Efficacy of minute virus of mice (MVM) inactivation utilizing high temperature short time (HTST) pasteurization and suitability assessment of pasteurized, concentrated glucose feeds in Chinese hamster ovary (CHO) cell expression systems

There is a growing need to provide effective adventitious agent mitigation for high risk upstream cell culture raw materials used for the production of biologics. It is also highly important in the growing fields of cell and gene therapies. Glucose is a critical raw material necessary for effective cell growth and productivity; however, glucose is the highest risk animal-origin-free raw material for viral contamination, and often the highest risk raw material in the upstream process as more companies move to chemically defined media.
This study examines the efficacy of utilizing High Temperature Short Time (HTST) pasteurization for inactivation of physiochemically resistant, worst-case parvovirus using a bench-scale HTST system. We demonstrated approximately six log inactivation of Minute Virus of Mice (MVM) in concentrated glucose feeds without impacting the subsequent performance of the glucose in a Chinese Hamster Ovary (CHO) expression system.

The NS1 protein of the parvovirus MVM Aids in the localization of the viral genome to cellular sites of DNA damage

The autonomous parvovirus Minute Virus of Mice (MVM) localizes to cellular DNA damage sites to establish and sustain viral replication centers, which can be visualized by focal deposition of the essential MVM non-structural phosphoprotein NS1. How such foci are established remains unknown. Here, we show that NS1 localized to cellular sites of DNA damage independently of its ability to covalently bind the 5′ end of the viral genome, or its consensus DNA binding sequence. Many of these sites were identical to those occupied by virus during infection.
However, localization of the MVM genome to DNA damage sites occurred only when wild-type NS1, but not its DNA-binding mutant was expressed. Additionally, wild-type NS1, but not its DNA binding mutant, could localize a heterologous DNA molecule containing the NS1 binding sequence to DNA damage sites. These findings suggest that NS1 may function as a bridging molecule, helping the MVM genome localize to cellular DNA damage sites to facilitate ongoing virus replication.

A comparison of mvM stress of inlays, onlays and endocrowns made from various materials and their bonding with molars in a computer simulation of mastication – FEA

Objectives: The purpose of the study was to compare the mvM stresses occurring in inlays, onlays and endocrowns made from different materials and their bonding with molars in a computer simulation of mastication.
Methods: The study was conducted using the finite elements method with contact elements. Sixteen 3D first molar models were created of a intact tooth – T; a tooth with a ceramic inlay – IN; a tooth with an onlay – ON; and a tooth with an endocrown – EN. The restorations were made of: Comp – resin nanoceramic; Hc – hybrid ceramic; Le – leucite ceramic; Dlit – lithium disilicate; and Zr – zirconia. Computer simulations of mastication were performed. The equivalent stresses according to the modified von Mises criterion (mvM) were calculated in model materials and contact stresses at the interface cement-dental tissue around the examined restorations.
Results: The highest equivalent mvM stresses were concentrated in buccal margins of inlays. The mvM stresses recorded in onlays were 1.6-5 times lower than those found in inlays, while in endocrowns they were 2.3-6.5 times lower. Around the onlays and endocrowns, in tooth structures and luting cement, mvM stresses were significantly lower compared to teeth restored with inlays. The tensile and shear contact stresses between inlays and teeth were several times lower than under another restorations. The highest stresses (58.5MPa) occurred in the zirconia inlay. The stresses observed in the enamel of a tooth restored with an INZr inlay were half those noted in INComp, and a third of those observed in cement. Tensile contact stresses at the interface between the INZr inlay and dental tissue were 4.5 times lower than in the INComp, and the shear stresses were more than 7 times lower.
Significance: The highest values and unfavorable of stress levels occurred in teeth restored with inlays. Cavities MOD in molars should be reconstructed with cusp-covering restorations. The endocrown in molars should withstand physiological loading. The higher the modulus of elasticity of the restoration material, the higher the stresses in the restorations, while the lower stresses were observed in the tooth structures, luting cement and at the interface between the restoration and the dental tissue. Ceramic restorations should provide better protection and marginal seal of the reconstructed tooth than composite ones.

