Tropomyosin is the major and predominant allergen among shellfish. This study developed an ultrasensitive immuno-PCR method for the quantification of crustacean tropomyosin in foods. The method couples sandwich ELISA with the real-time PCR (rtPCR) amplification of marker DNAs. Monoclonal anti-TPM antibody was the capture antibody, polyclonal rabbit anti-shrimp tropomyosin antibody was the detection antibody, while natural shrimp tropomyosin served as the standard. A double-stranded amino-DNA was covalently conjugated to a secondary anti-rabbit antibody and subsequently amplified and quantified via rtPCR. The quantification sensitivity of immuno-PCR was 20-fold higher than analogous ELISA, with LOQ 19.8 pg/mL. The developed immuno-PCR method is highly specific for the detection of crustacean tropomyosin and is highly precise in a broad concentration range. Tropomyosin recovery in the spiked vegetable soup was 87.7–115.6%. Crustacean tropomyosin was also quantified in commercial food products. The reported immuno-PCR assay is the most sensitive method for the quantification of crustacean tropomyosin and is the first immuno-PCR-based assay for the quantification of food allergen and food protein in general. The described method could be easily adapted for the specific and ultrasensitive immuno-PCR-based detection of traces of any food allergen that is currently being quantified with ELISA, which is of critical importance for people with food allergies.","AbstractOtherLang":null,"BibLvlCode":"AS","StandardTitle":"Ultrasensitive quantification of crustacean tropomyosin by immuno-PCR","OrigTitleLangCode":"en","OrigTitleLangCodeExtended":"eng","OrigTitleLangID":15,"DateLastModified":{"date":"2024-12-10 01:33:17.368041","timezone_type":1,"timezone":"+01:00"},"UserAccessRight":null,"UserAccID":null,"AuthorKeywords":"tropomyosin; immuno-PCR; crustacean allergen; ELISA; shellfish allergen; allergen quantification","OtherDescriptors":null,"Notes":null,"AnaPub":2023,"MonPub":null,"DateUpdate":"2024-03-26","DateCreate":"2024-03-26","SecASFANote":null,"ConfID":null,"PeerRev":1,"VlizCoreFlag":1,"WoScode":"WOS:001095516700001","VABBcode":null,"OpenAcc":1,"DOI":"10.3390/ijms242015410"},"refs":null,"anarec":{"AnaID":391473,"PubliDate":2023,"Pagination":"15410","XtraPublOfAnaID":null,"ISBN":null,"Volume":"24","Issue":"20","BRefMon":null,"BRefMonRR":null,"BRefXtra":null,"BRefXtraRR":null,"SerBRefID":270393,"SerRR":"International Journal of Molecular Sciences. 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