Purification of equine whole IgG snake antivenom by using an aqueous two phase system as a primary purification step

Vargas, Mariángela; Segura, Álvaro; Villalta, Mauren; Herrera, María; Gutiérrez, José Marı́a; León, Guillermo

doi:10.1016/j.biologicals.2014.10.003

Cited by 9 publications

(2 citation statements)

References 45 publications

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“…The whole IgGs are extracted from animal plasma, mostly equine or ovine, by various refinement strategies and either serve as the intermediate in further processing towards F(ab') 2 or Fab fragments or represent the final form of the active drug. The well-established and most commonly used industrial purification procedures on the large-scale are salting-out of IgGs with sodium or ammonium sulphate (ASP) and caprylic acid (CA) precipitation of non-IgG proteins [11,12]. The reported limitations of salt-mediated methods are not only the low recovery of antibody activity (less than 50%) [10] and the poor yield, but also the hardly reachable compliance with regulatory requirements concerning purity [11] and aggregate content [13,14].…”

Section: Introductionmentioning

confidence: 99%

Quality-Related Properties of Equine Immunoglobulins Purified by Different Approaches

Lukačević

Kurtović

Balija

et al. 2020

Toxins

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Whole IgG antivenoms are prepared from hyperimmune animal plasma by various refinement strategies. The ones most commonly used at industrial scale are precipitation by sodium or ammonium sulphate (ASP), and caprylic acid precipitation (CAP) of non-immunoglobulin proteins. The additional procedures, which have so far been used for experimental purposes only, are anion-exchange (AEX) and cation-exchange chromatography (CEX), as well as affinity chromatography (AC) using IgG’s Fc-binding ligands. These protocols extract the whole IgG fraction from plasma, which contains both venom-specific and therapeutically irrelevant antibodies. Such preparations represent a complex mixture of various IgG subclasses whose functional and/or structural properties, as well as relative distribution, might be affected differently, depending on employed purification procedure. The aim of this work was to compare the influence of aforementioned refinement strategies on the IgG subclass distribution, venom-specific protective efficacy, thermal stability, aggregate formation and retained impurity profile of the final products. A unique sample of Vipera ammodytes ammodytes specific hyperimmune horse plasma was used as a starting material, enabling direct comparison of five purification approaches. The highest purity was achieved by CAP and AC (above 90% in a single step), while the lowest aggregate content was present in samples from AEX processing. Albumin was the main contaminant in IgG preparations obtained by ASP and CEX, while transferrin dominantly contaminated IgG sample from AEX processing. Alpha-1B-glycoprotein was present in CAP IgG fraction, as well as in those from ASP- and AEX-based procedures. AC approach induced the highest loss of IgG(T) subclass. CEX and AEX showed the same tendency, while CAP and ASP had almost no impact on subclass distribution. The shift in IgG subclass composition influenced the specific protective efficacy of the respective final preparation as measured in vivo. AC and CEX remarkably affected drug’s venom-neutralization activity, in contrary to the CAP procedure, that preserved protective efficacy of the IgG fraction. Presented data might improve the process of designing and establishing novel downstream processing strategies and give guidance for optimization of the current ones by providing information on potency-protecting and purity-increasing properties of each purification principle.

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Section: Introductionmentioning

confidence: 99%

Quality-Related Properties of Equine Immunoglobulins Purified by Different Approaches

Lukačević

Kurtović

Balija

et al. 2020

Toxins

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“…Following 7 weeks at rest after immunization and bleeding, two groups of three horses each were immunized and bled as described above. Then, immediately after the bleeding on the third day, the first group received an intravenous infusion of equine albumin at a dose of 2 g/kg body weight, in a total volume of 8–9 L. Equine albumin was purified by the method of aqueous two-phase system ( Vargas et al, 2015 ) from plasma previously collected from the same group of horses. The solution was sterile, endotoxin-free and formulated at 10 g/dL total protein, with a purity of 70% ( Supplementary Fig.…”

Section: Methodsmentioning

confidence: 99%

Clinical effects of immunization, bleeding, and albumin-based fluid therapy in horses used as immunoglobulin source to produce a polyspecific antivenom (Echitab-plus-ICP) towards venoms of African snakes

et al. 2023

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Concept of sample-specific correction of immunoassay results for precise and accurate IgG quantification in horse plasma

Halassy

Kurtović

Balija

et al. 2019

Journal of Pharmaceutical and Biomedical Analysis

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Purification of equine whole IgG snake antivenom by using an aqueous two phase system as a primary purification step

Cited by 9 publications

References 45 publications

Quality-Related Properties of Equine Immunoglobulins Purified by Different Approaches

Quality-Related Properties of Equine Immunoglobulins Purified by Different Approaches

Clinical effects of immunization, bleeding, and albumin-based fluid therapy in horses used as immunoglobulin source to produce a polyspecific antivenom (Echitab-plus-ICP) towards venoms of African snakes

Concept of sample-specific correction of immunoassay results for precise and accurate IgG quantification in horse plasma

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