For one of my cell culture assays, i need to add additional calcium. The culture medium i use has 1.6mM calcium. Do I need to count this. I am interested i effective concentration suitable for the calmadulin protein in cells. Can you please suggest an optimum concentration of CaCl2 to add in to the 6 well plate?? May be 10uM final concentration??
The concentration of calcium found in classic media and their derivatives varies from 0 in BGJb Medium Fitton-Jackson Modification, developed to culture cartilagenous embryonic bone, to 2 mM in MCDB media 201, developed to culture Chick Embryo Fibroblasts, an attached cell line. As a general rule, the highest concentrations of calcium are found in serum supplemented, such as FBS, classic media developed for attached cell types. Sera generally protects cells from oxidative damage. Calcium levels are generally lower in serum-free media and media developed for primary and clonal cell culture. This may be due to the fact that primary cells and cells grown at clonal densities are susceptible to the toxic influx of calcium through cell membranes damaged by oxidative processes, prevalent in serum-free culture media, and enzymatic and mechanical processing. Since calcium affects cell signaling and differentiation as well as cell attachment, media developed for specific cell types may require substantially lowered calcium levels.
The following classic media contain 1.8 mM calcium: Basal Medium Eagle (BME); CMRL-1066 Medium; Dulbecco's Modified Eagle's Medium (DMEM); Glascow Minimum Essential Medium, GMEM; H-Y Medium (Hybri-Max®); Medium 199; Minimum Essential Medium Eagle (EMEM); NCTC Medium; Swim's S-77 Medium; and Williams Medium E. Click's Medium; MCDB media 131 and Iscove's Modified Dulbecco's Medium (IMDM) contain slightly less calcium at 1.67, 1.60 and 1.49 mM, respectively.
L-15 Medium and Ames' Medium contain 1.26 and 1.15 mM of calcium, respectively.
DMEM/Ham's Nutrient Mixture F-12 (50:50) is frequently used as a base media for the development of proprietary and specialty media to culture CHO cells for biomanufacturing. This media contains 1.05 mM calcium, which is very close to the level of calcium in MCDB media 105 and 110 at 1.0 mM. MCDB media was developed for clonal growth of human diploid cells. Other classic mediadeveloped for the culture of primary cells such as Nutrient Mixture Ham's F-12 Kaighn's Modification (F12K) (0.92 mM); McCoy's 5A Modified Medium (0.9 mM) and Waymouth Medium MB (0.82 mM) also contain close to1 mM of calcium.
MCDB medium 302 developed for CHO cells contains 0.60 mM calcium. Nutrient Mixtures, Ham's F-10 and F-12; and Serum-Free/Protein Free Hybridoma Medium (which is based on F-12) all contain 0.30 mM calcium. RPMI-1640 contains 0.42 mM calcium. The concentration of calcium used in media for CHO culture ranges from 0.30 to 1.05 mM. DMEM//F-12 (50:50) media with 1.05 mM calcium is more frequently used as a base for development of CHO media used in biomanufacturing.
MCDB medium 151, and 153 developed for keratinocytes contain very low levels of calcium in the range of 0.03 mM. The very low levels of calcium used for keratinocytes reveals that calcium may have important regulatory or differentiation affects on specific cell types.
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