FluxMPS™ Ham's F-12 Nutrient Mixture [2X] Family

verified Double-Strength Ham's F-12 · Developed by R.G. Ham, University of Colorado, 1965

FluxMPS™ Ham's F-12 Nutrient Mixture [2X] Family

Ham's F-12 nutrient mixture is a widely utilized basal medium for culturing a variety of mammalian cells, including Chinese Hamster Ovary (CHO) cells, cancer cells, primary cells, and chicken embryonic cells. Developed by R.G. Ham in 1965 as an enhancement over the earlier F-10 nutrient mixture, it contains a more comprehensive array of components such as zinc, putrescine, hypoxanthine, and thymidine compared to other basal media like DMEM. Formulated for serum-free culture of single cells and clonal growth, Ham's F-12 can also be supplemented with serum for other cell types. While lacking proteins, lipids, and growth factors, it requires supplementation with serum or serum-free additives to facilitate optimal growth. The medium employs a sodium bicarbonate buffer system, necessitating a 5-10% CO₂ environment to maintain physiological pH. Common modifications to Ham's F-12 include the addition or removal of L-glutamine, phenol red, and HEPES buffer. A popular variant, DMEM/F12, combines DMEM and F-12 in equal parts, creating a more nutrient-rich medium. This [2X] family supplies the formulation at double strength — 3,603.2 mg/L glucose in the concentrate, 1,801.6 mg/L at 1X — for preparations where an aqueous matrix, supplement, or treatment solution would otherwise dilute the basal medium. Every FluxMPS™ variant is purified through quadruple-stage 0.04 micron filtration, engineered for organ-on-chip (OoC), tissue-on-chip (ToC), and lab-on-chip (LoC) microfluidic culture systems.

14 variants across L-Glutamine, Sodium Bicarbonate, and Phenol Red inclusion/exclusion, plus specialty formulations without Folic Acid, without Calcium Chloride, and with L-Alanyl-L-Glutamine (Sodium Pyruvate included in every variant; no variant contains HEPES)
Catalog numbers: DCP-H122X, DCP-H12-Q2X, DCP-H12-B2X, DCP-H12-R2X, DCP-H12-QB2X, DCP-H12-QR2X, DCP-H12-BR2X, DCP-H12-QBR2X, DCP-H12-F2X, DCP-H12-BF2X, DCP-H12-RF2X, DCP-H12-BRF2X, DCP-H12-C2X, DCP-H12YA2X
2X concentrate: mix with an equal volume of a compatible solution or matrix for a final 1X medium
3,603.2 mg/L D-glucose, 292 mg/L L-glutamine, and 220.2 mg/L sodium pyruvate in the 2X (half of each at 1X); sodium bicarbonate 2,352 mg/L where included (1,176 mg/L at 1X)
One of the first defined media to incorporate copper and zinc as trace elements, plus putrescine, hypoxanthine, and thymidine, absent from DMEM
Overall, a versatile basal medium suitable for serum-free and low-serum culture of various cell types, especially CHO cells
Applications stated on the 2X product pages: CHO growth and clonal culture, epithelial cells, chondrocytes, selected primary cells, in vitro clonal-toxicity assays, serum-reduced or serum-free medium development, and semisolid and matrix-culture development
The base formulation for the widely used DMEM/F-12 1:1 hybrid
Sodium bicarbonate buffering system requiring a 5-10% CO₂ environment
FluxMPS™ quadruple-stage 0.04 micron filtration — finer than any ready-to-use cell culture media currently available
Available in a 500 mL size; 2-8°C storage away from bright light, do not freeze; fully customizable on request

SKU: HamsF-12NutrientMixture2X — 14 VARIANTS

FluxMPS™ Ham's F-12 Nutrient Mixture — 2X Liquid Cell Culture Medium

Concentration2X (double strength)
Working strength1X after 1:1 mixing
Glucose3,603.2 mg/L at 2X (1,801.6 mg/L at 1X)
L-GlutamineConfigurable (+/-)
Sodium PyruvateIncluded (all variants)
Sodium Bicarbonate / Phenol RedConfigurable (+/-)
Folic Acid / Calcium ChlorideConfigurable (-, specialty variants)
HEPESNot included (on request)
FiltrationQuadruple-stage, 0.04 µm final
Size500 mL
Storage2-8 C, away from light; do not freeze

RUOHam 1965 Foundational2X ConcentrateCustomizable

Product Selector

Select Your Ham's F-12 [2X] Configuration

Please select the supplement(s) of interest, then click Search. Check the supplement(s) you need below and press Search to instantly highlight every matching variant. Each row links straight through to its product page via the catalog number or the View button. Rows marked with an italic note carry a specialty modification (No Folic Acid, No Calcium Chloride, or L-Alanyl-L-Glutamine) beyond the four standard supplement flags.

At-a-glance supplement matrix — 14 variants

At-a-glance supplement matrix — click to view product page. Sodium Pyruvate is included in every variant of this family. HEPES buffering is available as a customization on request but is not a standard catalog option.

