Purification of flavivirus VLPs by a two-step chomatographic process

Lima, Túlio M.; Souza, Matheus Oliveira de; Castilho, Leda R.

doi:10.1016/j.vaccine.2019.05.066

Cited by 22 publications

(19 citation statements)

References 36 publications

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“…After harvest on day 3 of transfection, cell-free supernatants were obtained by centrifugation at 5,000 g for 10 min followed by microfiltration using 0.45 um membranes. Prior to use in sorting experiments, ZIKV VLPs were purified by liquid chromatography in 50 mM Tris pH 8.5 buffer, using Sartobind Q membrane adsorber (Sartorius) and Captocore 700 resin (GE Healthcare), as previously described ( 45 , 46 ). For confirmation of VLP size and morphology, VLPs were characterized by transmission electron microscopy (TEM) using a Hitachi 7600 microscope equipped with a lens-coupled CCD (HV = 80 kV).…”

Section: Methodsmentioning

confidence: 99%

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Functional Profiling of Antibody Immune Repertoires in Convalescent Zika Virus Disease Patients

et al. 2021

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The re-emergence of Zika virus (ZIKV) caused widespread infections that were linked to Guillain-Barré syndrome in adults and congenital malformation in fetuses, and epidemiological data suggest that ZIKV infection can induce protective antibody responses. A more detailed understanding of anti-ZIKV antibody responses may lead to enhanced antibody discovery and improved vaccine designs against ZIKV and related flaviviruses. Here, we applied recently-invented library-scale antibody screening technologies to determine comprehensive functional molecular and genetic profiles of naturally elicited human anti-ZIKV antibodies in three convalescent individuals. We leveraged natively paired antibody yeast display and NGS to predict antibody cross-reactivities and coarse-grain antibody affinities, to perform in-depth immune profiling of IgM, IgG, and IgA antibody repertoires in peripheral blood, and to reveal virus maturation state-dependent antibody interactions. Repertoire-scale comparison of ZIKV VLP-specific and non-specific antibodies in the same individuals also showed that mean antibody somatic hypermutation levels were substantially influenced by donor-intrinsic characteristics. These data provide insights into antiviral antibody responses to ZIKV disease and outline systems-level strategies to track human antibody immune responses to emergent viral infections.

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Section: Methodsmentioning

confidence: 99%

“…Transient transfection of HEK293F cells, harvest and clarification were carried out as described for ZIKV VLPs. Yellow fever VLPs were purified by liquid chromatography in 50 mM Tris pH 8.5 buffer, using Sartobind Q membrane adsorber (Sartorius) and Captocore 700 resin (GE Healthcare), as previously described ( 45 , 46 ).…”

Section: Methodsmentioning

confidence: 99%

Functional Profiling of Antibody Immune Repertoires in Convalescent Zika Virus Disease Patients

et al. 2021

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“…The aforementioned issues associated with the current YFV 17D vaccines has spawned a number of approaches to develop new YF vaccine strategies [ 6 , 16 ]. These include production of the live attenuated vaccine in Vero cells (ClinicalTrials.gov Identifier: NCT04142086), a beta-propiolactone inactivated YF vaccine [ 17 , 18 ] (ClinicalTrials.gov Identifier: NCT00995865), a Modified Vaccinia Ankara vectored recombinant YF vaccine [ 19 ] (ClinicalTrials.gov Identifier: NCT02743455) and (at a preclinical stage) a virus-like particle (VLP) YF vaccine manufactured in transfected HEK293 cells [ 20 ].…”

Section: Introductionmentioning

confidence: 99%

A Yellow Fever Virus 17D Infection and Disease Mouse Model Used to Evaluate a Chimeric Binjari-Yellow Fever Virus Vaccine

et al. 2020

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Despite the availability of an effective, live attenuated yellow fever virus (YFV) vaccine (YFV 17D), this flavivirus still causes up to ≈60,000 deaths annually. A number of new approaches are seeking to address vaccine supply issues and improve safety for the immunocompromised vaccine recipients. Herein we describe an adult female IFNAR-/- mouse model of YFV 17D infection and disease that recapitulates many features of infection and disease in humans. We used this model to evaluate a new YFV vaccine that is based on a recently described chimeric Binjari virus (BinJV) vaccine technology. BinJV is an insect-specific flavivirus and the chimeric YFV vaccine (BinJ/YFV-prME) was generated by replacing the prME genes of BinJV with the prME genes of YFV 17D. Such BinJV chimeras retain their ability to replicate to high titers in C6/36 mosquito cells (allowing vaccine production), but are unable to replicate in vertebrate cells. Vaccination with adjuvanted BinJ/YFV-prME induced neutralizing antibodies and protected mice against infection, weight loss and liver pathology after YFV 17D challenge.

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“…During the process of virus purification, due to the significant differences of molecular weight, density, charge properties between virus molecules and impurity molecules, most viruses can be purified by density gradient centrifugation and gel filtration chromatography (SEC) [27][28][29][30]. Compared with the density gradient ultracentrifugation method, gel chromatography is significantly faster, more consistent and can realize more likely automatically [31,32].…”

Section: Discussionmentioning

confidence: 99%

Purification of cell‐derived Japanese encephalitis virus by dual‐mode chromatography

Zhang

Luo

Teng

et al. 2020

Biotech and App Biochem

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Purification of the enveloped virus poses a challenge as one must retain viral infectivity to preserve immunogenicity. The traditional process of virus purification is time-consuming, laborious and hard to scale up. Here, a rapid, simple and extensible laboratory program for the purification of Japanese encephalitis virus (JEV) was developed by using differential centrifugation, ultrafiltration, Sepharose 4 fast flow gel chromatography, and Capto TM Core 700 chromatography. The entire process recovered 61.64% of the original virus, and the purified virus particles maintained good activity and immunogenicity. The purification process described has potential application in large-scale production of high-purity JEV.

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Purification of flavivirus VLPs by a two-step chomatographic process

Cited by 22 publications

References 36 publications

Functional Profiling of Antibody Immune Repertoires in Convalescent Zika Virus Disease Patients

Functional Profiling of Antibody Immune Repertoires in Convalescent Zika Virus Disease Patients

A Yellow Fever Virus 17D Infection and Disease Mouse Model Used to Evaluate a Chimeric Binjari-Yellow Fever Virus Vaccine

Purification of cell‐derived Japanese encephalitis virus by dual‐mode chromatography

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