Technical Note – Products in Development

CA_MLP_DexS_N1_V8_E

Cellulose resin MLP1000 DexS
Next generation cellulose resin with engineered large through pores optimized for
purification of large biomolecules including whole virus particles
Cellulose resin MLP1000 DexS is based on next-generation cellulose resin MLP1000, and sulfatedpolysaccharide is immobilized as a ligand. This resin is designed for isolation and purification ofinactivated virus for vaccine production.
The cellulose resin MLP1000 is a highly crosslinked cellulose particle that has a large continuous pore
structure. In the purification of large virus and virus-like particles, MLP1000 DexS exhibits a high binding
capacity and purification capabilities because the intraparticle surface of the resin can be used as anadsorption site.
Surface

Cross section

Fig.1. Cellulose resin MLP1000

How to request products under development
Cellulose resin MLP1000 DexS is currently under development. The quality of the Cellufinechromatography media is guaranteed by ISO 9001, but the quality of this product is subject to change asit is under development.
This product is NOT for sale. We can provide small volume samples for evaluation. For
pharmaceutical companies, diagnostics companies or other manufacturing companies and researchers
who would like to handle this product for manufacturing process, we will provide samples those whoconsenting to above fact.
For inquiries regarding samples, please contact the following.

JNC Corporation
Life Chemicals Division
2-1, Otemachi 2-Chome, Chiyoda-ku, Tokyo 100-8105, Japan
Phone +81-3-3243-6150, Fax+81-3-3234-6219
E-mail: cellufine@jnc-corp.co.jp
http://www.jnc-corp.co.jp/fine/en/cellufine/1

Technical Note – Products in Development

CA_MLP_DexS_N1_V8_E■ Characteristics of MLP1000
Cellulose resin MLP1000 is a highly crosslinked
cellulose particle with a large continuous porestructure. In the purification of large virus andvirus-like particles, MLP1000 DexS exhibits highbinding capacity and purification capabilities
because the intraparticle surface of the resin canbe used as an adsorption site.
Fig. 3. Flow characteristics ofMLP1000 DexS
After packing the resin into a 2.2 cm ID x 20 cm
column, pure water at 24°C ± 1°C was used as themobile phase, and the relationship between flow rateand pressure drop across the column was evaluated.

■ Purification of influenza virus

Chromatography

purificationwasperformed

using allantoic fluid containing influenza A virus
H1N1. The cellulose resin MLP1000 DexS wasFig. 2. Retention times of PEG and PEO

packed into the column and the allantoic fluid wasin MLP1000

directly loaded. Virus in the load, flow through,After packing the resin into 7.8 mm ID x 300 mm

wash and elution fractions was measured using ancolumn, polyethylene glycol (PEG) or polyethylene

HA assay. Figure 4 shows a comparison between

oxide (PEO) with different index molecular weights

MLP1000 DexS and a commercially available resin

was passed through the column at a flow rate of 0.4

A (dextran sulfate modified resin). MLP1000 DexSmL/min to confirm the retention time.

continued to adsorb the virus without a 10%
break-through

until

20mL

loading,

while

Table 1. Properties of MLP1000 DexScommercially available resin A showed a 10%

Particle size

Ave. 90 µm (40 – 150 µm)

break-through when about 4 mL of liquid wasLigand

Dextran sulfate

passed through the column.

Base resin

Cross linked cellulose

Pressure< 0.3MPa

In the case of general chromatography resin, thepore size is small and virus particles cannot enter
inside the particle. For this reason, the virus isadsorbed on only the surface of the resin, and the■ Flow property

binding sites are limited. In contrast, the cellulose

The cellulose resin MLP1000 DexS is aresin

highly crosslinked cellulose particle. Despite

MLP1000DexS

has

extremely

large

through-pores. This allows virus particles to enter

having extremely large open pores for virus

inside the resin. In this experiment, it did not show

particles to enter, the resin has high pressure

a 10% break-through even if the load sample

resistance and can be used in large columns

exceeded 20 mL, which is more than five times

to produce biopharmaceuticals (Fig. 3).