Name	Cat No.	L-Glutamine	Pyruvate	Bicarbonate	Phenol Red	Product Page
Ham's F-12	DCP-H122X	check	check	check	check	Viewarrow_forward
Ham's F-12 w/o Glutamine	DCP-H12-Q2X	remove	check	check	check	Viewarrow_forward
Ham's F-12 w/o Bicarbonate	DCP-H12-B2X	check	check	remove	check	Viewarrow_forward
Ham's F-12 w/o Phenol Red	DCP-H12-R2X	check	check	check	remove	Viewarrow_forward
Ham's F-12 w/o Glutamine, Bicarbonate	DCP-H12-QB2X	remove	check	remove	check	Viewarrow_forward
Ham's F-12 w/o Glutamine, Phenol Red	DCP-H12-QR2X	remove	check	check	remove	Viewarrow_forward
Ham's F-12 w/o Bicarbonate, Phenol Red	DCP-H12-BR2X	check	check	remove	remove	Viewarrow_forward
Ham's F-12 w/o Glutamine, Bicarbonate, Phenol Red	DCP-H12-QBR2X	remove	check	remove	remove	Viewarrow_forward
Ham's F-12 w/o Folic Acid (No Folic Acid)	DCP-H12-F2X	check	check	check	check	Viewarrow_forward
Ham's F-12 w/o Bicarbonate, Folic Acid (No Folic Acid)	DCP-H12-BF2X	check	check	remove	check	Viewarrow_forward
Ham's F-12 w/o Phenol Red, Folic Acid (No Folic Acid)	DCP-H12-RF2X	check	check	check	remove	Viewarrow_forward
Ham's F-12 w/o Bicarbonate, Phenol Red, Folic Acid (No Folic Acid)	DCP-H12-BRF2X	check	check	remove	remove	Viewarrow_forward
Ham's F-12 w/o Calcium Chloride (No Calcium Chloride; 2X only)	DCP-H12-C2X	check	check	check	check	Viewarrow_forward
Ham's F-12 + L-Alanyl-L-Glutamine (Has L-Alanyl-L-Glutamine)	DCP-H12YA2X	check	check	check	check	Viewarrow_forward

check Included remove Not included All variants: 2X concentration · 3,603.2 mg/L Glucose · Sodium Pyruvate · no HEPES · 500 mL

Customization available: All variants above use the standard 2X concentration (mix 1:1 for 1X) and 3,603.2 mg/L Glucose (1,801.6 mg/L at 1X). Other concentrations, additions of chemicals, compounds, proteins, supplements (including HEPES buffer), a different pH, and modifications are available on request — contact support@diagnocine.com.

About Ham's F-12

About Ham's F-12 Nutrient Mixture

Ham's F-10 and Ham's F-12 are chemically defined, nutrient-rich cell culture media developed by Richard G. Ham at the University of Colorado to support the clonal growth of single Chinese hamster ovary (CHO) cells, an objective that required sustaining individual cells at very low densities under defined conditions, without reliance on serum or undefined biological extracts. They belong to a larger family of sequentially numbered formulations (the "F" series) developed by Ham in the late 1950s-1960s, each iteration tested against clonal growth efficiency as the quantitative readout.

Ham's F-12 (1965)

Ham, R.G. (1965). "Clonal growth of mammalian cells in a chemically defined, synthetic medium." Proceedings of the National Academy of Sciences USA, 53(2): 288-293. PMID: 14283412.^[2] F-12 was developed from F-10 (1963-1965) to achieve true protein-free clonal growth of CHO cells. In the 1965 PNAS paper, Ham demonstrated for the first time that single CHO cells could proliferate into clones in a fully chemically defined, protein-free medium.

F-12 is a direct development of F-10, retaining the same qualitative amino acid and vitamin profiles but with substantially altered concentrations of several components, most notably an approximately 30-fold higher zinc sulfate level, along with the addition of putrescine and linoleic acid, thereby rendering it effective for low-density, protein-free CHO culture. Both media share the same inorganic salt framework, the same 10-vitamin panel (including biotin and vitamin B12, which are absent from DMEM), the same nucleoside/nucleotide precursors (hypoxanthine and thymidine), and the same organic supplements (lipoic acid and sodium pyruvate).

F-12 became a medium of choice for myeloma and hybridoma cloning, primary rat hepatocytes, rat prostate epithelial cells, clonal toxicity assays, and the base of the widely used DMEM/F-12 hybrid. A tribute to Ham notes that F-12 became the starting medium for the systematic definition of individual growth-factor requirements across cell types, the platform from which the MCDB series grew.^[3,4]

Ham's F-12 Nutrient Mixture is a nutrient-rich basal medium containing amino acids, vitamins, inorganic salts, glucose, trace elements, and other components for in vitro mammalian-cell research; its formulation includes zinc, putrescine, hypoxanthine, thymidine, sodium pyruvate, and lipid-related constituents. Originally developed for serum-free single-cell plating and clonal growth of CHO cells, it is also used with selected epithelial cells, chondrocytes, and other laboratory cell systems when appropriately supplemented.^[2,8] It provides a basal nutritional foundation for cell-growth, colony-formation, nutrient-response, and research-medium development studies. It contains no added proteins or growth factors, so serum or defined supplements should be selected according to the cell type and validated protocol; its historical serum-free CHO application does not establish universal serum-free suitability.^[10]

Why a 2X (Double-Strength) Format

Ham's F-12 Nutrient Mixture, 2X, is a double-strength version of the corresponding 1X basal formulation, intended for research preparations that require substantial dilution. When mixed with an equal volume of a compatible solution that contributes none of the corresponding basal ingredients, it is designed to provide those ingredients at their intended final 1X concentrations. Every formulated component is supplied at twice its working level: D-glucose 3,603.2 mg/L (1,801.6 mg/L at 1X), L-glutamine 292 mg/L (146 mg/L at 1X), sodium pyruvate 220.2 mg/L (110.1 mg/L at 1X), and, where included, sodium bicarbonate 2,352 mg/L (1,176 mg/L at 1X). Variants that leave out L-glutamine, sodium bicarbonate, Phenol Red, folic acid, or calcium chloride let those components be added fresh or set to the level a protocol requires.

The concentrate may be evaluated for preparation of semisolid or hydrogel-containing research cultures where an aqueous matrix component would otherwise dilute the basal medium.^[9] The purpose is dilution compensation, not routine culture at twice the usual nutrient and salt concentrations.

