MEM, with NEAA, Custom pH

FAQs

• Q:1 Can Neurobasal Medium alone or with B-27 Supplement be frozen to extend the shelf life of the product?

  Answer

  Freezing the culture medium is not recommended because a precipitate may form upon thawing; the inorganic salts and amino acids in the formulation may come out of solution when the temperature fluctuates, and once formed, these precipitates do not easily return to solution.

• Q:2 If the basic culture medium is frozen at -20℃, will it affect its use?

  Answer

  If the cell culture medium is accidentally frozen, observe whether there is precipitation after the medium is naturally thawed. If no precipitation is produced, the medium can be used normally. If precipitation occurs, it is recommended to discard it.

• Q:3 My cells are growing very slowly. What could be the cause of this?

  Answer

  Please see some common reasons below, and solutions to the issue: - Growth medium is not correct: Use pre-warmed growth medium as recommended by the supplier. - Serum in the growth medium is of poor quality: Use serum from a different lot. and choose good quality serum to ensure nutrition. - Cells have been passaged too many times: Use healthy, low-passage number cells. - Cells were allowed to grow beyond confluency: Passage mammalian cells when they are in the log-phase before they reach confluency. - Culture is contaminated with mycoplasma: Discard cells, media, and reagents. Obtain new stocks of cells, and use them with fresh media and reagents.

• Q:4 What factors can contribute to rapid cell death/culture failure?

  Answer

  There are a number of events that can contribute to this: 1. Incorrect CO2 levels:Monitor the level of CO2 manually with a Fyrite kit, available from Bacharach. Check if the manual readings concur with the readings displayed on the incubator. If the incubator has a trace readout, check the printout for fluctuations in CO2 level. Check the settings to ensure that CO2 levels are set at appropriate levels for your cell line (usually between 5 and 10%). Check line connections frequently for leaks. Avoid frequent opening and closing of incubator doors. 2. Temperature fluctuations in the incubator:Monitor the temperature of incubator with a good thermometer inside the incubator. 3. Amphotericin B or other preventive antibiotics/antimycotics are present at toxic concentrations:Use at recommended levels. 4. Humidity is incorrect:Check the water level in the water pan. Humidity is vital to appropriate gas exchange for many types of cells and media. 5. Incorrect osmotic pressure in medium:Check osmolality of complete medium. Most mammalian cells can tolerate an osmolality of 260 to 350 mOsm/kg. Additions of reagents such as HEPES and drugs may affect osmolality. 6. Contamination by microorganisms:Bacterial and fungal contaminations are usually easily visible; symptoms of mycoplasma contamination are more subtle, and careful monitoring of culture morphology and regular testing are necessary to detect this type of contamination. 7. Inappropriate medium is being used:Double-check that the medium used is appropriate for your cell type and culture application. For example, ensure that the medium being used for serum-free culture is actually designed for serum-free culture; make sure that appropriate selective drugs are used at appropriate levels; check the expiration dates for the reagents being used; and store medium at appropriate temperatures in the dark.