The FBS or FCS, used in the cell cultures normally contains enough B12 required by the cells. If you would like to study the effect of B12 on something, you need to be very careful in designing the experiments. Whatever the amount of B12 you provide may be redundant in the cell culture. You need to dig the literature before you start doing the experiments. The following article may be useful to you. Good luck

http://www.pnas.org/content/106/51/21930

Amended on 30 May 2018

I have recently added biotin (vitamin H) at 5 μg/mL to the culture medium for Caco-2, since I have determined by HPLC-photometric method that the biotin concentration of children's serum is c.a. 3 μg/mL (please see files; JCB Fucoidan transport and Netherlands biotin). I have recently recognized that the serum biotin concentration (2 μg/mL) is very similar value to Km of serum biotinidase (9 μM) as BAQ was used as substrate (Mr 371) (please see files; JMBT alopecia and The Fascio effect). Therefore, vitamin B12 may possibly be present in the serum at a similar μg/mL level to biotin.

If your data of serum vitamin B12 concentration has been obtained by the notorious CLEIA (Chemiluminescent Enzyme Immunoassay) method, you must not believe in those CLEIA values. Notorious immunoassays (RIA and ELISA) usually give falsely small values.

The report of the above mentioned article (PNAS) is also not true, since non quantitative UPLC-MS analysis and parametric statistics are used. Further, PNAS is not a Journal without any referees.

I have previously found that fucoidan concentration is c.a. 1,000-fold higher as compared to notorious ELISA method (please see file; JCB Fucoidan transport). This is because (1) direct photometry reduces 3-fold than HPLC-photometric method, due to many interfering molecules in the cuvette, (2) immunoglobulin G (IgG) has unexpectedly wide binding abilities to many molecules; i.e., I have found that avidin binds biotin/lipoic acid/amino acids, and (3) binding reaction is depended on monomer-multimer states of binding protein (please see file; The Fascio effect).

If vitamin B12 concentration of serum has been determined by using reliable HPLC-photometric method, you can add it at a similar concentration. Please do not use the values of notorious HPLC-MS method, since MS is unexpectedly a non-quantitative detector at all (my unpublished experience, and the personal communication from a kind technical expert of Shimadzu Co., Kyoto, Japan).

By the way, lipoic acid (thioctic acid) has been successfully added to human cells, but it killed the rat cells (our unpublished observation at Gunma University, Maebashi, Gunma, Japan). Rat can biosynthesize lipoic acid, and lipoic acid is not the vitamin of rat. On the other hand, humans can not synthesize lipoic acid, and lipoic acid is surely vitamin of humans.

Similarly, vitamin C (ascorbic acid) is vitamin or not biosynthesized in humans and guinea pigs, but other rodents can synthesize ascorbate.

Therefore, it is an important issue that the species differences are usual in vitamins among animals (please see the file; Wide Range of Biotin).

C1 x V1 = C2 x V2