{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,12,16]],"date-time":"2025-12-16T12:59:46Z","timestamp":1765889986851,"version":"3.48.0"},"reference-count":28,"publisher":"MDPI AG","issue":"12","license":[{"start":{"date-parts":[[2025,12,16]],"date-time":"2025-12-16T00:00:00Z","timestamp":1765843200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100004663","name":"National Key Research and Development Program of China","doi-asserted-by":"publisher","award":["2023YFF1304900"],"award-info":[{"award-number":["2023YFF1304900"]}],"id":[{"id":"10.13039\/501100004663","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100008871","name":"Yunnan Provincial Science and Technology Department","doi-asserted-by":"publisher","award":["202403AC100010"],"award-info":[{"award-number":["202403AC100010"]}],"id":[{"id":"10.13039\/501100008871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100008871","name":"Yunnan Provincial Science and Technology Department","doi-asserted-by":"publisher","award":["202301AT070343"],"award-info":[{"award-number":["202301AT070343"]}],"id":[{"id":"10.13039\/501100008871","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Toxins"],"abstract":"<jats:p>The preparation of an antibody to treat snake envenomation requires a large amount of snake venom. In China, only four types of anti-snake venom sera are clinically available, and the production and immunization strategies for clinically approved anti-snake venom sera still mainly rely on detoxified antigens, which is a mature technical route commonly adopted by domestic pharmaceutical enterprises. At present, researchers immunize animals with low doses of certain snake venom toxic components or prokaryotically expressed toxic components to reduce the amount of venom needed, and use prepared antisera for their specific investigation purposes. However, it is unclear if low-dose immunized antibody titers and toxin-neutralizing activities are consistent with those of high-dose detoxified crude venom immunized antibodies. In this study, we developed a method for the preparation of highly effective rabbit polyclonal antisera while saving a large amount of toxin. Rabbit polyclonal antisera prepared by low-dose natural \u03b1-bungarotoxin (\u03b1-BGT) had strong neutralizing effects on the toxin itself and achieved the same antibody titers as antisera prepared with high doses of detoxified \u03b1-BGT. Antigen of A maltose binding protein (MBP) fused with \u03b1-BGT (MBP-\u03b1-BGT) expressed in prokaryotes had low antibody titer and low neutralizing activity. This study provides an effective dosage selection guide and methods for the preparation of polyclonal antibodies and antiserum for investigation purposes.<\/jats:p>","DOI":"10.3390\/toxins17120601","type":"journal-article","created":{"date-parts":[[2025,12,16]],"date-time":"2025-12-16T12:47:26Z","timestamp":1765889246000},"page":"601","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Characterization of Alpha-Bungarotoxin Antibodies Prepared by Different Strategies"],"prefix":"10.3390","volume":"17","author":[{"given":"Huijuan","family":"Lu","sequence":"first","affiliation":[{"name":"Engineering Laboratory of Peptides of Chinese Academy of Sciences, Key Laboratory of Bioactive Peptides of Yunnan Province and State Key Laboratory of Genetic Evolution & Animal Models, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming 650201, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Guowen","family":"Zhang","sequence":"additional","affiliation":[{"name":"Engineering Laboratory of Peptides of Chinese Academy of Sciences, Key Laboratory of Bioactive Peptides of Yunnan Province and State Key Laboratory of Genetic Evolution & Animal Models, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming 650201, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lin","family":"Zhao","sequence":"additional","affiliation":[{"name":"Engineering Laboratory of Peptides of Chinese Academy of Sciences, Key Laboratory of Bioactive Peptides of Yunnan Province and State Key Laboratory of Genetic Evolution & Animal Models, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming 650201, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ying","family":"Yuan","sequence":"additional","affiliation":[{"name":"Department of Emergency, The First People\u2019s Hospital of Yunnan Province, The Affiliated Hospital of Kunming University of Science and Technology, Kunming 650034, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0007-3378-3071","authenticated-orcid":false,"given":"Bing","family":"Gong","sequence":"additional","affiliation":[{"name":"Department of Emergency, The First People\u2019s Hospital of Yunnan Province, The Affiliated Hospital of Kunming University