Removal of small non-enveloped viruses by nanofiltration.
OBJECTIVE
Nanofiltration is one of the most effective virus reduction methods in the manufacturing process of plasma products. However, it is difficult to remove small viruses from high molecular weight protein preparations like immunoglobulin G or factor VIII complex by nanofiltration, because the size of the protein is similar to that of viruses. In order to separate the viruses from these proteins by nanofiltration, it is necessary to change the size of either one. In this study, we report that such non-enveloped viruses as human parvovirus B19 (B19), human encephalomyocarditis virus (EMC) or porcine parvovirus (PPV) aggregate in the presence of certain kinds of amino acids and could be easily removed by nanofiltration.


METHODS
0.3 M Glycine (or other amino acid) solution spiked with viruses was subjected to dead-end single filtration with a 35-nm pore-size filter. Virus removal by nanofiltration was either evaluated by PCR or by infectivity assay.


RESULTS
B19 in a 0.3 M glycine solution was reduced to 1:10(7.5) (7.5-log) by nanofiltration with a 35-nm pore-sized filter, whereas in PBS it was not reduced. Similarly, B19 was also reduced when suspended in other amino acids solutions. This effect was also confirmed with the other small non-enveloped viruses EMC or PPV. When 5% globulin or 5% albumin was added to a 0.3 M glycine solution, the removal rate was decreased.


CONCLUSIONS
These data suggest that viruses in the presence of certain kinds of amino acids could be aggregated and effectively removed by a filter that has a pore size larger than the size of the viruses.
Yokoyama
T
T
Japanese Red Cross, Plasma Fractionation Center, Chitose, Hokkaido, Japan. kenkyuu@pfc.jrc.or.jp
Murai
K
K
Murozuka
T
T
Wakisaka
A
A
Tanifuji
M
M
Fujii
N
N
Tomono
T
T
D000818:Animals; D001798:Blood Proteins; D036022:Capsid Proteins / Q000302:isolation & purification; D002460:Cell Line / Q000821:virology; D002522:Chlorocebus aethiops; D004686:Encephalomyocarditis virus / Q000254:growth & development / Q000302:isolation & purification / Q000648:ultrastructure; D005998:Glycine; D006510:Hepatitis B Antibodies / Q000302:isolation & purification; D006863:Hydrogen-Ion Concentration; D008854:Microscopy, Electron; D008970:Molecular Weight; D036103:Nanotechnology / Q000295:instrumentation*; D010316:Particle Size; D016732:Parvovirus B19, Human / Q000254:growth & development / Q000302:isolation & purification / Q000648:ultrastructure; D028323:Parvovirus, Porcine / Q000254:growth & development / Q000302:isolation & purification / Q000648:ultrastructure; D016133:Polymerase Chain Reaction; D012996:Solutions; D013552:Swine; D014462:Ultrafiltration / Q000295:instrumentation / Q000379:methods*; D014709:Vero Cells / Q000821:virology; D014779:Virus Replication; D014780:Viruses / Q000254:growth & development / Q000302:isolation & purification* / Q000648:ultrastructure
D001798:Blood Proteins; D036022:Capsid Proteins; D006510:Hepatitis B Antibodies; D012996:Solutions; C095547:capsid protein VP1, parvovirus B19; C095548:capsid protein VP2, parvovirus B19; D005998:Glycine