A CRISPR/Cas12a electrochemical biosensing to detect pig mtDNA D-loop for ensuring food authenticity
Ensuring safe and accurately labeled food products is crucial for communities worldwide. One significant concern in food safety is the unintended inclusion of pork in products, which impacts food labeling, religious dietary practices, legal standards, and public health. Detecting such contamination...
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Elsevier
2025-02-01
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| Series: | Sensing and Bio-Sensing Research |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2214180425000212 |
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| author | Muhammad Ihda H.L. Zein Clianta Yudin Kharismasari Ari Hardianto Salma Nur Zakiyyah Riezki Amalia Mehmet Ozsoz Mara Mirasoli Irkham Yeni Wahyuni Hartati |
| author_facet | Muhammad Ihda H.L. Zein Clianta Yudin Kharismasari Ari Hardianto Salma Nur Zakiyyah Riezki Amalia Mehmet Ozsoz Mara Mirasoli Irkham Yeni Wahyuni Hartati |
| author_sort | Muhammad Ihda H.L. Zein |
| collection | DOAJ |
| description | Ensuring safe and accurately labeled food products is crucial for communities worldwide. One significant concern in food safety is the unintended inclusion of pork in products, which impacts food labeling, religious dietary practices, legal standards, and public health. Detecting such contamination is challenging due to complex food processing, requiring advanced methods for accurate identification. A CRISPR/Cas12a-based electrochemical biosensor has garnered attention for its rapidity, portability, high sensitivity, and specificity, leveraging trans-cleavage activity for precise detection. This study utilized a modified electrode, SPCE/Ceria/STV/Biotin-ssDNA-Methylene Blue (MB), characterized by SEM/EDX and voltammetry methods to examine morphology and electrochemical behavior. Optimization using the Box-Behnken Design (BBD) yielded ideal conditions: 30 min STV incubation, 1.5 μM probe concentration, 240 min probe incubation, 0.1 % BSA concentration, 30 min target incubation, and 2 μL target volume. These parameters enabled the development of an ultrasensitive biosensor with detection and quantification limits of 4 fM and 71 fM, respectively. Operating within a range of 10 nM to 100 fM, the biosensor achieved 1.1 % RSD. This CRISPR-based biosensor successfully detected pig mtDNA in raw and processed meat samples, achieving a 100.82 % recovery rate for corned samples and maintaining stability for 14 days. These findings highlight the biosensor's potential as a rapid, stable, ultrasensitive, and ultraspecific alternative for food monitoring. Its robustness makes it particularly suitable for ensuring the authenticity of animal-derived food products susceptible to counterfeiting. |
| format | Article |
| id | doaj-art-da85ded23183454bad5627fa0defa57e |
| institution | Kabale University |
| issn | 2214-1804 |
| language | English |
| publishDate | 2025-02-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Sensing and Bio-Sensing Research |
| spelling | doaj-art-da85ded23183454bad5627fa0defa57e2025-02-06T05:11:46ZengElsevierSensing and Bio-Sensing Research2214-18042025-02-0147100755A CRISPR/Cas12a electrochemical biosensing to detect pig mtDNA D-loop for ensuring food authenticityMuhammad Ihda H.L. Zein0Clianta Yudin Kharismasari1Ari Hardianto2Salma Nur Zakiyyah3Riezki Amalia4Mehmet Ozsoz5Mara Mirasoli6 Irkham7Yeni Wahyuni Hartati8Department of Chemistry, Faculty of Mathematics and Natural Sciences, Universitas Padjadjaran, Bandung 45363, Indonesia; Department of Chemistry “Giacomo Ciamician”, Alma Mater Studiorum - University of Bologna, Bologna 40126, ItalyDepartment of Chemistry, Faculty of Mathematics and Natural Sciences, Universitas Padjadjaran, Bandung 45363, IndonesiaDepartment of Chemistry, Faculty of Mathematics and Natural Sciences, Universitas Padjadjaran, Bandung 45363, IndonesiaDepartment of Chemistry, Faculty of