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TnBP⁄Triton X-45 treatment of plasma for transfusion efficiently inactivates hepatitis C virus.

Chou ML, Burnouf T, Chang SP, Hung TC, Lin CC, Richardson CD, Lin LT - PLoS ONE (2015)

Bottom Line: SD treatment effectively inactivated HCVcc within 30 min, as demonstrated by the baseline level of reporter signals, total loss of viral infectivity, and absence of viral protein NS5A.SD specifically targeted HCV particles to render them inactive, with essentially no effect on plasma protein content and hemostatic function.Therefore, treatment by 1% TnBP / 1% Triton X-45 at 31°C is highly efficient to inactivate HCV in plasma for transfusion, showing its capacity to enhance the safety of therapeutic plasma products.

View Article: PubMed Central - PubMed

Affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.

ABSTRACT
Risk of transmission of hepatitis C virus (HCV) by clinical plasma remains high in countries with a high prevalence of hepatitis C, justifying the implementation of viral inactivation treatments. In this study, we assessed the extent of inactivation of HCV during minipool solvent/detergent (SD; 1% TnBP / 1% Triton X-45) treatment of human plasma. Luciferase-tagged infectious cell culture-derived HCV (HCVcc) particles were used to spike human plasma prior to treatment by SD at 31 ± 0.5°C for 30 min. Samples were taken before and after SD treatment and filtered on a Sep-Pak Plus C18 cartridge to remove the SD agents. Risk of cytotoxicity was assessed by XTT cell viability assay. Viral infectivity was analyzed based on the luciferase signals, 50% tissue culture infectious dose viral titer, and immunofluorescence staining for HCV NS5A protein. Total protein, cholesterol, and triglyceride contents were determined before and after SD treatment and C18 cartridge filtration. Binding analysis, using patient-derived HCV clinical isolates, was also examined to validate the efficacy of the inactivation by SD. SD treatment effectively inactivated HCVcc within 30 min, as demonstrated by the baseline level of reporter signals, total loss of viral infectivity, and absence of viral protein NS5A. SD specifically targeted HCV particles to render them inactive, with essentially no effect on plasma protein content and hemostatic function. More importantly, the efficacy of the SD inactivation method was confirmed against various genotypes of patient-derived HCV clinical isolates and against HCVcc infection of primary human hepatocytes. Therefore, treatment by 1% TnBP / 1% Triton X-45 at 31°C is highly efficient to inactivate HCV in plasma for transfusion, showing its capacity to enhance the safety of therapeutic plasma products. We propose that the methodology used here to study HCV infectivity can be valuable in the validation of viral inactivation and removal processes of human plasma-derived products.

No MeSH data available.


Related in: MedlinePlus

Flowchart of SD treatment and C18 filtration procedure on human plasma for HCV infectivity assay.
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pone.0117800.g001: Flowchart of SD treatment and C18 filtration procedure on human plasma for HCV infectivity assay.

Mentions: Plasma samples (thawed at 31°C) were evaluated for their potential cytotoxicity on Huh-7.5 cells prior to all experiments. Plasma samples were spiked with the HCVcc stock at a ratio of 9:1 (plasma:HCV stock) in accordance with regulatory guidelines and international recommendations [22]. For inactivation by SD, virus-spiked samples were treated with SD agents at a final concentration of 1% (v/v), each followed by vortexing and incubation for 30 min at 31°C under mild mixing. The treatment was stopped by processing the samples (0.3 ml) on Sep-Pak Plus C18 cartridge 55–105 μm cartridges (Waters Corporation, Milford, MA, USA) containing 130 mg of octadecyl (C18) material, which is known to quickly remove the SD agents and stop the reaction [23]. Samples containing plasma only, HCV-spiked plasma, or HCV-spiked plasma treated by SD minipool with or without subsequent C18 cartridge filtration were then evaluated for viral infectivity. A schematic of the procedure is shown in Fig. 1.