10% DBC of a commercially available resin A.

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Technical Note – Products in Development

CA_MLP_DexS_N1_V8_E

The recovery rate and purity of inactivatedinfluenza A virus are summarized in Table 2.
With chromatography using the cellulose
resin MLP1000 DexS, the liquid volume wasreduced from 20 mL to 1.5 mL and the virus
could be concentrated up to 13-fold. The
amount of protein impurities was reduced
from 10,100 µg to 975 µg, resulting in aFig. 4. Dynamic binding capacity of Inactivated

90.3% reduction. It was also able to reduce

Influenza Virus

the amount of dsDNA to 22% in the viral

Experimental conditions

elution fraction, achieving a final virus

Resin: Cellulose resin MLP1000 DexS,

recovery rate of 93%. The cellulose resin

Commercially available resin AMLP1000 DexS shows a high binding

Column: I.D. 5.0 x 15 mm (0.3 mL)

capacity and excellent product recovery

Load sample: Allantoic fluid containing inactivatedunder high flow conditions.

influenza A virus
Virus strain: A/Hyogo/YS/2011(H1N1)
Flow rate: 0.5 ml/min (150 cm/h, R.T. 0.6 min)Equilibration: 10 mM Na phosphate, 120 mM NaCl,
pH 7.4
Wash: 10 mM Na phosphate, 120 mM NaCl, pH 7.4
Elution: 10 mM Na phosphate, 2 M NaCl, pH 7.4

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Technical Note – Products in Development

CA_MLP_DexS_N1_V8_E

Table 2. Summary of recovery and purity of inactivated influenza virus1) Cellulose resin MLP1000 DexS
Process

Volumeml

Virus activity
HA activity

ProteinsµgdsDNAµg

Recovery%

Allantoic fluid
Flow throughWash
Elution202061.5

99,600400
1,560
92,88010,100
9,220
786
975

6.4
3.6
0.3
1.410012932) Commercially available resin AProcess

Volumeml

Virus activity
HA activity

Proteins
µg

dsDNA
µg

Recovery%

Allantoic fluid
Flow throughWash
Elution

2020
6
1.5

99,60073,6006,720
12,90010,100
7,520
780
330

6.4
4.60.180.710074713■ Purification of Human coronavirus OC43

Human coronavirus OC43 (hCoV OC43) is acommon cold-causing virus of approximately 100
nm in size belonging to the same beta
coronavirus genus as SARS-CoV-2 that causes
COVID-19 infection. Following the example of thepurification of influenza virus, we present an
example of the purification of human coronavirus
OC43 cultured in Vero cells by chromatography.
Thechromatographicpurification

wasFig. 5. Dynamic binding capacity of Inactivatedperformed using cell culture supernatant fluid

Human coronavirus OC43

containing hCoV OC43. The cellulose resinExperimental conditions

MLP1000 DexS was packed into the column andResin: Cellulose resin MLP1000 DexS,

the cell culture supernatant fluid was directly

Commercially available resin Aloaded. The virus in the load, flow through, washColumn: I.D. 5.0 x 15 mm (0.3 mL)and elution fractions was measured using an HA

Load sample: cell culture supernatant fluid containing

assay. Figure 5 shows that the MLP1000 DexS

inactivated hCoV OC43

adsorbs a larger amount of the virus than theFlow rate: 0.3 ml/min (90 cm/h, R.T. 1.0 min)commercially available resin A (dextran sulfate

Equilibration: 10 mM Na phosphate, 150 mM NaCl,modified resin). While the commercially available

pH 7.4

resin A showed a 10% break-through when about

Wash: 10 mM Na phosphate, 150 mM NaCl, pH 7.4

18 mL loading, the MLP1000 DexS continued toElution: 10 mM Na phosphate, 2 M NaCl, pH 7.4

adsorb the virus without a 10% break-througheven after 34.5 mL was loaded.

