Production and purification of Clostridium perfringens type C beta-toxin and IgG and IgY antitoxins

IF 4.6 Q2 MATERIALS SCIENCE, BIOMATERIALS ACS Applied Bio Materials Pub Date : 2023-12-30 DOI:10.1016/j.anaerobe.2023.102817

Guilherme Guerra Alves , Ronnie Antunes Assis , Victor Santos do Amarante , Carlos Augusto de Oliveira Júnior , Rodrigo Otávio Silveira Silva , Luiz Guilherme Dias Heneine , Francisco Carlos Faria Lobato

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Abstract

Objectives

This study aimed to produce and purify Clostridium perfringens type C beta-toxin, sheep anti-beta toxin immunoglobulin G (IgG) and chicken immunoglobulin Y (IgY).

Methods

Two methods were used for beta-toxin purification: single-step metal affinity chromatography (MAC) using zinc as a chelator and ion exchange chromatography (IEX). The purified and inactivated beta-toxoids were then administered to sheep and chickens in order to produce IgG and IgY.

Results

All assays using the IEX failed. In contrast, MAC purified more than 21 mg of toxin per run in a single-step protocol. The purified and inactivated beta-toxoids were then administered to sheep and chickens, and IgG and IgY were purified with a high yield, medium antibody titer of 50 IU/mL, and high avidity (73.2 %).

Conclusions

C. perfringens type C beta-toxin and sheep or chicken anti-beta toxin IgG and IgY antibodies were successfully produced and purified using a simple protocol. This protocol can be used for the production of components used in the diagnosis and research of necrotic enteritis caused by C. perfringens type C, as well as for the evaluation of existing vaccines and the development of new preventive methods against this disease.

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生产和纯化 C 型产气荚膜梭菌 beta 型毒素以及 IgG 和 IgY 抗毒素

目的本研究旨在生产和纯化 C 型产气荚膜梭菌 beta-毒素、绵羊抗 beta-毒素免疫球蛋白 G (IgG) 和鸡免疫球蛋白 Y (IgY)。方法采用两种方法纯化 beta-毒素：以锌为螯合剂的单步金属亲和层析法 (MAC) 和离子交换层析法 (IEX)。然后将纯化和灭活的 beta 毒素注射给羊和鸡，以产生 IgG 和 IgY。相比之下，MAC 在单步方案中每次运行可纯化超过 21 毫克的毒素。然后将纯化和灭活的 C 型毒素给绵羊和鸡注射，纯化出的 IgG 和 IgY 产率高，抗体滴度为 50 IU/mL，亲和力高（73.2%）。该方案可用于生产诊断和研究 C 型产气荚膜杆菌引起的坏死性肠炎的成分，也可用于评估现有疫苗和开发预防该疾病的新方法。

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