TnBP⁄Triton X-45 treatment of plasma for transfusion efficiently inactivates hepatitis C virus.

Chou ML, Burnouf T, Chang SP, Hung TC, Lin CC, Richardson CD, Lin LT - PLoS ONE (2015)

Flowchart of SD treatment and C18 filtration procedure on human plasma for HCV infectivity assay.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4320006&req=5

pone.0117800.g001: Flowchart of SD treatment and C18 filtration procedure on human plasma for HCV infectivity assay.
Mentions: Plasma samples (thawed at 31°C) were evaluated for their potential cytotoxicity on Huh-7.5 cells prior to all experiments. Plasma samples were spiked with the HCVcc stock at a ratio of 9:1 (plasma:HCV stock) in accordance with regulatory guidelines and international recommendations [22]. For inactivation by SD, virus-spiked samples were treated with SD agents at a final concentration of 1% (v/v), each followed by vortexing and incubation for 30 min at 31°C under mild mixing. The treatment was stopped by processing the samples (0.3 ml) on Sep-Pak Plus C18 cartridge 55–105 μm cartridges (Waters Corporation, Milford, MA, USA) containing 130 mg of octadecyl (C18) material, which is known to quickly remove the SD agents and stop the reaction [23]. Samples containing plasma only, HCV-spiked plasma, or HCV-spiked plasma treated by SD minipool with or without subsequent C18 cartridge filtration were then evaluated for viral infectivity. A schematic of the procedure is shown in Fig. 1.

Bottom Line: SD treatment effectively inactivated HCVcc within 30 min, as demonstrated by the baseline level of reporter signals, total loss of viral infectivity, and absence of viral protein NS5A.SD specifically targeted HCV particles to render them inactive, with essentially no effect on plasma protein content and hemostatic function.Therefore, treatment by 1% TnBP / 1% Triton X-45 at 31°C is highly efficient to inactivate HCV in plasma for transfusion, showing its capacity to enhance the safety of therapeutic plasma products.

View Article: PubMed Central - PubMed

Affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.

ABSTRACT
Risk of transmission of hepatitis C virus (HCV) by clinical plasma remains high in countries with a high prevalence of hepatitis C, justifying the implementation of viral inactivation treatments. In this study, we assessed the extent of inactivation of HCV during minipool solvent/detergent (SD; 1% TnBP / 1% Triton X-45) treatment of human plasma. Luciferase-tagged infectious cell culture-derived HCV (HCVcc) particles were used to spike human plasma prior to treatment by SD at 31 ± 0.5°C for 30 min. Samples were taken before and after SD treatment and filtered on a Sep-Pak Plus C18 cartridge to remove the SD agents. Risk of cytotoxicity was assessed by XTT cell viability assay. Viral infectivity was analyzed based on the luciferase signals, 50% tissue culture infectious dose viral titer, and immunofluorescence staining for HCV NS5A protein. Total protein, cholesterol, and triglyceride contents were determined before and after SD treatment and C18 cartridge filtration. Binding analysis, using patient-derived HCV clinical isolates, was also examined to validate the efficacy of the inactivation by SD. SD treatment effectively inactivated HCVcc within 30 min, as demonstrated by the baseline level of reporter signals, total loss of viral infectivity, and absence of viral protein NS5A. SD specifically targeted HCV particles to render them inactive, with essentially no effect on plasma protein content and hemostatic function. More importantly, the efficacy of the SD inactivation method was confirmed against various genotypes of patient-derived HCV clinical isolates and against HCVcc infection of primary human hepatocytes. Therefore, treatment by 1% TnBP / 1% Triton X-45 at 31°C is highly efficient to inactivate HCV in plasma for transfusion, showing its capacity to enhance the safety of therapeutic plasma products. We propose that the methodology used here to study HCV infectivity can be valuable in the validation of viral inactivation and removal processes of human plasma-derived products.

No MeSH data available.


Related in: MedlinePlus