| Human parvovirus B19 is the only known parvovirus causing human diseases. B19 is a non-enveloped virus with a diameter of about 18~20nm and containing a single-stranded linear DNA genome of about 5.5kb, and it can resist the heat-inactivation treatment. B19 infection has a worldwide distribution with its incidence is very high in the general population and B19 epidemics occurs from late winter to early spring. B19 virus has a strong resistance to the current virus inactivation processes due to its unique characteristics, so for the quality control of plasma products, screening of plasma source material is an important means to lower the viral load. The PCR technology is simple, intuitive, reproducible, specific and accurate quantitative. In the first part of the article, we take advantage of the technology to investigate the situation of human parvovirus B19 DNA load in plasma-derived medicinal products, in order to improve their production process and give basis to their quality control. Among the 2276 individual plasma samples, 9 samples were positive, and 4 were confirmed positive after repeated test of the 9 positive samples for 3 times. Tested the plasma-derived medicinal products, 84 batches of intravenous immunoglobulin were all negative; One batch was confirmed positive in 14 batches of human fibrinogen; 9 batches were confirmed positive in the 25 batches of factor VIII and 18 batches were positive in 24 batches of prothrombin complex. The positive frequency in the source plasma and plasma products are similar to those reported in the foreign literatures. This study will build up a good foundation for further researches.Traditional process of blood products inactivation such as heating, S/D, and low pH method have good effection on coated virus, but for nonenvelop virus inactivation, there always been a controversial. In recent years, it emerges many methods aiming at inactivating uncoated virus such as nanometer film filtering method, photochemical method, but all those have poor effection on human parvovirus B19. In the second part of the study, we explord a new method of short-wave ultraviolet C inactivation process, using UVC inactivated UVivatec instrument ( exploited by German Bayer and Satouris ). The porcine parvovirus ( PPV ) as an indicator virus in PPC was inactivated with short-wave UVC at dosage of 200, 250 and 300 J/m2 respectively. The virus titer was determined by micro-cell culture method. Validate the inactivation effect, short-wave UVC showed good effectiveness. The titer of PPV inactivated with short-wave UVC decreased by more than 4.0 log, accorded with the virus inactivated validation guidelines requirements.In the third part of the study, according with the 2010 Pharmacopoeia III, detecting activities of samples before and after UVC treated, we find that the recovery rate of coagulation factor activity in absence of stabilizer was more than 70%. To exclude subtle changes in the structures of proteins, the PPC samples were evaluated for structures and functions by 2-DE and HPLC. HPLC proved that the polymer contents of coagulation factor-proteins and the chromatographic behaviores of the samples after UVC irradiation showed no significant change as compared with that before irradiation. 2-DE showed that the number of protein spots after irradiation was basically consistent with that before irradiation.Human parvovirus B19 incidence is very high in the general population, and it has potential pathogenic threats to immune deficiency patients. A single plasma unit containing the virus has the potential risk of contamination of plasma pools, the inactivation effection of nonenvelop virus by short-wave ultraviolet UVC is good. To exclude short-wave ultraviolet UVC irradiation make proteins form polymers or cracked into small protein fragments, thereby causing changes in the activity of functional effects, we use active detection and 2-DE and SEC-HPLC to study the functions and structures of the samples after treated. So that a variety of detection methods complement each other to obtain a more satisfactory result. The activities of samples before and after the UVC inactivation are stable and the activity recovery is over 70%, the difference from before and after inactivation, and between different inactivating doses are small, and the activities are within the reference range of the pharmacopoeia. UVC inactivation of non-enveloped virus showed prominent advantages, and also provides us with new ideas and methods. |
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