Download Product Information N2 Supplement (100X)

 Product Information N2 Supplement (100X) A chemically defined, animal‐free, supplement used for expansion of undifferentiated neural stem and progenitor cells in culture. Can also be used for maintenance of post‐mitotic primary neurons in culture. Catalog No. Unit Size Shipping Storage Safety Information GSM‐3300 5 mL Frozen on dry ice Store at ‐20C until ready to use BioSafety Level: 1 Safety conditions pertaining to well characterized agents not known to cause disease in healthy adult humans and of minimal potential hazard to laboratory personnel and the environment should be used. Lab coats and gloves should be worn at all times. Face protection is recommended when thawing a frozen vial stored in a liquid nitrogen freezer. Intended Use N2 Supplement is an animal‐free, chemically‐defined supplement used to maintain neural stem and progenitor cells in an undifferentiated state. Ideally used with DMEM/F12. While N2 can be used to grow primary neurons, our data indicates that NeuralQ® with GS21 is better suited for this application. It is not recommended for use with PluriQ™ ES‐DMEM. For Research Use Only (RUO). Caution: Not intended for human or animal diagnostic or therapeutic uses. Fig. 1 ‐ Primary Rat Neural Stem Cells (GSC‐8010) were cultured in DMEM/F12 with N2 supplement. After 3 passages, cells were co‐labeled with Sox2 (green) and Nestin (red). Formulation Component rH Insulin Human apo‐Transferrin Putrescine Sodium selenite Progesterone Concentration (100X supplement) 2500 µg/mL 10,000 µg/mL 10 mM 3 µM 2 µM Product Dilution 1. Thaw 100x N2 Supplement overnight at 4°C. a. If thawed N2 is cloudy or a precipitate is seen, swirl the bottle in a 37°C water bath for several minutes until it clears. b. Thawed N2 can be stored at 4°C for up to two weeks. Do not refreeze/rethaw the supplement! 2. Supplement the DMEM/F12 medium with 2 mM L‐Glutamine or L‐Alanyl‐L‐Glutamine, 1X Non‐
Essential Amino Acids (NEAA), and D(+)Glucose to 4500 mg/Liter final concentration. a. For example, Life Tech DMEM/F12 (cat. #10565) contains 3151 mg/L D‐glucose; to prepare 500 mL: i. Weigh out 0.675 g D(+)Glucose ii. Dissolve in 10 mL DMEM/F12 iii. Sterilize via 0.2 micron filter, add to 470 mL DMEM/F12 iv. Add 5 mL 200 mM L‐Alanyl‐L‐Glutamine (e.g. Corning Glutagro) v. Add 5 mL 100X NEAA 3. Add N2 Supplement to 1X final concentration. 4. Add FGF2 (GSR‐2001) to 20 ng/mL. For some applications you may also add EGF to 20 ng/mL. 5. The complete 1X N2 medium can be stored at 4°C for up to three weeks. To preserve FGF2 activity, avoid repeatedly warming the medium. Instead, dispense and warm only what is needed for the day. References Bottenstein JE, Sato GH. Growth of a rat neuroblastoma cell line in serum‐free supplemented medium. Proc Natl Acad Sci U S A. 1979 Jan;76(1):514‐7.