Mathematics and Natural Sciences, Universitas Padjadjaran, Bandung 45363, IndonesiaDepartment of Pharmacology and Clinical Pharmacy, Universitas Padjadjaran, Bandung 45363, IndonesiaDepartment of Chemistry, Faculty of Mathematics and Natural Sciences, Universitas Padjadjaran, Bandung 45363, Indonesia; Department of Biomedical Engineering, Near East University, Mersin 99138, TurkeyDepartment of Chemistry “Giacomo Ciamician”, Alma Mater Studiorum - University of Bologna, Bologna 40126, ItalyDepartment of Chemistry, Faculty of Mathematics and Natural Sciences, Universitas Padjadjaran, Bandung 45363, Indonesia; Corresponding authors.Department of Chemistry, Faculty of Mathematics and Natural Sciences, Universitas Padjadjaran, Bandung 45363, Indonesia; Corresponding authors.Ensuring safe and accurately labeled food products is crucial for communities worldwide. One significant concern in food safety is the unintended inclusion of pork in products, which impacts food labeling, religious dietary practices, legal standards, and public health. Detecting such contamination is challenging due to complex food processing, requiring advanced methods for accurate identification. A CRISPR/Cas12a-based electrochemical biosensor has garnered attention for its rapidity, portability, high sensitivity, and specificity, leveraging trans-cleavage activity for precise detection. This study utilized a modified electrode, SPCE/Ceria/STV/Biotin-ssDNA-Methylene Blue (MB), characterized by SEM/EDX and voltammetry methods to examine morphology and electrochemical behavior. Optimization using the Box-Behnken Design (BBD) yielded ideal conditions: 30 min STV incubation, 1.5 μM probe concentration, 240 min probe incubation, 0.1 % BSA concentration, 30 min target incubation, and 2 μL target volume. These parameters enabled the development of an ultrasensitive biosensor with detection and quantification limits of 4 fM and 71 fM, respectively. Operating within a range of 10 nM to 100 fM, the biosensor achieved 1.1 % RSD. This CRISPR-based biosensor successfully detected pig mtDNA in raw and processed meat samples, achieving a 100.82 % recovery rate for corned samples and maintaining stability for 14 days. These findings highlight the biosensor's potential as a rapid, stable, ultrasensitive, and ultraspecific alternative for food monitoring. Its robustness makes it particularly suitable for ensuring the authenticity of animal-derived food products susceptible to counterfeiting.http://www.sciencedirect.com/science/article/pii/S2214180425000212CRISPR/Cas12aElectrochemical biosensorFood monitoringOptimizationBBD (box-Benhken design) |
| spellingShingle | Muhammad Ihda H.L. Zein Clianta Yudin Kharismasari Ari Hardianto Salma Nur Zakiyyah Riezki Amalia Mehmet Ozsoz Mara Mirasoli Irkham Yeni Wahyuni Hartati A CRISPR/Cas12a electrochemical biosensing to detect pig mtDNA D-loop for ensuring food authenticity Sensing and Bio-Sensing Research CRISPR/Cas12a Electrochemical biosensor Food monitoring Optimization BBD (box-Benhken design) |
| title | A CRISPR/Cas12a electrochemical biosensing to detect pig mtDNA D-loop for ensuring food authenticity |
| title_full | A CRISPR/Cas12a electrochemical biosensing to detect pig mtDNA D-loop for ensuring food authenticity |
| title_fullStr | A CRISPR/Cas12a electrochemical biosensing to detect pig mtDNA D-loop for ensuring food authenticity |
| title_full_unstemmed | A CRISPR/Cas12a electrochemical biosensing to detect pig mtDNA D-loop for ensuring food authenticity |
| title_short | A CRISPR/Cas12a electrochemical biosensing to detect pig mtDNA D-loop for ensuring food authenticity |
| title_sort | crispr cas12a electrochemical biosensing to detect pig mtdna d loop for ensuring food authenticity |
| topic | CRISPR/Cas12a Electrochemical biosensor Food monitoring Optimization BBD (box-Benhken design) |
| url | http://www.sciencedirect.com/science/article/pii/S2214180425000212 |
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