Ham’s F12 medium

Media frequently employed in the culture of continuous mammalian cell lines include Eagle s medium, MEM (Eagle, 1959) Eagle s medium modified by Dulbecco, DMEM (Dulbecco and Freeman, 1959) RPMI 1640 medium (Moore et ah, 1967) CMRL 1066 medium (Parker et al., 1957) and Ham s F12 medium (Ham, 1965). For the cultivation of adherent continuous cell lines the following basal media are suitable CMRL 1066, MCDB 411, DMEM. F12, MCDB 301, and IMDM. For non-transformed cells, the media DMEM, IMDM, MCDB 104, 105, 202, 401, and 501 are suitable (Freshney, 1992). Each of these basal formulations may be supplemented with serum or other specific proteins. 

The AR42J cell line expressing somatostatin receptor subtype 2 was obtained from ATCC (Manassas, VA, USA). The cells were maintained in Kaighn s modification of Ham s F12 medium (F12K) with 2mM L-glutamine, 10% foetal bovine serum and 1.5 g/L of sodium bicarbonate in a humidified atmosphere at 5% CO2 and 37°C. 

Cell seeding. Human fetal osteoblasts (hFOB 1.19, CRL-11372, American Type Culture Collection) were seeded at an initial density of 1 x 104 cells cm 2 in a medium containing a 1 1 mixture of Dulbecco s Modified Eagle s Medium (DMEM) without phenol red and Ham s F12 medium and supplemented with 0.3 mg/ml G418, antibiotics (100 U/ml penicillin and 100 pg/ml streptomycin) and 10% foetal bovine serum. Cells were grown for 48 h at 37°C in an humidified atmosphere of 5% C02 on the cell culture plastic supports (control), and sucrose thin films deposited on glass substrates (SI substrate). Prior to cell culture, the samples were sterilized by UV light exposure. 

For the cultivation of airway epithelial cells, the cells are usually plated onto tissue culture plates coated with collagen or extracellular matrix components. Hormonally defined Ham s F12 medium contained 1 % penicillin-streptomycin, 5 ig/ml insulin, 5 ig/ml transferrin, 25 ng/ml epidermal growth factor, 15 ig/ml epithehal growth factor supplement, 2 X 10" M triiodothyronin, and 10" M hydrocortisone. 

To induce and maintain differentiated morphology, in a double chamber culture system described by Clark etal. (1995) bronchial epithelial cells were plated on the upper chamber of Trans Well culture plates in hormonally supplemented Ham s F12 medium. After 7 days the epithelial cells showed a ciliated differentiation (Takizawa 1999). 

A range of conventional media can be used but the commonest choices are Dulbecco s MEM (DMEM), Iscove s modified DMEM (IMDM), RPMI-1640, or a 1 1 mixture of DMEM and Ham s F12 medium. 

Ham s F10 medium was very carefully formulated for cloning diploid hamster ovary cells (Ham, 1963) and was later modified — Ham s F12 (Appendix 1 Table 16) (Ham, 1965) — to support... 

To prepare the agarose cell cultures, 24 well plates are coated with 0.2 ml/well of a 1 1 mixture of the 2 % agarose solution with preheated Ham s F12 and left at room temperature to gel. Then 0.1 ml/well (0.2 ml/well for radiolabeling) of a 1 1 mixture of the above described cell suspension with the 2 % agarose solution is added. Care has to be taken to maintain an even cell suspension, and not to overheat the cells during this procedure. After gel formation at room temperature, the multiwell plates are placed in the incubator, and 0.5 ml/well medium is added either 4 h later or the following day. The medium consists of Ham s F12 supplemented with 5 % FCS and 25 xg/ml ascorbic acid, and is changed every second day. 

Fetlocks of freshly slaughtered steers (age 18 to 20 months) are skinned and the metacarpo-phalangeal joint opened under semi-sterile conditions. With a sterile punch (as used for obtaining skin biopsies) full thickness disks of cartilage are obtained from all accessible cartilaginous areas and their wet weight is assessed. In each well, 1 ml of medium is added, consisting of Ham s F12 supplemented with 10 % FCS... 

RPMI-1640 (DM) Dulbecco s modification of Eagle s medium (F12 and F12K) Ham s 1 medium and Kaighn s ... 

As an example, the main compounds of a 1 1 mixture of IMDM (Iscove s Modification of Dulbecco s Medium) and Ham s F12 are given as follows ... 

Ham s medium F12 has been designed specifically for cloning cells (Ham, 1965) and should be supplemented with 10-30% foetal bovine serum. Great care must be taken to prevent the medium becoming too alkaline by loss of C02. 

Use the type of medium the cells have been growing in as control. Test 3-4 commercially available balanced media, such as F12/DMEM (Ham s nutrient mixture/Dulbecco s modified Eagle s medium), DMEM, IMDM (Iscove s modified Dulbecco s medium), HB-CHO (Hana Biological Chinese hamster ovary medium) or RPMI-1640 (Roswell Park Memorial Institute). 

Vanadium compounds have a toxic effect on pneumocytes similar to chromates. Although generally less soluble the vanadium compounds studies by Popper et al. (1992) were ingested or phagocytosed by rat pneumocytes II cultured as monolayers in Ham s medium F12. V2O5 was the most toxic, while VO2 was the least toxic oxide. Superoxide dismut-ase, reduced glutathione, EDTA, and DMSO when added to the culture medium had no influence on the survival of the cultured pneumocytes, while... 

Growth medium, Dulbecco s Modified Eagles Medium/Hams F12, supplemented with 10% fetal calf serum and glutamine, penicillin, streptomycin, and fungizone. 

Linoleic acid and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) were obtained from Sigma Chemical Co. (St. Louis, MO). Dulbeccos Modified Eagle s Medium/Hams F-12 nutrient mixture (DMEM/F12) containing 2 pM linoleic acid, fetal bovine serum (FBS), trypsin-EDTA, penicillin-streptomycin, selenium and transferrin were purchased from Gibco BRL (Rockville, MD). Other chemicals used were of reagent grade. 

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