Richard G. Ham and the Serum-Free Cloning Problem

By the late 1950s, growing a single mammalian cell into a clone essential for genetic analysis, mutagenesis, and selection of stable lines still depended on high serum concentrations or feeder cells. Ham recognized that the barrier was not a single missing factor but an incompletely defined nutrient environment whose inadequacies were masked by the serum's complex activity.

Working with near-diploid Chinese hamster (CHO) cells, the line initiated by Theodore T. Puck at the University of Colorado in 1957,^[8] Ham systematically developed a series of nutrient mixtures using single-cell plating efficiency (the fraction of individually plated cells that formed visible colonies) as his quantitative endpoint. Each formulation number marked an iteration in this optimization.

Ham's F-10 (1963) — The Direct Precursor

Ham, R.G. (1963). "An improved nutrient solution for diploid Chinese hamster and human cell lines." Experimental Cell Research, 29: 515-526. PMID: 13952250.^[1] F-10 enabled consistent single-cell cloning of Chinese hamster cells under low-serum conditions, and serum-free growth in the presence of defined proteins (serum albumin and fetuin). It was one of the first defined media to incorporate copper (CuSO₄) and zinc (ZnSO₄) as trace elements, and included hypoxanthine, thymidine, and lipoic acid. F-10 was also found to support human diploid cells, chromosome analysis of white blood cells, primary explants of rat, rabbit, and chicken tissues, and chick embryo cells.

Composition

Ham's F-12 [2X] Composition

Concentration scale: The 1X column gives the standard nutrient mixture F-12 (with L-glutamine; without sodium bicarbonate) at the working scale, from the Ham's F-12 family source (the source table nevertheless lists sodium bicarbonate at 1176 mg/L; kept as given). The 2X column gives the values listed on the 2X product pages for the concentrate as supplied; each component reaches the 1X value after 1:1 mixing. The complete DCP-H122X lists 47 components; each left-out component lowers that count by one.

All concentrations in mg/L. Users are advised to review the literature for recommendations regarding medium supplementation and physiological growth requirements specific for different cell lines. Per-lot Certificate of Analysis (CoA) requests can be directed to support@diagnocine.com.

Component Summary

Concentration	2X (double strength); 1X after 1:1 mixing
D-Glucose	3,603.2 mg/L at 2X (1,801.6 mg/L at 1X)
L-Glutamine	292 mg/L at 2X where included; L-Alanyl-L-Glutamine 434.4 mg/L in DCP-H12YA2X
Sodium Pyruvate	220.2 mg/L at 2X (all variants)
Buffer	NaHCO3 2,352 mg/L at 2X where included; no HEPES
Phenol Red	2.48 mg/L at 2X where included
Appearance	Red; Phenol Red-free variants pale yellow

Inorganic Salts

Salt	CAS No.	1X (mg/L)	2X as supplied (mg/L)
Calcium chloride dihydrate (CaCl₂ · 2H₂O) (not in DCP-H12-C2X)	10035-04-8	44.100	88.200
Copper sulfate pentahydrate (CuSO₄ · 5H₂O)	7758-99-8	0.0025	0.0050
Ferrous sulfate heptahydrate (FeSO₄ · 7H₂O)	7782-63-0	0.834	1.668
Magnesium chloride anhydrous (MgCl₂)	7786-30-3	57.650	115.300
Potassium chloride (KCl)	7447-40-7	223.600	447.200
Sodium chloride (NaCl)	7647-14-5	7599.000	15198.000
Sodium phosphate dibasic anhydrous (Na₂HPO₄)	7558-79-4	142.040	284.080
Zinc sulfate heptahydrate (ZnSO₄ · 7H₂O)	7446-20-0	0.863	1.726

F-12 uses only dibasic sodium phosphate (Na₂HPO₄), in contrast to F-10's dual KH₂PO₄ + Na₂HPO₄ system. F-10 uses MgSO₄ (anhydrous, 74.64 mg/L); F-12 uses MgCl₂ (anhydrous, 57.65 mg/L), reducing the sulfate load.

Amino Acids

Amino Acid	CAS No.	1X (mg/L)	2X as supplied (mg/L)
Glycine	56-40-6	7.500	15.000
L-Alanine	56-41-7	8.910	17.820
L-Arginine hydrochloride	1119-34-2	210.700	421.400
L-Asparagine monohydrate	5794-13-8	15.010	30.020
L-Aspartic acid	56-84-8	13.300	26.600
L-Cysteine hydrochloride monohydrate	7048-04-6	35.120	70.240
L-Glutamic acid	56-86-0	14.700	29.400
L-Glutamine (not in the w/o L-Glutamine variants; replaced by L-Alanyl-L-Glutamine in DCP-H12YA2X)	56-85-9	146.000	292.000
L-Alanyl-L-Glutamine (DCP-H12YA2X only; equimolar to the L-Glutamine above)	39537-23-0	217.2	434.4
L-Histidine hydrochloride monohydrate	5934-29-2	20.960	41.920
L-Isoleucine	73-32-5	3.940	7.880
L-Leucine	61-90-5	13.100	26.200
L-Lysine hydrochloride	657-27-2	36.500	73.000
L-Methionine	63-68-3	4.480	8.960
L-Phenylalanine	63-91-2	4.960	9.920
L-Proline	147-85-3	34.500	69.000
L-Serine	56-45-1	10.500	21.000
L-Threonine	72-19-5	11.900	23.800
L-Tryptophan	73-22-3	2.040	4.080
L-Tyrosine disodium salt dihydrate	122666-87-4	7.810	15.620
L-Valine	72-18-4	11.700	23.400

L-Glutamine in F-12 base: Unlike F-10, standard F-12 includes L-glutamine (146 mg/L = 1 mM at 1X) as part of the base formulation.

