antibiotic useful in research for the selection of animal cells that express the E. coli gene coding for XGPRT (xanthine guanine phosphoribosyltransferase). Mycophenolic Acid is also used as immunosuppressive agent becasue it acts by inhibiting lymphocyte proliferation and antibody production. Cells transfected with the E. coli GPT gene can be selected for growth in medium containing GPT Selection Reagent. Each Kit Contains:• 500X Mycophenolic Acid containing ETOH (12.5mg/mL) 2 x 1mL vials• 1 bottle (10mL) of 100X Aminopterin solution which consists of: - xanthine 25 mg/mL - hypoxanthine: 1.5 mg/mL - aminopterin: 0.2mg/mL - thymidine: 1 mg/mL - in 0.08M NaOH
• Application: 1. Seed approximately 106 cells on 100 mm culture plates in Dulbecco′s modified Eagle medium (DMEM) containing 5% fetal calf serum and penicillin/streptomycin, and allow them to grow for twenty-four hours. 2. Transfect the culture with 10-20 μg of the plasmid-EcoGPT DNA. 3. After 3 days at 37°C in DMEM containing 5% fetal calf serum, trypsinize the cell monolayers and disperse 5 x 105 cells on fresh 100 mm culture plates in DMEM containing 10% dialyzed fetal calf serum supplemented with the MPA and Aminopterin solutions. This will give a final solution of approximately: 250 μg/mL xanthine, 15 μg/mL hypoxantine, 10 μg/mL thymidine, 2 μg/mL aminopterin, and 25 μg/mL mycophenolic acid. Dialyzed serum is used to speed the selection process. Selection will be slower with undialyzed serum. 4. Twenty-four hours later, replace the medium with fresh medium containing the same supplements, and fluid change. 5. Repeat every 3 days. Colonies are visible in 7-10 days, typically. Note: Other media bases can also be used such as Ham′s F12. Much depends upon the cell type being used.
• Storage and Stability: Store at -20°C
• Disclaimer: Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.