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The current treatment, horse serum-derived antivenom, has unpredictable side effects and presents manufacturing challenges. This study focused on developing new-generation snake venom antidotes by using microbial phage display technology to derive nanobodies from an alpaca immunized with attenuated\n              <jats:italic>N. atra<\/jats:italic>\n              venom. Following confirmation of the immune response in the alpaca, we amplified\n              <jats:italic>\n                V\n                <jats:sub>H<\/jats:sub>\n                H\n              <\/jats:italic>\n              genes from isolated peripheral blood mononuclear cells and constructed a phage display\n              <jats:italic>\n                V\n                <jats:sub>H<\/jats:sub>\n                H\n              <\/jats:italic>\n              library of 1.0 \u00d7 10\n              <jats:sup>7<\/jats:sup>\n              transformants. After four rounds of biopanning, the enriched phages exhibited increased binding activity to\n              <jats:italic>N. atra<\/jats:italic>\n              venom. Four nanobody clones with high binding affinities were selected: aNAH1, aNAH6, aNAH7, and aNAH9. Specificity testing against venom from various snake species, including two Southeast Asian cobra species, revealed nanobodies specific to the genus\n              <jats:italic>Naja<\/jats:italic>\n              . An\n              <jats:italic>in vivo<\/jats:italic>\n              mouse venom neutralization assay demonstrated that all nanobodies prolonged mouse survival and aNAH6 protected 66.6% of the mice from the lethal dosage. These findings highlight the potential of phage display-derived nanobodies as valuable antidotes for\n              <jats:italic>N. atra<\/jats:italic>\n              venom, laying the groundwork for future applications in snakebite treatment.\n            <\/jats:p>\n            <jats:sec>\n              <jats:title>IMPORTANCE<\/jats:title>\n              <jats:p>Chinese cobra venom bites present a formidable medical challenge, and current serum treatments face unresolved issues. Our research applied microbial phage display technology to obtain a new, effective, and cost-efficient treatment approach. Despite interest among scientists in utilizing this technology to screen alpaca antibodies against toxins, the available literature is limited. This study makes a significant contribution by introducing neutralizing antibodies that are specifically tailored to Chinese cobra venom. We provide a comprehensive and unbiased account of the antibody construction process, accompanied by thorough testing of various nanobodies and an assessment of cross-reactivity with diverse snake venoms. These nanobodies represent a promising avenue for targeted antivenom development that bridges microbiology and biotechnology to address critical health needs.<\/jats:p>\n            <\/jats:sec>\n          <\/jats:sec>","DOI":"10.1128\/aem.00121-24","type":"journal-article","created":{"date-parts":[[2024,7,9]],"date-time":"2024-07-09T13:00:46Z","timestamp":1720530046000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":6,"title":["Phage display-derived alpaca nanobodies as potential therapeutics for\n            <i>Naja atra<\/i>\n            snake envenomation"],"prefix":"10.1128","volume":"90","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-5452-5537","authenticated-orcid":false,"given":"Wei-Chu","family":"Wang","sequence":"first","affiliation":[{"name":"Ph.D. Program in Medical Biotechnology, College of Medical Science and Technology, Taipei Medical University, Taipei, 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