Using a Non-Infectious MVM Surrogate For Assessing Viral Clearance During Downstream Process Development.

Viral contamination is an inherent risk during the manufacture of biopharmaceuticals. As such, biopharmaceutical companies must demonstrate the viral clearance efficacy of their downstream process steps prior to clinical trials and commercial approval. This is accomplished through expensive and logistically challenging spiking studies which utilize live mammalian viruses. These hurdles deter companies from analyzing viral clearance during process development and characterization.
We utilized a non-infectious Minute Virus of Mice – Mock Virus Particle (MVM-MVP) as a surrogate spiking agent during small scale Viral Filtration (VF) and Anion Exchange Chromatography (AEX) studies. For VF experiments, in-process mAb material was spiked and processed through Asahi Kasei P15, P20, P35 and BioEX nanofilters. Across each filter type, flux decay profiles and Log Reduction Values (LRV) were nearly identical for either particle. For AEX experiments, loads were conditioned with various amounts of sodium chloride (9, 20, 23 and 41 mS/cm), spiked with either particle and processed through a Q-SFF packed column. LRV results met our expectations of predicting MVM removal.

Mouse Anti-Varicella Zoster Virus (VZV/chickenpox) IgM ELISA kit, 96 tests, Quantitative

520-240-MVM Alpha Diagnostics 1 Kit 773 EUR

Rabbit Anti-Mouse Minute Virus (MVM) capsid protein VP2 antiserum

MVMVP21-S Alpha Diagnostics 100 ul 457 EUR

RecombiVirus Rat Minute Virus (MVM) IgG ELISA Kit, 96 tests

AE-300910-1 Alpha Diagnostics 1 kit 567 EUR

RecombiVirus Mouse Minute Virus (MVM) IgG ELISA Kit, 96 tests, Quantitative

AE-300900-1 Alpha Diagnostics 1 Kit 567 EUR

Recombinant purified Mouse Minute Virus (MVM) capsid protein VP2 control for Western blot

MVMVP21-C Alpha Diagnostics 100 ul 286 EUR

Mouse Anti-Mouse Minute Virus (MVM) capsid protein VP2 antibody negative control serum

MVMVP21-MNC Alpha Diagnostics 1 ml 225 EUR

Mouse Anti-Mouse Minute Virus (MVM) capsid protein VP2 antibody positive control serum

MVMVP21-MPC Alpha Diagnostics 1 ml 225 EUR

Rat Anti-Mouse Minute Virus (MVM) capsid protein VP2 antibody negative control serum

MVMVP21-RNC Alpha Diagnostics 1 ml 225 EUR

Rat Anti-Mouse Minute Virus (MVM) capsid protein VP2 antibody positive control serum

MVMVP21-RPC Alpha Diagnostics 1 ml 225 EUR

Recombinant (E. coli) Mouse Minute Virus (MVM) capsid protein VP2 (>95%, His-tag, 66 Kda)

MVMVP25-R-10 Alpha Diagnostics 10 ug 408 EUR

The effects of buffer condition on the fouling behavior of MVM virus filtration of an Fc-fusion protein.

A combined pore blockage and cake filtration model was applied to the virus filtration of an Fc-fusion protein using the three commercially available filters, F-1, F-2, and F-3 in a range of buffer conditions including sodium-phosphate and tris-acetate buffers with and without 200 mM NaCl at pH 7.5. The fouling behaviors of the three filters for the feed solutions spiked with minute virus of mice were described well by this combined model for all the solution conditions.
This suggests that fouling of the virus filters is dominated by the pore blockage mechanism during the initial stage of the filtration and transformed to the cake filtration mechanism during the later stage of the filtration. Both flux and transmembrane resistance can be described well by this model. The pore blockage rate and the rate of increase of protein layer resistance over blocked pores are found to be affected by membrane properties as well as the solution conditions resulting from the modulation of interactions between virus, protein, and membrane by the solution conditions.

Leave a Reply

Your email address will not be published.