Vitamins

Vitamin	CAS No.	1X (mg/L)	2X as supplied (mg/L)
Biotin	58-85-5	0.0073	0.0146
Choline chloride	67-48-1	13.960	27.920
D-Ca-Pantothenate	137-08-6	0.480	0.960
Folic acid (not in the w/o Folic Acid variants)	59-30-3	1.320	2.640
Nicotinamide	98-92-0	0.037	0.074
Pyridoxine hydrochloride	58-56-0	0.062	0.124
Riboflavin	83-88-5	0.038	0.076
Thiamine hydrochloride	67-03-8	0.340	0.680
Vitamin B12	68-19-9	1.360	2.720
myo-Inositol	87-89-8	18.000	36.000

Vitamin B6 form: F-12 uses pyridoxine hydrochloride, as in high-glucose DMEM (Gibco), and in contrast to CMRL 1066 and GMEM, which use pyridoxal hydrochloride. Biotin and Vitamin B12: F-12 contains biotin and vitamin B12 — shared with CMRL 1066 but absent from DMEM, MEM, and GMEM.

Other Components

Component	CAS No.	1X (mg/L)	2X as supplied (mg/L)
D-Glucose	50-99-7	1801.600	3603.200
Hypoxanthine sodium salt	45738-97-4	4.770	9.540
Linoleic acid	60-33-3	0.084	0.168
Lipoic acid	1077-28-7	0.210	0.420
Phenol red sodium salt (not in the w/o Phenol Red variants)	34487-61-1	1.240	2.480
Putrescine dihydrochloride	333-93-7	0.161	0.322
Sodium pyruvate	113-24-6	110.100	220.200
Thymidine	50-89-5	0.730	1.460
Sodium bicarbonate (NaHCO₃) (not in the w/o Sodium Bicarbonate variants)	144-55-8	1176.000	2352.000

Background

Key Structural Differences: F-10 to F-12

Context on the biochemical rationale behind the F-10-to-F-12 evolution that underlies the Coon's Modification base formulation.

The zinc difference is the defining feature. The approximately 30-fold increase in ZnSO₄ · 7H₂O from F-10 (0.029) to F-12 (0.863 mg/L) was the single most important change enabling protein-free CHO cloning. Zinc is required for numerous metalloenzymes, receptors, and transcription factors; at very low cell densities, the per-volume zinc requirement is not met by trace contamination, so it must be supplied at a defined, elevated concentration.

Putrescine supports polyamine-dependent proliferation. Putrescine (0.161 mg/L) is the diamine precursor to spermidine and spermine, essential for DNA synthesis, ribosome function, and cell division, partially bypassing the polyamine supply that serum normally provides.

Linoleic acid is an essential fatty acid. Linoleic acid (0.084 mg/L, free fatty acid) supplies the essential omega-6 fatty acid for membrane phospholipid synthesis that serum albumin-bound fatty acids would otherwise provide; cells cannot synthesize it de novo.

myo-Inositol is 33x higher in F-12 (18.0 vs 0.541 mg/L). Inositol is a component of phosphatidylinositol, a key membrane phospholipid and signaling intermediate whose requirement rises sharply in single-cell cultures.

Choline is 20x higher in F-12 (13.96 vs 0.698 mg/L), supporting phosphatidylcholine synthesis and the higher membrane biosynthetic demand of rapidly proliferating single-cell clones under protein-free conditions.^[5,6]

Related Formulation

Comparison to Ham's F-10 (The Direct Precursor)

Ham's F-10 is provided here for reference and comparison; it is a distinct, separately formulated product from the F-12 family sold above. All values in this comparison are at 1X working strength.

Side-by-Side Composition Comparison

Salt	F-10 (mg/L)	F-12 (mg/L)	Notes
CaCl₂ · 2H₂O	44.100	44.100	Identical
CuSO₄ · 5H₂O	0.0025	0.0025	Identical
FeSO₄ · 7H₂O (ferrous)	0.834	0.834	Identical
MgSO₄ anhydrous	74.640	—	F-10 only
MgCl₂ anhydrous	—	57.650	F-12 only; Mg source change
KCl	285.000	223.600	Lower in F-12
KH₂PO₄	83.000	—	F-10 only
NaCl	7400.000	7599.000	Slightly higher in F-12
Na₂HPO₄	153.700	142.040	Both are slightly lower in F-12
ZnSO₄ · 7H₂O	0.029	0.863	Approximately 30x higher in F-12 — critical difference

Amino Acid	F-10 (mg/L)	F-12 (mg/L)	Change
Glycine	7.510	7.500	Approximately same
L-Alanine	8.910	8.910	Identical
L-Arginine HCl	211.000	210.700	Approximately same
L-Asparagine · H₂O	15.010	15.010	Identical
L-Aspartic acid	13.300	13.300	Identical
L-Cysteine · HCl · H₂O	35.130	35.120	Approximately same
L-Glutamic acid	14.700	14.700	Identical
L-Glutamine	(add separately)	146.000	Included in F-12 base
L-Histidine · HCl · H₂O	21.000	20.960	Approximately same
L-Isoleucine	2.600	3.940	Higher in F-12 (+52%)
L-Leucine	13.100	13.100	Identical
L-Lysine · HCl	29.300	36.500	Higher in F-12 (+25%)
L-Methionine	4.480	4.480	Identical
L-Phenylalanine	4.960	4.960	Identical
L-Proline	11.500	34.500	3x higher in F-12
L-Serine	10.500	10.500	Identical
L-Threonine	3.570	11.900	3.3x higher in F-12
L-Tryptophan	0.600	2.040	3.4x higher in F-12
L-Tyrosine	2.610	7.810	3x higher in F-12
L-Valine	3.500	11.700	3.3x higher in F-12

Vitamin	F-10 (mg/L)	F-12 (mg/L)	Change
Biotin	0.024	0.0073	Lower in F-12
Choline chloride	0.698	13.960	20x higher in F-12
D-Ca-Pantothenate	0.715	0.480	Lower in F-12
Folic acid	1.320	1.320	Identical
Nicotinamide	0.615	0.037	Lower in F-12
Pyridoxine · HCl	0.206	0.062	Lower in F-12
Riboflavin	0.376	0.038	Lower in F-12
Thiamine · HCl	1.000	0.340	Lower in F-12
Vitamin B12	1.360	1.360	Identical
myo-Inositol	0.541	18.000	33x higher in F-12

Vitamin B6 form: Both F-10 and F-12 use pyridoxine hydrochloride, as in high-glucose DMEM (Gibco), and in contrast to CMRL 1066 and GMEM, which use pyridoxal hydrochloride. Biotin and Vitamin B12: Both F-10 and F-12 contain biotin and vitamin B12 — shared with CMRL 1066 but absent from DMEM, MEM, and GMEM.

