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Here, we report an entropy-guided aptamer engineering strategy that removes redundant regions while preserving essential stem-loop recognition motifs, optimizing the thermodynamic balance of target binding, and yielding truncated aptamers with 3.3- and 3.6-fold improved SGI-derived affinities toward PHD and \u03b1-AMT, respectively. The engineered aptamers were integrated into a dual-mode luminescence resonance energy transfer (LRET)\/surface-enhanced Raman scattering (SERS) platform comprising upconversion nanoparticles (UCNPs), and Fe3O4@AuNS. A single target-binding-driven aptamer-cDNA structural-switching event synchronously regulated both optical outputs, enabling internally cross-validated multiplex quantification with low-pg\/mL detection limits over 0.01\u2013100 ng\/mL (R2 &amp;gt; 0.99), recoveries of 92.0\u201399.2%, and relative standard deviations below 5% in mushroom samples. This strategy provides a generalizable framework for sensitive detection of structurally constrained food toxins and multiplex food safety analysis.<\/jats:p>","DOI":"10.1021\/acs.jafc.6c07518","type":"journal-article","created":{"date-parts":[[2026,9,1]],"date-time":"2026-09-01T09:55:25Z","timestamp":1788256525000},"page":"27904-27923","source":"Crossref","is-referenced-by-count":0,"title":["Entropy-Engineered Aptamer Truncation Enables Dual-Mode\nLRET\/SERS Aptasensing for Multiplex Detection of Phalloidin and \u03b1-Amanitin\nin Mushroom Samples"],"prefix":"10.1021","volume":"74","author":[{"given":"Ali","family":"Raza","sequence":"first","affiliation":[{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"State Administration for Market Regulation , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Faizan","family":"Ul Haq","sequence":"additional","affiliation":[{"name":"Yangzhou University , , , ,","place":["Yangzhou, Jiangsu, China, 225127"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Khubaib","family":"Ali","sequence":"additional","affiliation":[{"name":"Yangzhou University , , , ,","place":["Yangzhou, Jiangsu, China, 225127"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mehwish","family":"Mumtaz","sequence":"additional","affiliation":[{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2360-0987","authenticated-orcid":true,"given":"Muhammad","family":"Shoaib","sequence":"additional","affiliation":[{"name":"Zhejiang University , , 866 Yuhangtang Road , ,","place":["Hangzhou, China, 310058"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaodong","family":"Lu","sequence":"additional","affiliation":[{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"State Administration for Market Regulation , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yating","family":"Song","sequence":"additional","affiliation":[{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"State Administration for Market Regulation , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Li","family":"Ji","sequence":"additional","affiliation":[{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"State Administration for Market Regulation , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xi","family":"Yu","sequence":"additional","affiliation":[{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"State Administration for Market Regulation , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shuaishuai","family":"Wei","sequence":"additional","affiliation":[{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"State Administration for Market Regulation , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wenbo","family":"Gao","sequence":"additional","affiliation":[{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"State Administration for Market Regulation , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3868-8125","authenticated-orcid":true,"given":"Zhouping","family":"Wang","sequence":"additional","affiliation":[{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"State Administration for Market Regulation , , , ,","place":["Wuxi, Jiangsu, China, 214122"]},{"name":"Jiangnan University , , , ,","place":["Wuxi, Jiangsu, China, 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Rep."},{"key":"2026090917074896700_cit3","doi-asserted-by":"publisher","first-page":"107639","DOI":"10.1016\/j.toxicon.2024.107639","article-title":"Wild mushroom\npoisoning: A case study of amatoxin-containing mushrooms and implications\nfor public health","volume":"240","author":"He","year":"2024","journal-title":"Toxicon"},{"issue":"10","key":"2026090917074896700_cit4","doi-asserted-by":"publisher","first-page":"717","DOI":"10.1080\/15563650.2023.2268981","article-title":"2022 Annual report of the national poison data system\u00ae(NPDS)\nfrom America\u2019s Poison Centers\u00ae: 40th annual report","volume":"61","author":"Gummin","year":"2023","journal-title":"Clin. Toxicol."},{"issue":"14","key":"2026090917074896700_cit5","doi-asserted-by":"publisher","first-page":"5500","DOI":"10.1002\/anie.201914620","article-title":"A Convergent\nTotal Synthesis of the\nDeath Cap Toxin \u03b1-Amanitin","volume":"59","author":"Siegert","year":"2020","journal-title":"Angew. Chem.,\nInt. Ed."