Chromatography

CFT ™ Ceramic Fluoroapatite MediaCFT Type II is available in a 40 μm particle size
and is sintered at high temperatures to produce
physically and chemically stable bioprocess
media. CFT Type II maintains its highest IgGbinding capacity (34–37 mg/ml) at pH 5.0–6.0(Figure 1) and over a range of flow rates (Figure 2).
When process requirements call for bufferedconditions at low pH values, solubility and lifespanwill be less of a concern with CFT media thanwith CHT ceramic hydroxyapatite (Figure 3). To
determine which type of apatite provides optimal
chromatographic performance, an evaluation of33.6

34.7

36.7

34.7
27.0

4.4
4.5

5.0

5.5

6.0

pH

6.5

7.0

7.5

8.0

Fig 1. IgG binding vs. pH using CFT ceramic fluoroapatite media.
45
34.7

35

33.5

28.4

25
15
500

50

100

150

200

250

300

350

400

450

500

Flow rate, cm/hFig 2. IgG binding vs. flow rate using CFT ceramic fluoroapatite media.
200

CFT Type II, 40 µm

30

2131100 Cycles

100080 Cycles
60 Cycles
40 Cycles
20 Cycles
1 Cycle020051015Retention time, min

CHT Type II, 40 µm

3100 Cycles

100080 Cycles
60 Cycles
40 Cycles
20 Cycles
1 Cycle0510
15Retention time, min

10030

211202020Fig 3. Separation of a mixture of three proteins demonstrated
stability of CFT vs. CHT over 100 cycles at pH 5.0.

20100Conductivity, mS/cm

CFT ceramic fluoroapatite possesses separation
characteristics similar to those of the more widely
known CHT™ ceramic hydroxyapatite. However,when CFT is used, the process scientist canperform purification across a range of lower pHvalues to obtain optimal results for the targetedbiomolecule. CFT can be used under stringent
chromatography conditions to separate acidic
proteins with minimal compromise to the solubility
or lifespan of the apatite. Its tensile strength,chemical durability, and density enable it toprovide the throughput and reproducibility
required for process-scale manufacturingof biopharmaceuticals.

454035302520
151050

Conductivity, mS/cm	Inorganic calcium
phosphate backbone
for distinct selectivities

CFT and CHT media should be performed withattention to buffer pH and selectivity.

lgG, mg/mln	Rigid particles forfast cleaning andequilibration

CFT ceramic fluoroapatite is a rigid, spherical,
macroporous support used in the purification of
biologically significant compounds. CFT is acomposite of fluoroapatite and hydroxyapatite
prepared by chemically converting hydroxyapatite
nanocrystals to fluoroapatite with a fluorine
reagent. The chemical conversion allows CFT
media to withstand the rigors of protein separations
requiring acidic-buffered conditions (pH 5–6.5).

lgG, mg/mln	Sintered at hightemperature for aheavy-duty durable
supportObtain High Selectivity Under Demanding
Low pH Conditions

Absorbance at 280 nmn	Acidic protein
separation forapplications requiring
pH as low as 5.0

Absorbance at 280 nmnSpecifications
Functional groups					Particle sizes						Recommended linear flow rate			Operating pH range					Chemical compatibility																																					Ca 2+, PO43–, F –40 ± 4 µm
300 cm/hr
5–14
2 M NaOH, 			
6 M guanidine-HCl, 		
8 M urea, 				0.1 M sodium acetate, 		pH 5.7

Regeneration				
Normal conditions					400 mM sodium 											phosphate, pH 7.4
Difficult conditions					400–1,000 mM sodium								phosphate, pH 11–12
Sanitization							1–2 M NaOH or KOH
Autoclavability (121ºC, 20 min)			YesPacking density (g/ml packed bed)			
0.86 g/ml
Dynamic binding capacity				14–21.5 mg lysozyme/g
Typical IgG binding capacities 300 cm/hr
33 mg/ml			
Nominal pore diameter					600–800 Å			Maximum operating pressure			
55 bar (800 psi)		
Note: A small amount (up to 5 mM) of sodium phosphate should beadded to all unbuffered solutions as a counterion.