Component	F-10 (mg/L)	F-12 (mg/L)	Change
D-Glucose	1100.000	1801.600	Higher in F-12 (approximately 10 mM vs approximately 6 mM)
Hypoxanthine (Na salt)	4.080	4.770	Approximately same
Lipoic acid	0.210	0.210	Identical
Linoleic acid	—	0.084	F-12 only
Putrescine · 2HCl	—	0.161	F-12 only
Sodium pyruvate	110.000	110.100	Approximately same
Thymidine	0.730	0.730	Identical
NaHCO₃	1200.000	1176.000	Approximately same
Phenol red Na salt	1.300	1.240	Approximately same

Comparison

Media Lineage Comparison

Values in this lineage table refer to 1X working strength.

Feature	Ham's F-12 (2X family, at 1X)	Ham's F-10	DMEM (High Glucose)
Developer & year	R.G. Ham, 1965	R.G. Ham, 1963	Dulbecco & Freeman, 1959
Parent formulation	F-10 (direct)	Earlier "F" series	BME
Trace metals (Cu, Zn, Fe)	Cu + Zn + Fe	Cu + Zn + Fe	Fe only (ferric nitrate)
Hypoxanthine	Yes, 4.77 mg/L	Yes, 4.08 mg/L	No
Thymidine	Yes, 0.73 mg/L	Yes, 0.73 mg/L	No
Lipoic acid	Yes, 0.21 mg/L	Yes, 0.21 mg/L	No
Putrescine	Yes, 0.161 mg/L	No	No
Linoleic acid	Yes, 0.084 mg/L	No	No
Biotin	Yes, 0.0073 mg/L	Yes, 0.024 mg/L	No
Vitamin B12	Yes, 1.36 mg/L	Yes, 1.36 mg/L	No
Vitamin B6 form	Pyridoxine HCl	Pyridoxine HCl	Pyridoxine HCl (Gibco)
myo-Inositol	18.0 mg/L	0.541 mg/L	Present (lower)
Glucose	1801.6 mg/L (approximately 10 mM)	1100 mg/L (approximately 6.1 mM)	4500 mg/L (25 mM)
Sodium pyruvate	110 mg/L	110 mg/L	110 mg/L (optional)
NaHCO₃	1176 mg/L	1200 mg/L	3700 mg/L
Serum-free design	Full (protein-free CHO)	Partial (with proteins)	No
Osmolality (with NaHCO₃)	260-300 mOsm/kg	300-340 mOsm/kg	approximately 320-355 mOsm/kg

Ham's F-12: 1X vs. 2X

Feature	Ham's F-12, 1X	Ham's F-12, 2X
Supplied concentration	1X (ready to use)	2X (double strength)
Preparation before use	None	Mix with an equal volume of a compatible solution or matrix (aqueous agar, agarose, or hydrogel component, supplements, or treatment solutions) for a final 1X medium
Component concentrations as supplied	As listed in the 1X column of the composition tables	Twice the 1X value (2X column of the composition tables), before dilution
Composition at working strength	Standard F-12 formulation	Standard F-12 formulation, after 1:1 dilution
Glucose	1801.6 mg/L	3,603.2 mg/L (1,801.6 mg/L after 1:1 dilution)
L-Glutamine (where included)	146 mg/L	292 mg/L (146 mg/L after 1:1 dilution)
Sodium bicarbonate (where included)	1176 mg/L	2,352 mg/L (1,176 mg/L after 1:1 dilution)
Catalog variants	13	14 (adds a 2X-only variant without Calcium Chloride, DCP-H12-C2X)
Catalog number suffix	...1X	...2X (e.g., DCP-H122X)
Filtration	Quadruple-stage, 0.04 micron final	Quadruple-stage, 0.04 micron final
Sizes	500 mL / 1000 mL	500 mL
Storage	2-8°C, away from bright light	2-8°C, away from bright light; do not freeze

Validated Applications

Validated Cell Types and Applications

Ham's F-12

CHO Cells

Primary application; protein-free clonal growth; serum-free bioproduction standard. Ham used CHO cells from the Puck line; the widely used proline-requiring subclone is CHO-K1.

Mouse L Cells

Serum-supplemented maintenance.

HeLa Cells

Serum-free culture with defined hormones (insulin, transferrin, hydrocortisone, EGF, FGF).

Myeloma & Hybridoma Cells

A medium of choice for cloning; widely used in monoclonal antibody work (Coon's modification of F-12 was developed for virally fused hybrid cells).

Primary Rat Hepatocytes

F-12 + supplements are a standard.

Rat Prostate Epithelial Cells

F-12-based defined systems.

Normal Rat Kidney (NRK) Cells

Supported with appropriate supplementation.

Clonal Toxicity Assays

CHO cells in F-12 at single-cell plating density.

Cancer Cells & Chicken Embryonic Cells

Ham's F-12 is also used for culturing cancer cells and chicken embryonic cells more broadly.

