Abbasali Salarifar, Pooria Safarzadeh Kozani, Mohammad Javad Rasaee
{"title":"制备免疫球蛋白不同化学分馏方法的比较。","authors":"Abbasali Salarifar, Pooria Safarzadeh Kozani, Mohammad Javad Rasaee","doi":"10.1080/15321819.2025.2450664","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Application of antibodies in therapeutics and diagnostics are growing Continually. Herein, we aimed to find the most qualified immunoglobulin (Ig) chemical preparation method.</p><p><strong>Methods: </strong>A rabbit was immunized against recombinant SARS-CoV-2 nucleocapsid (NP) and reactive polyclonal antibodies were prepared using the ammonium sulfate (AS), caprylic acid (CA), polyethylene glycol (PEG), and caprylic acid/ammonium sulfate (CA/AS) methods. Different antibody solutions were analyzed by SDS-PAGE and subsequently quantified by ImageJ software for further analysis in terms of Ig purity, Ig recovery, and albumin impurity. Ultimately, the prepared antibodies were assessed via Western blotting and ELISA to evaluate their ability to bind NP.</p><p><strong>Results: </strong>Prepared Ig solutions via the CA/AS method had the highest Ig purity (followed by CA, PEG, and AS) and lowest albumin impurity (followed by CA, AS, and PEG). The PEG method had the highest recovery followed by AS, CA, and CA/AS methods. Moreover, antibodies prepared via different methods demonstrated comparable binding capacities to NP in ELISA and Western blotting.</p><p><strong>Conclusions: </strong>CA/AS, closely followed by CA, proved to be the most qualified method for the preparation of Ig yielding the highest Ig purity while the PEG method resulted in the highest Ig recovery rate.</p>","PeriodicalId":15990,"journal":{"name":"Journal of immunoassay & immunochemistry","volume":" ","pages":"169-185"},"PeriodicalIF":0.0000,"publicationDate":"2025-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"A comparison between different chemical fractionation methods for immunoglobulin preparation.\",\"authors\":\"Abbasali Salarifar, Pooria Safarzadeh Kozani, Mohammad Javad Rasaee\",\"doi\":\"10.1080/15321819.2025.2450664\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>Application of antibodies in therapeutics and diagnostics are growing Continually. Herein, we aimed to find the most qualified immunoglobulin (Ig) chemical preparation method.</p><p><strong>Methods: </strong>A rabbit was immunized against recombinant SARS-CoV-2 nucleocapsid (NP) and reactive polyclonal antibodies were prepared using the ammonium sulfate (AS), caprylic acid (CA), polyethylene glycol (PEG), and caprylic acid/ammonium sulfate (CA/AS) methods. Different antibody solutions were analyzed by SDS-PAGE and subsequently quantified by ImageJ software for further analysis in terms of Ig purity, Ig recovery, and albumin impurity. Ultimately, the prepared antibodies were assessed via Western blotting and ELISA to evaluate their ability to bind NP.</p><p><strong>Results: </strong>Prepared Ig solutions via the CA/AS method had the highest Ig purity (followed by CA, PEG, and AS) and lowest albumin impurity (followed by CA, AS, and PEG). The PEG method had the highest recovery followed by AS, CA, and CA/AS methods. Moreover, antibodies prepared via different methods demonstrated comparable binding capacities to NP in ELISA and Western blotting.</p><p><strong>Conclusions: </strong>CA/AS, closely followed by CA, proved to be the most qualified method for the preparation of Ig yielding the highest Ig purity while the PEG method resulted in the highest Ig recovery rate.</p>\",\"PeriodicalId\":15990,\"journal\":{\"name\":\"Journal of immunoassay & immunochemistry\",\"volume\":\" \",\"pages\":\"169-185\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2025-03-04\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Journal of immunoassay & immunochemistry\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.1080/15321819.2025.2450664\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2025/1/11 0:00:00\",\"PubModel\":\"Epub\",\"JCR\":\"Q2\",\"JCRName\":\"Health Professions\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of immunoassay & immunochemistry","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1080/15321819.2025.2450664","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2025/1/11 0:00:00","PubModel":"Epub","JCR":"Q2","JCRName":"Health Professions","Score":null,"Total":0}
A comparison between different chemical fractionation methods for immunoglobulin preparation.
Background: Application of antibodies in therapeutics and diagnostics are growing Continually. Herein, we aimed to find the most qualified immunoglobulin (Ig) chemical preparation method.
Methods: A rabbit was immunized against recombinant SARS-CoV-2 nucleocapsid (NP) and reactive polyclonal antibodies were prepared using the ammonium sulfate (AS), caprylic acid (CA), polyethylene glycol (PEG), and caprylic acid/ammonium sulfate (CA/AS) methods. Different antibody solutions were analyzed by SDS-PAGE and subsequently quantified by ImageJ software for further analysis in terms of Ig purity, Ig recovery, and albumin impurity. Ultimately, the prepared antibodies were assessed via Western blotting and ELISA to evaluate their ability to bind NP.
Results: Prepared Ig solutions via the CA/AS method had the highest Ig purity (followed by CA, PEG, and AS) and lowest albumin impurity (followed by CA, AS, and PEG). The PEG method had the highest recovery followed by AS, CA, and CA/AS methods. Moreover, antibodies prepared via different methods demonstrated comparable binding capacities to NP in ELISA and Western blotting.
Conclusions: CA/AS, closely followed by CA, proved to be the most qualified method for the preparation of Ig yielding the highest Ig purity while the PEG method resulted in the highest Ig recovery rate.
期刊介绍：
The Journal of Immunoassay & Immunochemistry is an international forum for rapid dissemination of research results and methodologies dealing with all aspects of immunoassay and immunochemistry, as well as selected aspects of immunology. They include receptor assay, enzyme-linked immunosorbent assay (ELISA) in all of its embodiments, ligand-based assays, biological markers of ligand-receptor interaction, in vivo and in vitro diagnostic reagents and techniques, diagnosis of AIDS, point-of-care testing, clinical immunology, antibody isolation and purification, and others.
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