{"title":"Multiplexed Biomarker Detection Using DNA Payloads: Design, Assembly, and Analysis.","authors":"Matthew Aquilina, Katherine E Dunn","doi":"10.1007/978-1-0716-4394-5_16","DOIUrl":null,"url":null,"abstract":"<p><p>Most biomarker assays are typically designed to detect a single molecule type (DNA, RNA, proteins, etc.) in a single assay. This means that monitoring a diverse biomarker panel could quickly become a complex endeavor, requiring different techniques, labs, and expertise. In this chapter, we describe a method for multiplexed biomarker detection from a single sample using variable-length DNA payload chains as the output signal. Through the course of the assay, payloads are systematically disassembled in the presence of specific biomarkers. The resulting distinctly sized fragments then yield characteristic gel electrophoresis band patterns, which can be detected and quantified using image analysis algorithms. We detail the entire process for constructing DNA payloads and conducting a biomarker detection assay, including the sequence design, laboratory assembly, running the assay, and final image analysis.</p>","PeriodicalId":18490,"journal":{"name":"Methods in molecular biology","volume":"2901 ","pages":"203-226"},"PeriodicalIF":0.0000,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Methods in molecular biology","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1007/978-1-0716-4394-5_16","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"Biochemistry, Genetics and Molecular Biology","Score":null,"Total":0}
引用次数: 0
Abstract
Most biomarker assays are typically designed to detect a single molecule type (DNA, RNA, proteins, etc.) in a single assay. This means that monitoring a diverse biomarker panel could quickly become a complex endeavor, requiring different techniques, labs, and expertise. In this chapter, we describe a method for multiplexed biomarker detection from a single sample using variable-length DNA payload chains as the output signal. Through the course of the assay, payloads are systematically disassembled in the presence of specific biomarkers. The resulting distinctly sized fragments then yield characteristic gel electrophoresis band patterns, which can be detected and quantified using image analysis algorithms. We detail the entire process for constructing DNA payloads and conducting a biomarker detection assay, including the sequence design, laboratory assembly, running the assay, and final image analysis.
期刊介绍：
For over 20 years, biological scientists have come to rely on the research protocols and methodologies in the critically acclaimed Methods in Molecular Biology series. The series was the first to introduce the step-by-step protocols approach that has become the standard in all biomedical protocol publishing. Each protocol is provided in readily-reproducible step-by-step fashion, opening with an introductory overview, a list of the materials and reagents needed to complete the experiment, and followed by a detailed procedure that is supported with a helpful notes section offering tips and tricks of the trade as well as troubleshooting advice.
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