Ham's F-12 [2X] — Research Applications

Cell Lines

CHO Growth & Clonal Culture

Developed for serum-free single-cell plating and clonal growth of Chinese hamster ovary cells; a reference basal medium for CHO research.^[2,8]

CHOSingle-cell platingClonal growth

Cell Biology

Epithelial Cells & Chondrocytes

Used with selected epithelial cells, chondrocytes, and other primary cells when appropriately supplemented after dilution to 1X.^[2]

EpithelialChondrocytesPrimary cells

Toxicology

Clonal-Toxicity Assays

Colony-formation readouts in a clonal-growth medium support in vitro clonal-toxicity and nutrient-response studies.^[2]

Colony formationClonal toxicityNutrient response

Media Development

Serum-Reduced & Semisolid Media

A double-strength base for serum-reduced or serum-free medium development and for semisolid or hydrogel cultures.^[9,10]

Serum-freeHydrogelsSoft agar

Microfluidics

Organ-on-a-Chip & MPS

Microchannel-safe medium for perfused organ-, tissue-, and body-on-a-chip platforms.^[13,14,15]

OoCToCMPS

Live-Cell Imaging

Microscopy & Optical Sensing

Fewer particles mean cleaner fields for colony counting, confocal imaging, and on-chip biosensors; the Phenol Red-free variants add no Phenol Red background.

ConfocalColony countingBiosensors

Specialty-formulation uses (from the 2X product pages): Folic acid-free versions give a folate-free base for folate-receptor uptake studies, antifolate drug research, and folate-depletion experiments; add a folate source back if long-term growth requires it. The calcium chloride-free version gives a calcium-free base for calcium-switch and cell-junction assays, gentle cell dissociation, suspension adaptation, and suspension-culture studies; add calcium back at the levels your protocol requires. In the L-alanyl-L-glutamine version the dipeptide replaces free L-glutamine and resists breakdown in liquid medium, so less ammonia builds up and the glutamine supply holds over long cultures, extended perfusion runs, and slow-growing 3D or semisolid cultures.

Shared Role: Base for Defined Media Development

F-12 is the base for the DMEM/F-12 1:1 hybrid (combining DMEM's high amino acid and glucose content with F-12's trace elements, lipids, and low osmolality), one of the most widely used media in serum-free and stem cell culture. It is also the starting formulation from which Ham and colleagues derived the MCDB series for specific primary cell types.

Ham's F-10 (Precursor, Reference)

F-10 is validated for CHO/Chinese hamster cells (founding application; clonal growth at low serum), human diploid cells, white blood cell chromosome analysis, primary rat/rabbit/chicken explants, chick embryo cells, mouse L cells, HeLa cells, and mammalian embryo culture (bovine, equine, porcine, human), often as F-10 + HEPES.

Why FluxMPS™

Why FluxMPS™ Ham's F-12 Nutrient Mixture [2X]

filter_alt

Quadruple-Stage 0.04 Micron Filtration

Purified to 0.04 microns — finer than any ready-to-use cell culture media currently available — removing the microscopic particulates and protein aggregates that silently block micro-channels.

device_hub

Built for OoC / ToC / LoC Platforms

Engineered from the ground up for Organ-on-Chip, Tissue-on-Chip, and Lab-on-Chip platforms, where the medium itself is part of the instrument and cannot be an afterthought.

water_drop

Particulate & Aggregate Removal

Eliminates particulates and protein aggregates that disrupt laminar flow and generate false biological signals before the media ever reaches the chip.

visibility

Optical Clarity

Supports real-time imaging and integrated biosensing on Organ-on-Chip platforms, where optical clarity of the medium is essential to data quality.

verified_user

FDA-Recognized Physiological Modeling Standards

Formulated to support FDA-recognized physiological modeling standards, providing a validated, reproducible media foundation for regulatory-facing studies.

science

Drug Discovery, Toxicology & Translational Research

A validated foundation for drug discovery, toxicology screening, and translational research, where downstream data must stand up to regulatory scrutiny.

Filtration Technology

Quadruple-Stage Filtration System

Every FluxMPS™ Ham's F-12 Nutrient Mixture [2X] variant passes through the same four-stage architecture before reaching your chip: two 0.1 micron pre-filtration stages followed by two 0.04 micron final filtration stages.

01

Pre-Filtration Stage 1 0.1 µm

Initial coarse particulate removal.
02

Pre-Filtration Stage 2 0.1 µm

Secondary particulate and aggregate reduction.
03

Final Filtration Stage 1 0.04 µm

Fine filtration below standard 0.22 micron practice.
04

Final Filtration Stage 2 0.04 µm

Final polish for microfluidic-grade clarity.

Engineered for Flow, Not Just Growth

Every component is optimized for consistent, laminar flow performance across complex micro-channel geometries, capillary-bed and vascular simulations, and long-term automated perfusion studies running continuously for weeks.

0.04µm

Final polish

4

Total filtration stages

FluxMPS™ Ham's F-12 Nutrient Mixture [2X] Family quadruple-stage filtration system diagram ? two 0.1 micron pre-filtration stages followed by two 0.04 micron final filtration stages, engineered for organ-on-a-chip (OoC), tissue-on-a-chip (ToC), and lab-on-a-chip microfluidic cell culture media applications by Diagnocine.

FAQ

Frequently Asked Questions

The family covers combinations of L-Glutamine, Sodium Bicarbonate, and Phenol Red, plus specialty formulations without Folic Acid, without Calcium Chloride, and with L-Alanyl-L-Glutamine; Sodium Pyruvate is included in every variant, and no variant contains HEPES. Use the supplement matrix above: check the box for each supplement your protocol requires, press Search, and the matching row (or rows) will highlight in mint green so you can click straight through via the catalog number link or the View button.

