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Cellufine Sulphate |
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Affinity Chromatography Media
For concentration, purification and depyrogenation of virus, viral/microbial antigens and heparin-binding proteins
Advances in vaccines and clinical diagnostics have created an increasing demand for large volumes of highly purified and concentrated virus and viral or microbial antigens.
Cellufine Sulfate affinity media is a simple, rapid and effective means for concentration, purification and depyrogenation of these important products.
Cellufine Sulfate eliminates cumbersome, time-consuming and potentially unsafe classical ultra-centrifugation and density gradient methods.
It can also provide a significant improvement in concentration and purity. Cellufine Sulfate can reduce or eliminate the expense, ligand leakage and
reproducibility problems associated with immobilized dextran sulfate, chondroitin sulfate or heparin.
Elution of the bound product is affected through simple stepwise or gradient increases in ionic strength.
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| Cellufine Sulfate |
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 Operating Instructions |
| Technical data sheet |
| Technical note |
| Technical Note (Column Packing) |
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Features
Affinity for a wide range of live, killed or disrupted viruses, viral or microbial antigens and heparin-binding proteins
Closed column operation assures safety and product sterility
Endotoxins do not bind, allowing a rapid and contaminant free depyrogenation
Rigid, high-strength beads
Autoclavable |
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Benefits
More effective than ultracentrifugation at removing contaminants from culture media and host cells
Avoids excessive product handling and safety concerns, particularly with viral preparations
Simultaneous concentration and purification improve yield, reduce processing steps, time and costs
Gentle binding and elution conditions provide high capacity and product yield
Resists compression, providing rapid flow for high-speed processing, even in large columns, making it easily scalable
Resistant to chemical depyrogenation with base and chemically sterilizable with formalin |
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| Characteristics |
| Support Matrix |
Cellulose |
| Particle Size |
ca. 40 - 130 um |
| Particle Shape |
Spherical |
| Gel Exclusion Limit |
3kD |
| Activated Group |
Sulfate Ester |
| Total Sulfur |
>700 ug/g dry |
Protein
Lysozyme :
Hepatitis B Surface Antigen : |
>3 mg/ml
7 mg/ml |
| Environmental Resistance |
Resistant to 0.1M NaOH,
0.1 % of 37 % Formalin |
| Operating Pressure |
<2 bar (30 psi) |
| Autoclavable |
In suspension at neutral pH;
30 min at 121oC |
| Supplied |
Suspension in 20 % Ethanol |
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The nearly rigid properties of the spherical cellulose support matrix allow
outstanding flow properties, particularly in large production columns.
| Column A: |
90 x 200 mm |
| Column B: |
350 x 200 mm |
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| Virus, Viral/Microbial Antigens |
| Viruses |
Viral/Microbial Agents |
| Rabies*
Influenza*
Japanese Enchephalitis*
Feline Leukemia
Feline Herpes
Feline Calicivirus
Respiratory Syncytial Virus
Human Herpes Simplex
Human Measles
Human Parainfluenza |
Herpes Simplex gA and gB
Glycoprotein Subunits*
Hepatitis B Surface Antigen
Filamentous Hemagglutinin from
B. pertussis *
Leucocytosis Promoting Factor
Hemagglutinin* |
*These applications are covered by US and foreign process patents.
Please inquire regarding details and licensing arrangements. |
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Purification of Rabies Virus
The example in Figure 3 illustrates the high degree of concentration, purification and yields obtained with Cellufine Sulfate on typical viral preparations. |
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| Column : |
50 x 70 mm (140 ml) |
| Buffer : |
0.01M Phosphate (pH 7.2) |
| Eluant : |
1M NaCl/0.01M Phosphate (pH 7.2) |
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Load |
Eluate |
| Volume (ml) |
4,200 |
50 |
| Virus titer |
32 |
4,096 |
| Protein (ug/ml) |
8.5 |
14 |
| Yield (%) |
100 |
152 |
| Purification factor |
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79x |
| Concentration factor |
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126x |
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Purification of Influenza Virus
Hen's egg allantoic fluid was loaded directly onto a 33.3 mL gel bed and 94.5%virus was recovered in the eluate fraction. |
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Volume
(ml) |
Virus
Titer |
TCA-N
ug/ml |
Recovery
(%) |
Fold
Purification |
| Allantoic Fluid |
4200 |
77 |
337.1 |
100 |
1 |
| Wash |
6700 |
1 |
209.2 |
2.1 |
- |
| Eluate |
170 |
1797 |
448.0 |
94.5 |
20.1 |
| Column : |
50 x 170 mm |
| Buffer : |
0.01M Phosphate pH 7.4 |
| Wash : |
0.01M Phosphate pH 7.2 + 0.2M NaCl |
| Elution : |
0.01M Phosphate pH 7.0 + 1.5M NaCl |
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Antigenic Protein Purification and Depyrogenation
Cellufine Sulfate is ideal for depyrogenating virus and other microbial extracts because it does not bind endotoxins. Figure 4 shows the purification of filamentous hemagglutinin (FHA) from the whooping cough bacterium Bordetella pertussis. |
| Column : |
16 x 70 mm (20 ml) |
| Sample : |
800 ml B. pertussis culture fluid
(endotoxin titer > 1015 by Limulus lysate test) |
| Buffer : |
0.01M Phosphate (pH 7.6) |
| Eluant : |
1M NaCl/0.01M Phosphate (pH 7.6) |
| FHA Yield : |
94% |
| Purification Factor : |
20x |
| Concentration Factor : |
28x (30 ml product) |
| Endotoxin : |
Below standard level by Limulus lysate, rabbit pyrogen and mouse toxicity tests |
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Protein Purification
Cellufine Sulfate mimics the affinity of heparin or dextran sulfate for many proteins. It can function as an affinity support for selected plasma proteins, cellular growth factors and lipases. Its capacity comparable to conventional heparin gels. |
| Binding Proteins |
Non-binding Proteins |
| Antithrombin III
β-Lipoprotein
Complement C5, C6, C8
Complement C3 Activator
Tryspin
Tryspin Inhibitor
Chymotrypsinogen
Lysozyme
Urease
Catalase
Factor IX |
Albumin
α-Lipoprotein
Complement C3, C9
Complement C1, C3b
Inactivators
IgG
Ceruloplasmin
α2-Macroglobulin
RNase
Bacitracin
Glucose Oxidase |
| *Binding and elution are extremely rapid and very fine separations can be generated in gradient mode. |
| Purification of Partially Purified Casein Kinase II from Calf Thymus |
| Column : |
10 x 20 mm |
| Sample : |
7 ml |
| Buffer : |
50mM Tris-HCl (pH 7.9)
+ 50mM MgCl2
+ 0.1mM EDTA
+ 0.1mM PMS
+ 0.5mM DTT
+ 25 % glycerol |
| Eluant : |
0.05 - 1.0M NaCl in buffer |
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