of Science and Technology, Kunming 650034, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Bin","family":"Han","sequence":"additional","affiliation":[{"name":"Department of Emergency, The First People\u2019s Hospital of Yunnan Province, The Affiliated Hospital of Kunming University of Science and Technology, Kunming 650034, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2621-5782","authenticated-orcid":false,"given":"Wen-Hui","family":"Lee","sequence":"additional","affiliation":[{"name":"Engineering Laboratory of Peptides of Chinese Academy of Sciences, Key Laboratory of Bioactive Peptides of Yunnan Province and State Key Laboratory of Genetic Evolution & Animal Models, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming 650201, China"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,12,16]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Kasturiratne, A., Wickremasinghe, A.R., de Silva, N., Gunawardena, N.K., Pathmeswaran, A., Premaratna, R., Savioli, L., Lalloo, D.G., and de Silva, H.J. (2008). The global burden of snakebite: A literature analysis and modelling based on regional estimates of envenoming and deaths. PLoS Med., 5.","DOI":"10.1371\/journal.pmed.0050218"},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Lin, B., Zhang, J.R., Lu, H.J., Zhao, L., Chen, J., Zhang, H.F., Wei, X.S., Zhang, L.Y., Wu, X.B., and Lee, W.H. (2020). Immunoreactivity and neutralization study of Chinese Bungarus multicinctus antivenin and lab-prepared anti-bungarotoxin antisera towards purified bungarotoxins and snake venoms. PLoS Negl. Trop. Dis., 14.","DOI":"10.1371\/journal.pntd.0008873"},{"key":"ref_3","first-page":"518","article-title":"Clinical features of 60 consecutive ICU-treated patients envenomed by Bungarus multicinctus","volume":"40","author":"Hung","year":"2009","journal-title":"Southeast Asian J. Trop. Med. Public Health"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"609","DOI":"10.1016\/S0041-0101(98)00199-8","article-title":"Primary structure of g-bungarotoxin, a new postsynaptic neurotoxin from venom of Bungarus multicinctus","volume":"37","author":"Aird","year":"1999","journal-title":"Toxicon"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"235","DOI":"10.1146\/annurev.bi.42.070173.001315","article-title":"Neurotoxins of animal venoms: Snakes","volume":"42","author":"Tu","year":"1973","journal-title":"Annu. Rev. Biochem."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"113","DOI":"10.3109\/10409238309102792","article-title":"Conformational properties of the neurotoxins and cytotoxins isolated from Elapid snake venoms","volume":"14","author":"Dufton","year":"1983","journal-title":"CRC Crit. Rev. Biochem."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"2400","DOI":"10.1073\/pnas.77.5.2400","article-title":"Three-dimensional structure of the \u201clong\u201d neurotoxin from cobra venom","volume":"77","author":"Walkinshaw","year":"1980","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1254\/jphs.94.1","article-title":"Three-Finger \u03b1-Neurotoxins and the Nicotinic Acetylcholine Receptor, Forty Years On","volume":"94","author":"Nirthanan","year":"2004","journal-title":"J. Pharmacol. Sci."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"e20210040","DOI":"10.1590\/1678-9199-jvatitd-2021-0040","article-title":"Compositional and toxicological investigation of pooled venom from farm-raised Naja atra","volume":"28","author":"Xiao","year":"2022","journal-title":"J. Venom Anim. Toxins Incl. Trop. Dis."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"105311","DOI":"10.1016\/j.actatropica.2019.105311","article-title":"Geographical variations in king cobra (Ophiophagus hannah) venom from Thailand, Malaysia, Indonesia and China: On venom lethality, antivenom immunoreactivity and in vivo neutralization","volume":"203","author":"Tan","year":"2020","journal-title":"Acta Trop."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"393","DOI":"10.1007\/s13181-010-0051-4","article-title":"A controlled clinical trial of a novel antivenom in patients envenomed by Bungarus multicinctus","volume":"6","author":"Ha","year":"2010","journal-title":"J. Med. Toxicol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"108937","DOI":"10.1016\/j.biocon.2020.108937","article-title":"Will the COVID-19 outbreak be a turning point for China\u2019s wildlife protection: New developments and challenges of wildlife conservation in China","volume":"254","author":"Huang","year":"2021","journal-title":"Biol. Conserv."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"46","DOI":"10.1007\/s40656-021-00406-6","article-title":"COVID-19, a critical juncture in China\u2019s wildlife protection?","volume":"43","author":"Xu","year":"2021","journal-title":"Hist. Philos. Life Sci."