Mix 2X Ham's F-12 with an equal volume of a compatible solution that contributes none of the basal ingredients — an aqueous agar, agarose, or hydrogel component, supplements, or treatment solutions — so glucose (3,603.2 mg/L to 1,801.6 mg/L), amino acids, vitamins, and salts end up at their 1X concentration.^[9] After 1:1 mixing every component falls to half its supplied level: L-glutamine 292 mg/L to 146 mg/L, sodium pyruvate 220.2 mg/L to 110.1 mg/L, and sodium bicarbonate 2,352 mg/L to 1,176 mg/L where included. The purpose is dilution compensation, not routine culture at twice the usual nutrient and salt concentrations. Serum, antibiotics, and other supplements can be added before or after filter sterilization, observing sterility precautions.

Ham's F-12 is a widely utilized basal medium for culturing Chinese Hamster Ovary (CHO) cells, cancer cells, primary cells, chicken embryonic cells, mouse L cells, HeLa cells, myeloma and hybridoma cells, primary rat hepatocytes, rat prostate epithelial cells, and normal rat kidney (NRK) cells. The 2X product pages also list selected epithelial cells and chondrocytes, in vitro clonal-toxicity assays, serum-reduced or serum-free medium development, and semisolid and matrix-culture development.

F-12 (1965) is a direct development of F-10 (1963), developed to achieve true protein-free clonal growth of CHO cells. The key differences are an approximately 30-fold higher zinc sulfate concentration, added putrescine and linoleic acid (absent from F-10), and large increases in several amino acids and two vitamins (choline, myo-inositol). See the Comparison to Ham's F-10 section above for the full side-by-side breakdown.

In addition to the standard supplement configuration matrix, this family includes four variants without Folic Acid (combined with Bicarbonate and Phenol Red options), one variant without Calcium Chloride, and one variant formulated with L-Alanyl-L-Glutamine, a stable glutamine dipeptide. Each is identified by an italic note in the matrix Name column and carries its own catalog number and product page. Folic acid is left out so you can control folate exposure yourself — for example in folate-receptor uptake studies, antifolate drug work, or folate-depletion experiments where the 2.64 mg/L (1.32 mg/L at 1X) in standard 2X Ham's F-12 would compete or mask the effect; add folic acid or another folate back if your cells need it. Calcium chloride is left out to give a calcium-free base for calcium-switch, cell-dissociation, cadherin-junction, and suspension-culture work; standard 2X Ham's F-12 carries 88.2 mg/L calcium chloride dihydrate (44.1 mg/L, about 0.3 mM, at 1X). DCP-H12YA2X carries 434.4 mg/L L-alanyl-L-glutamine (217.2 mg/L at 1X), equimolar to the 292 mg/L L-glutamine in standard 2X Ham's F-12; cells cleave the dipeptide to release L-glutamine as they need it.

L-glutamine breaks down in liquid medium over time, releasing ammonia that can stress sensitive cultures; a glutamine-free base lets you add fresh L-glutamine or a stable dipeptide just before use. Our complete 2X formulation carries 292 mg/L L-glutamine (146 mg/L at 1X). Phenol Red is left out because it adds background in fluorescence and absorbance readouts and has weak estrogen-like activity that can confound hormone-sensitive work; track pH with a meter or indicator strip.^[12] Phenol Red-free variants are pale yellow; the others are red.

Formulated for serum-free culture of single cells and clonal growth, Ham's F-12 can also be supplemented with serum for other cell types. While lacking proteins, lipids, and growth factors, it requires supplementation with serum or serum-free additives to facilitate optimal growth. Its historical serum-free CHO use does not establish universal serum-free suitability.^[2,10]

The medium employs a sodium bicarbonate buffer system, necessitating a 5-10% CO₂ environment. For the bicarbonate-containing 2X variants, use a CO₂-controlled incubator once diluted to 1X (1,176 mg/L sodium bicarbonate at 1X; classically equilibrated with 10% CO₂, commonly 5%) and keep time in room air short. For the bicarbonate-free variants it depends on the buffer you add: with sodium bicarbonate, use a CO₂-controlled incubator; with a non-volatile buffer such as HEPES, CO₂ is not needed for pH control. Leaving sodium bicarbonate out lets the solution or matrix you mix with the 2X carry the buffer, or lets you set your own bicarbonate level for your CO₂ tension.

DMEM/F-12 is a popular variant that combines DMEM and F-12 in equal parts, creating a more nutrient-rich medium. F-12 is the base for this widely used 1:1 hybrid, combining DMEM's high amino acid and glucose content with F-12's trace elements, lipids, and low osmolality.

HEPES is not a standard catalog option for this family; no 2X variant contains HEPES, which keeps it out of HEPES-sensitive assays. Common modifications to Ham's F-12 include the addition or removal of HEPES buffer, and it is available as a customization on request via support@diagnocine.com.

Yes. Every variant is quadruple-stage filtered to a 0.04 micron final pore size, which lowers the risk of clogging microchannels, valves, and membranes in organ-on-a-chip, hydrogel-based 3D, and microfluidic perfusion systems.^[13,14,15]

Yes. Standard concentration for this family is 2X, with 3,603.2 mg/L glucose (1,801.6 mg/L at 1X). Other concentrations, additions of chemicals, compounds, proteins, supplements, a different pH, and modifications are available on request via support@diagnocine.com. A 10 nm (0.01 micron) ultra-filtered grade for automated bioreactors, liquid-handling robots, and closed perfusion loops is also made on request — email your catalog number and volume.

All 2X variants are supplied in a 500 mL size. Store at 2-8°C, away from bright light; do not freeze. A lot-specific Certificate of Analysis is available on request at support@diagnocine.com.

