SpyDirect: A Novel Biofunctionalization Method for High Stability and Longevity of Electronic Biosensors

Author:

Guo Keying1ORCID,Grünberg Raik1ORCID,Ren Yuxiang1,Chang Tianrui1,Wustoni Shofarul1ORCID,Strnad Ondrej2ORCID,Koklu Anil1ORCID,Díaz‐Galicia Escarlet1ORCID,Agudelo Jessica Parrado1ORCID,Druet Victor1ORCID,Castillo Tania Cecilia Hidalgo1,Moser Maximilian3ORCID,Ohayon David1ORCID,Hama Adel1ORCID,Dada Ashraf4ORCID,McCulloch Iain3ORCID,Viola Ivan2ORCID,Arold Stefan T.15ORCID,Inal Sahika1ORCID

Affiliation:

1. Computational Bioscience Research Center (CBRC), Biological and Environmental Science and Engineering King Abdullah University of Science and Technology (KAUST) Thuwal 23955‐6900 Saudi Arabia

2. Computer, Electrical and Mathematical Science and Engineering KAUST Thuwal 23955‐6900 Saudi Arabia

3. Department of Chemistry University of Oxford Oxford OX1 3TA UK

4. King Faisal Specialist Hospital & Research Centre (KFSH‐RC) Jeddah 21499 Saudi Arabia

5. Centre de Biologie Structurale (CBS), INSERM, CNRS Université de Montpellier Montpellier F‐34090 France

Abstract

AbstractElectronic immunosensors are indispensable tools for diagnostics, particularly in scenarios demanding immediate results. Conventionally, these sensors rely on the chemical immobilization of antibodies onto electrodes. However, globular proteins tend to adsorb and unfold on these surfaces. Therefore, self‐assembled monolayers (SAMs) of thiolated alkyl molecules are commonly used for indirect gold–antibody coupling. Here, a limitation associated with SAMs is revealed, wherein they curtail the longevity of protein sensors, particularly when integrated into the state‐of‐the‐art transducer of organic bioelectronics—the organic electrochemical transistor. The SpyDirect method is introduced, generating an ultrahigh‐density array of oriented nanobody receptors stably linked to the gold electrode without any SAMs. It is accomplished by directly coupling cysteine‐terminated and orientation‐optimized spyTag peptides, onto which nanobody‐spyCatcher fusion proteins are autocatalytically attached, yielding a dense and uniform biorecognition layer. The structure‐guided design optimizes the conformation and packing of flexibly tethered nanobodies. This biolayer enhances shelf‐life and reduces background noise in various complex media. SpyDirect functionalization is faster and easier than SAM‐based methods and does not necessitate organic solvents, rendering the sensors eco‐friendly, accessible, and amenable to scalability. SpyDirect represents a broadly applicable biofunctionalization method for enhancing the cost‐effectiveness, sustainability, and longevity of electronic biosensors, all without compromising sensitivity.

Funder

King Abdullah University of Science and Technology

Publisher

Wiley

Subject

General Physics and Astronomy,General Engineering,Biochemistry, Genetics and Molecular Biology (miscellaneous),General Materials Science,General Chemical Engineering,Medicine (miscellaneous)

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