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Technical Note – Products in Development

CA_MLP_DexS_N1_V8_E

In this human coronavirus adsorption experiment, the cellulose resin MLP1000 DexS showed theextremely high adsorption capacity with less than a 3% break-through, when a 10 % break-throughwas observed with the commercially available resin.
The recovery rate and purity of the chromatographically purified inactivated human coronaviruses aresummarized in Table 3. With the chromatographic purification using the cellulose resin MLP1000 DexS,
the amount of protein impurities was reduced from 6394 µg to 170 µg, resulting in a 97.3% reduction. It
was also able to reduce the amount of dsDNA to 9% in the viral elution fraction, achieving a final virus
recovery rate of 91%, which showed a high recovery performance.
It was shown that MLP1000 DexS enables more efficient purification of the human coronavirus
OC43 belonging to the same beta coronavirus genus as SARS-Cov-2 as well as the influenza virus.
Table 3. Summary of recovery and purity of inactivated human coronavirus OC433) Cellulose resin MLP1000 DexS
Process
supernatant fluid
Flow throughWash
Elution(gradient, isocratic)

Volumeml

Virus activity
HA activity

Proteinsµg

dsDNAµg

Recovery%

34.5
34.5
4.51201861020
906394
4376
57

52.79
55.53
2.25

100
0.80.110.5108990

170

4.95

91

Volumeml

Virus activity
HA activity

ProteinsµgdsDNAµg

Recovery%

34.5
34.5
4.5

12018625770
600

69605479
59

45.48
50.3
2.4

100
210.5

10.583550

1648.94704) Commercially available resin AProcess
Allantoic fluid
Flow throughWash
Elution(gradient, isocratic)

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Technical Note – Products in Development

CA_MLP_DexS_N1_V8_E■ Conclusion

Cellulose resin MLP1000 is a next-generation chromatography resin with large pores thatoverturns conventional wisdom. Its huge through-pores allow giant biomolecules such as virus todiffuse into the particle inside. Conventional chromatography resins have poor pressure resistancewhen their pores are large enough to allow entry of whole virus particles, making it difficult to use atlarge process scale. In contrast, the cellulose resin MLP1000 has extremely high-pressure
resistance and can be used in large bioprocess columns.
Recently there has been growing increasing trend in the use large biomolecules as
pharmaceuticals. For example, adeno-associated virus (AAV) is used in gene therapy, and herpes
simplex virus (HSV) is used in antitumor virotherapy. In addition, exosomes, which are particles ofabout 100 nm, have been vigorously studied in the field of regenerative medicine. The mRNA based
COVID-19 vaccines are also large molecules. Such biomolecules have difficulty entering inside thepores of conventional chromatography resin, limiting adsorption performance.
Cellulose resin MLP1000 has an extremely large pore size that allows virus particles to diffuse
within the inside structure. As a result, the virus can access the intraparticle surface, leading to an
extremely high dynamic binding capacity. The bottleneck of conventional chromatography media
can be eliminated.
JNC Corporation is working on the development of this new chromatography resin in response to
the demand for next-generation modality purification. We will continue to contribute to thedevelopment of medical care by providing new purification processes for large biopharmaceuticals.
■ Technical information: Reference

K. Kadoi, E. Iwamoto, T. Nakama, Fabrication and characterization of a cellulose monolith-likeparticle for virus purification, Biochem Eng J. 192 (2023) 108849.
https://doi.org/10.1016/j.bej.2023.108849.

JNC Corporation
Life Chemicals Division
Shin Otemach Bldg.,2-1 Otemachi 2-Chome,Chiyoda-ku, Tokyo 100-8105 Japan
TEL：+81-3-3243-6150 Fax：+81-3-3243-6219
e-mail: cellufine@jnc-corp.co.jp
http://www.jnc-corp.co.jp/fine/en/cellufine/6