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Ratanabanangkoon, K., Tan, K.Y., Eursakun, S., Tan, C.H., Simsiriwong, P., Pamornsakda, T., Wiriyarat, W., Klinpayom, C., and Tan, N.H. (2016). A Simple and Novel Strategy for the Production of a Pan-specific Antiserum against Elapid Snakes of Asia. PLoS Negl. Trop. Dis., 10.","DOI":"10.1371\/journal.pntd.0004565"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"17063","DOI":"10.1038\/nrdp.2017.63","article-title":"Snakebite envenoming","volume":"3","author":"Calvete","year":"2017","journal-title":"Nat. Rev. Dis. Primers"},{"key":"ref_16","unstructured":"Bon, C., Burnouf, T., Gutierrez, J.M., Padilla, A., and Warrell, D.A. (2013, October 30). WHO Guidelines for the Production, Control and Regulations of Snake Antivenoms Immunoglobulins. Available online: https:\/\/www.who.int\/publications\/m\/item\/snake-antivenom-immunoglobulins-annex-5-trs-no-1004."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1405","DOI":"10.1016\/j.toxicon.2010.02.012","article-title":"Antivenom: The most cost-effective treatment in the world?","volume":"55","author":"Brown","year":"2010","journal-title":"Toxicon"},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Gutierrez, J.M. (2018). Global Availability of Antivenoms: The Relevance of Public Manufacturing Laboratories. Toxins, 11.","DOI":"10.3390\/toxins11010005"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"526","DOI":"10.2471\/BLT.13.132431","article-title":"A multicomponent strategy to improve the availability of antivenom for treating snakebite envenoming","volume":"92","author":"Burnouf","year":"2014","journal-title":"Bull World Health Organ."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"30","DOI":"10.1016\/j.toxicon.2006.09.005","article-title":"Strengthening antivenom production in Central and South American public laboratories: Report of a workshop","volume":"49","author":"Higashi","year":"2007","journal-title":"Toxicon"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"61","DOI":"10.1051\/jbio\/2009042","article-title":"Emergency immunotherapy: Snake and scorpion antivenoms","volume":"204","author":"Chippaux","year":"2010","journal-title":"Biol. Aujourdhui."},{"key":"ref_22","first-page":"1","article-title":"Progress and improvement of the manufacturing process of snake antivenom","volume":"68","author":"Zolfagharina","year":"2013","journal-title":"Arch. Razi Inst."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1016\/j.jprot.2016.02.028","article-title":"Proteomic characterization and comparison of venoms from two elapid snakes (Bungarus multicinctus and Naja atra) from China","volume":"138","author":"Shan","year":"2016","journal-title":"J. Proteomics"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"76","DOI":"10.1016\/0003-2697(85)90442-7","article-title":"Measurement of protein using bicinchoninic acid","volume":"150","author":"Smith","year":"1985","journal-title":"Anal. Biochem."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"659","DOI":"10.1096\/fj.07-9574LSF","article-title":"Dose translation from animal to human studies revisited","volume":"22","author":"Nihal","year":"2008","journal-title":"FASEB J."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"680","DOI":"10.1038\/227680a0","article-title":"Cleavage of structural proteins during the assembly of the head of bacteriophage T4","volume":"227","author":"Laemmli","year":"1970","journal-title":"Nature"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"299","DOI":"10.1016\/j.toxicon.2016.06.023","article-title":"Rapid and selective detection of experimental snake envenomation\u2014Use of gold nanoparticle based lateral flow assay","volume":"119","author":"Pawade","year":"2016","journal-title":"Toxicon"},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Wang, Y., Zhang, J., Zhang, D., Xiao, H., Xiong, S., and Huang, C. (2018). Exploration of the Inhibitory Potential of Varespladib for Snakebite Envenomation. Molecules, 23.","DOI":"10.3390\/molecules23020391"}],"container-title":["Toxins"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2072-6651\/17\/12\/601\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,12,16]],"date-time":"2025-12-16T12:55:01Z","timestamp":1765889701000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2072-6651\/17\/12\/601"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,12,16]]},"references-count":28,"journal-issue":{"issue":"12","published-online":{"date-parts":[[2025,12]]}},"alternative-id":["toxins17120601"],"URL":"https:\/\/doi.org\/10.3390\/toxins17120601","relation":{},"ISSN":["2072-6651"],"issn-type":[{"value":"2072-6651","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,12,16]]}}}