},{"key":"2026090917074896700_cit6","unstructured":"NCBI.\n          \n          PubChem. Compound Summary for CID 9543442, Alpha-Amanitin; National Center for Biotechnology Information, 2026."},{"issue":"44","key":"2026090917074896700_cit7","doi-asserted-by":"publisher","first-page":"27891","DOI":"10.1021\/acs.jafc.5c09325","article-title":"Recent Advances in Detection Methods\nand Toxicological Mechanisms of Amatoxins","volume":"73","author":"Tian","year":"2025","journal-title":"J.\nAgric. Food Chem."},{"key":"2026090917074896700_cit8","doi-asserted-by":"publisher","first-page":"104784","DOI":"10.1016\/j.tifs.2024.104784","article-title":"T-2 mycotoxin: From\noccurrence and toxicokinetics to recent advances in aptasensor-based\ndetection strategies and future perspectives for enhanced food safety","volume":"156","author":"Raza","year":"2025","journal-title":"Trends Food Sci. 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Model."},{"key":"2026090917074896700_cit20","doi-asserted-by":"publisher","first-page":"117947","DOI":"10.1016\/j.bios.2025.117947","article-title":"A cascade amplification platform integrating entropy-driven DNA nanomachine\nwith CRISPR\/Cas12a for microRNA-21 and Listeria monocytogenes detection","volume":"290","author":"Ding","year":"2025","journal-title":"Biosens. Bioelectron."},{"issue":"47","key":"2026090917074896700_cit21","doi-asserted-by":"publisher","first-page":"52976","DOI":"10.1021\/acsami.0c17200","article-title":"Application\nof NaYF4:Yb\/Er\/Tm UCNPs\nin Array-Based Sensing of Neurotransmitters: From a Single Particle\nto a Multichannel Sensor Array","volume":"12","author":"Abbasi-Moayed","year":"2020","journal-title":"ACS Appl. Mater.\nInterfaces"},{"issue":"9","key":"2026090917074896700_cit22","doi-asserted-by":"publisher","first-page":"1733","DOI":"10.1021\/acsaom.3c00194","article-title":"Toward Upconversion Luminescence\nEnhancement with Dye Sensitization","volume":"2","author":"Liu","year":"2024","journal-title":"ACS Appl.\nOpt. Mater."},{"key":"2026090917074896700_cit23","doi-asserted-by":"publisher","first-page":"141575","DOI":"10.1016\/j.foodchem.2024.141575","article-title":"Development of novel\nFRET aptasensor based on the quenching ability of iron oxide-gold\nnanostars for the detection of aflatoxin M1","volume":"464","author":"Haq","year":"2025","journal-title":"Food Chem."},{"issue":"23","key":"2026090917074896700_cit24","doi-asserted-by":"publisher","first-page":"12393","DOI":"10.1021\/acsanm.5c02344","article-title":"Multibranched\nMagnetic Core\u2013Shell Gold Nanostars for In Situ Solution-Based\nSERS Detection","volume":"8","author":"Atta","year":"2025","journal-title":"ACS Appl. 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Acta"},{"key":"2026090917074896700_cit34","doi-asserted-by":"publisher","first-page":"132171","DOI":"10.1016\/j.foodchem.2022.132171","article-title":"Aptamer truncation\nstrategy assisted by molecular docking and sensitive detection of\nT-2 toxin using SYBR Green I as a signal amplifier","volume":"381","author":"Ma","year":"2022","journal-title":"Food Chem."},{"issue":"3","key":"2026090917074896700_cit35","doi-asserted-by":"publisher","first-page":"130","DOI":"10.1007\/s00604-024-06199-2","article-title":"High-affinity truncated aptamers\nfor detection of Cronobacter spp\nwith magnetic separation-assisted DNAzyme-driven 3D DNA walker","volume":"191","author":"Yang","year":"2024","journal-title":"Microchim. 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Actuators, B"},{"issue":"19","key":"2026090917074896700_cit38","doi-asserted-by":"publisher","first-page":"2419885","DOI":"10.1002\/adfm.202419885","article-title":"Mesoporous-Confined\nAu Thorn Nano-Pockets Enable of Precisely Capturing\nTarget Molecules for Universal SERS Detection","volume":"35","author":"Liao","year":"2025","journal-title":"Adv. Funct. Mater."},{"key":"2026090917074896700_cit39","doi-asserted-by":"publisher","first-page":"280","DOI":"10.1016\/j.bios.2016.03.005","article-title":"Dual-excitation upconverting\nnanoparticle and quantum dot aptasensor for multiplexed food pathogen\ndetection","volume":"81","author":"Kurt","year":"2016","journal-title":"Biosens. Bioelectron."},{"issue":"2","key":"2026090917074896700_cit40","doi-asserted-by":"publisher","first-page":"441","DOI":"10.1039\/D2TB01701F","article-title":"Novel metal\nenhanced dual-mode fluorometric and SERS aptasensor incorporating\na heterostructure nanoassembly for ultrasensitive T-2 toxin detection","volume":"11","author":"Khan","year":"2023","journal-title":"J. Mater. Chem. B"},{"key":"2026090917074896700_cit41","doi-asserted-by":"publisher","DOI":"10.1002\/adma.202309625","article-title":"Advancing\nMycotoxin Detection in Food And Feed: Novel Insights from Surface-Enhanced\nRaman Spectroscopy (SERS)","volume":"36","author":"Logan","year":"2024","journal-title":"Adv. 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