always be sanitized with 1–2 M potassium or sodiumhydroxide. For more detailed information on handling andusing CFT media, refer to the instruction manual.
Storage and Shelf Life
CFT ceramic fluoroapatite should be stored in 0.1–1.0 M NaOH
at room temperature. When sealed in the original container,
unused CFT ceramic fluoroapatite may be stored indefinitely
in dry form at room temperature.
Technical AssistanceA regulatory support file on CFT is available uponrequest. Bio-Rad Laboratories Inc., is an ISO 9001
registered corporation. For additional information andtechnical assistance, contact your local Bio-Rad office.
In the USA and Canada, call 1-800-4BIORAD.
Visit us on the Web at www.bio-rad.com for more
information on Bio-Rad’s complete line of process
chromatography supports and other products for life
science research and production.

Reference
Mechanism of Action andSchubert S and Freitag R (2009). Investigation of the interaction
Standard Chromatographymechanism of the recombinant human antibody MDJ8 and its
The unique selectivity of CFT ceramic fluoroapatite is
fragments with chromatographic apatite phases. J Chromatogr A 1216,3831–3840.
attributed to its multiple interactions with biomolecules.
Amino groups are attracted to phosphate sites (P-sites) but Schubert S and Freitag R (2007). Comparison of ceramic hydroxy- andfluoroapatite versus protein A/G–based resins in the isolation of arepelled by calcium sites (C-sites); the situation is reversed recombinant human antibody from cell culture supernatant.
for carboxyl groups. Although amine binding to P-sites and J Chromatogr A 1142, 106–113.
the initial attraction of carboxyls to C-sites are electrostatic, Ordering Information
the actual binding of carboxyls to C-sites involves
Catalog #Description
formation of much stronger coordination complexes
158-5200
CFT Ceramic Fluoroapatite, Type II, 40 μm, 10 g
157-5000
CFT Ceramic Fluoroapatite, Type II, 40 μm, 100 g
between C-sites and clusters of protein carboxyls.
157-5100
CFT Ceramic Fluoroapatite, Type II 40 μm, 1 kg
Phosphoryl groups on proteins and other solutes interact
157-5500
CFT Ceramic Fluoroapatite, Type II 40 μm, 5 kg
even more strongly with C-sites than do carboxyl groups.
Related Items
The role of fluoride ions in the chromatographic interaction
CHT Ceramic Hydroxyapatite, Type II, 20 μm
with biomolecules is unclear, but the partial substitution of158-2200
CHT Ceramic Hydroxyapatite, 10 ghydroxyl groups by fluoride ions improves the stability of CFT
157-2000
CHT Ceramic Hydroxyapatite, 100 g
157-2100
CHT Ceramic Hydroxyapatite, 1 kg
ceramic fluoroapatite under increasingly acidic conditions.
Standard chromatography is performed using low ionicstrength phosphate buffers (potassium or sodium) to bind
acidic, basic, and neutral proteins. A gradient of increasing
concentrations of sodium chloride or of phosphate is used
to elute bound molecules. CFT ceramic fluoroapatite should
be regenerated after each run with high strength phosphatebuffer. Other effective cleaning-in-place (CIP) agents include
1–2 M potassium or sodium chloride, 400 mM trisodium
phosphate, 6 M urea or guanidine-HCl, and pure organic
solvents. Prior to storage, CFT ceramic fluoroapatite should

157-2500

CHT Ceramic Hydroxyapatite, 5 kg

CHT Ceramic Hydroxyapatite, Type II, 40 μm
158-4200
CHT Ceramic Hydroxyapatite, 10 g
157-4000
CHT Ceramic Hydroxyapatite, 100 g
157-4100
CHT Ceramic Hydroxyapatite, 1 kg
157-4500
CHT Ceramic Hydroxyapatite, 5 kg
CHT Ceramic Hydroxyapatite, Type II, 80 μm
158-8200
CHT Ceramic Hydroxyapatite, 10 g
157-8000
CHT Ceramic Hydroxyapatite, 100 g
157-8100
CHT Ceramic Hydroxyapatite, 1 kg
157-8500
CHT Ceramic Hydroxyapatite, 5 kg

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Life ScienceGroup

Bulletin 3111

US/EG

Rev B09-0354

0609 Sig 0308