References

Verified Bibliography

Ham, R.G. (1963). An improved nutrient solution for diploid Chinese hamster and human cell lines. Experimental Cell Research, 29: 515-526. PMID: 13952250. DOI: 10.1016/S0014-4827(63)80014-2
Ham, R.G. (1965). Clonal growth of mammalian cells in a chemically defined, synthetic medium. Proceedings of the National Academy of Sciences USA, 53(2): 288-293. PMID: 14283412. DOI: 10.1073/pnas.53.2.288
Ham, R.G. & McKeehan, W.L. (1979). Media and growth requirements. Methods in Enzymology, 58: 44-93. DOI: 10.1016/S0076-6879(79)58127-2
Barnes, D. & Sato, G. (1980). Serum-free cell culture: a unifying approach. Cell, 22(3): 649-655. DOI: 10.1016/0092-8674(80)90540-1
McKeehan, W.L., Hamilton, W.G. & Ham, R.G. (1976). Selenium is an essential trace nutrient for the growth of WI-38 diploid human fibroblasts. Proc. Natl. Acad. Sci. USA, 73(6): 2023-2027. DOI: 10.1073/pnas.73.6.2023
Yao, T. & Asayama, Y. (2017). Animal-cell culture media: History, characteristics, and current issues. Reproductive Medicine and Biology, 16(2): 99-117. PMC5661806
Eagle, H. (1955). Nutrition needs of mammalian cells in tissue culture. Science, 122(3168): 501-504. DOI: 10.1126/science.122.3168.501
Tjio, J.H. & Puck, T.T. (1958). Genetics of somatic mammalian cells. II. Chromosomal constitution of cells in tissue culture. Journal of Experimental Medicine, 108(2): 259-268. DOI: 10.1084/jem.108.2.259
Borowicz, S., Van Scoyk, M., Avasarala, S., et al. (2014). The soft agar colony formation assay. Journal of Visualized Experiments, (92): e51998. DOI: 10.3791/51998
Barnes, D. & Sato, G. (1980). Methods for growth of cultured cells in serum-free medium. Analytical Biochemistry, 102(2): 255-270. DOI: 10.1016/0003-2697(80)90151-7
Morton, H.J. (1970). A survey of commercially available tissue culture media. In Vitro, 6(2): 89-108. DOI: 10.1007/BF02616111
Berthois, Y., Katzenellenbogen, J.A. & Katzenellenbogen, B.S. (1986). Phenol red in tissue culture media is a weak estrogen: implications concerning the study of estrogen-responsive cells in culture. Proc. Natl. Acad. Sci. USA, 83(8): 2496-2500. DOI: 10.1073/pnas.83.8.2496
Bhatia, S.N. & Ingber, D.E. (2014). Microfluidic organs-on-chips. Nature Biotechnology, 32(8): 760-772. DOI: 10.1038/nbt.2989
Low, L.A., Mummery, C., Berridge, B.R., Austin, C.P. & Tagle, D.A. (2021). Organs-on-chips: into the next decade. Nature Reviews Drug Discovery, 20(5): 345-361. DOI: 10.1038/s41573-020-0079-3
Leung, C.M., de Haan, P., Ronaldson-Bouchard, K., et al. (2022). A guide to the organ-on-a-chip. Nature Reviews Methods Primers, 2: 33. DOI: 10.1038/s43586-022-00118-6
Drexler, H.G. & Uphoff, C.C. (2002). Mycoplasma contamination of cell cultures: incidence, sources, effects, detection, elimination, prevention. Cytotechnology, 39(2): 75-90. DOI: 10.1023/A:1022913015916
Halldorsson, S., Lucumi, E., Gómez-Sjöberg, R. & Fleming, R.M.T. (2015). Advantages and challenges of microfluidic cell culture in polydimethylsiloxane devices. Biosensors and Bioelectronics, 63: 218-231. DOI: 10.1016/j.bios.2014.07.029

References [1]-[6] are the Ham's F-12 family bibliography; references [7]-[17] are cited on the 2X product pages.

FluxMPS™ Platform

FluxMPS™ — Precision Cell Culture Media for Microphysiological Systems

Built for the architecture of the future. Not the flask of the past.

Traditional cell culture media were formulated for static well plates and flasks — environments that tolerate impurities, precipitates, and particle loads that would immediately compromise a microfluidic system. FluxMPS™ was designed from the ground up for Organ-on-Chip (OoC), Tissue-on-Chip (ToC), and Lab-on-Chip (LoC) platforms, where the medium itself is part of the instrument.

Purity That Protects Your Platform

FluxMPS™ is purified to 0.04 microns — finer than any ready-to-use cell culture media currently available. At this level, the microscopic particulates and protein aggregates that silently block micro-channels, disrupt laminar flow, and generate false biological signals are eliminated before the media ever reaches your chip.

The result: your platform stays operational, your data stays clean, and your biology drives the result — not your media.

Engineered for Flow, Not Just Growth

The name FluxMPS™ reflects its core design principle. Every component is optimized for consistent, laminar flow performance across:

Complex micro-channel geometries
Capillary-bed and vascular simulations
Long-term automated perfusion studies running continuously for weeks

Zero-clogging performance is not a feature — it is the baseline specification.

Applications & Performance

Application	What FluxMPS™ Delivers
Microfluidics	Stable shear stress; no channel blockage
Metabolic Tracing	Ultra-pure matrix with no contaminant interference
Long-term Perfusion	Consistent formulation stability over weeks of continuous flow
Organ-on-Chip	Optical clarity for real-time imaging and integrated biosensing

Regulatory Foundation

FluxMPS™ is formulated to support FDA-recognized physiological modeling standards, providing a validated, reproducible media foundation for drug discovery, toxicology screening, and translational research. When your downstream data needs to stand up to regulatory scrutiny, your upstream media cannot be an afterthought.

The Bottom Line

Microfluidic platforms are precision instruments. They require precision inputs.

FluxMPS™ is the only ready-to-use cell culture medium engineered specifically to meet that standard — protecting your chip, your cells, and your science.

FluxMPS™ Ham's F-12 Nutrient Mixture [2X] Family

Select See the table below