Label-free electrochemical immunosensor for porcine gelatin using a boron-doped diamond electrode <i>via</i> diazonium salt electrografting
At a Glance
Section titled âAt a Glanceâ| Metadata | Details |
|---|---|
| Publication Date | 2025-06-09 |
| Journal | Journal of Electrochemical Science and Engineering |
| Authors | Irkham Irkham, Fadli Taufik Abdillah, Muhammad Ihda H.L. Zein, Adisyahputra, N. ZAHRA |
| Analysis | Full AI Review Included |
Technical Documentation & Analysis: Label-Free BDD Immunosensors for Food Authentication
Section titled âTechnical Documentation & Analysis: Label-Free BDD Immunosensors for Food AuthenticationâExecutive Summary
Section titled âExecutive SummaryâThis research successfully demonstrates a highly sensitive, label-free electrochemical immunosensor for the detection of porcine gelatin, leveraging the unique properties of Boron-Doped Diamond (BDD) electrodes.
- Core Achievement: Development of a rapid, highly specific, and label-free electrochemical platform for porcine gelatin detection, critical for food authentication and Halal verification.
- Material Platform: Utilizes a Boron-Doped Diamond (BDD) electrode, confirming BDDâs superior stability and wide potential window for complex biosensing applications.
- Surface Chemistry: Achieved stable, covalent immobilization of recognition elements (Protein A and anti-gelatin antibodies) via robust aryl diazonium salt electrografting (4-aminobenzoic acid).
- Analytical Performance: Demonstrated high sensitivity with a low detection limit (LoD) of 142.15 pg mL-1.
- Detection Method: Label-free detection relies on monitoring the current suppression of a redox probe ([Fe(CN)6]3-/4-) via Differential Pulse Voltammetry (DPV) upon specific antigen-antibody binding.
- Specificity: The sensor exhibited high selectivity, showing no significant cross-reactivity with closely related bovine gelatin, validating its reliability for discriminating animal sources.
- 6CCVD Value Proposition: 6CCVD specializes in providing custom, high-quality BDD materials necessary to replicate and scale this robust, functionalized biosensor platform.
Technical Specifications
Section titled âTechnical Specificationsâ| Parameter | Value | Unit | Context |
|---|---|---|---|
| Sensing Platform | Boron-Doped Diamond (BDD) | Electrode | Used for electrochemical immunosensing |
| Detection Limit (LoD) | 142.15 | pg mL-1 | Sensitivity for porcine gelatin |
| Optimal Antibody Incubation Time | 60 | min | Reduced from 8 hours for rapid detection |
| Optimal Gelatin Incubation Time | 15 | min | Rapid antigen binding time |
| DPV Potential Range | -0.3 to +0.7 | V | Used with [Fe(CN)6]3-/4- redox probe |
| DPV Scan Rate | 0.008 | V s-1 | Measurement parameter |
| Linear Correlation Coefficient (R) | 0.9854 | N/A | Confirms excellent linearity of calibration curve |
| BDD Bare Peak Current | 0.237 | mA | Baseline DPV response before modification |
| Final Sensor Peak Current (Target Bound) | 0.073 | mA | Current after porcine gelatin binding (signal suppression) |
| Specificity Test Result | < 7.3 | % Current Change | No significant cross-reactivity with bovine gelatin |
Key Methodologies
Section titled âKey MethodologiesâThe label-free immunosensor relies on a multi-step surface functionalization process using the BDD electrode as the base platform.
- Diazonium Salt Preparation: 4-aminobenzoic acid (4-ABA) was dissolved in 37% HCl, cooled to 0 °C, and mixed with NaNO2 to generate the diazonium salt precursor.
- BDD Electrografting: The diazonium salt was covalently electrografted onto the BDD surface using 11 consecutive cyclic voltammetry scans (0 V to +0.8 V, 100 mV s-1 scan rate) to form a stable, robust organic layer containing carboxyl groups.
- Protein A Immobilization: The carboxyl groups were activated using EDC/NHS coupling chemistry, facilitating the stable amide bond formation with Protein A (0.5 mg mL-1) to ensure oriented antibody attachment.
- Antibody Immobilization: Anti-porcine gelatin antibodies (0.1 g mL-1) were immobilized via interaction with Protein A, followed by an optimized incubation time of 60 minutes.
- Surface Blocking: Bovine Serum Albumin (BSA) solution (1%) was applied for 30 minutes to block any remaining unreacted sites, minimizing non-specific adsorption and enhancing sensor selectivity.
- Target Detection: Standard porcine gelatin solution (0.5 ng mL-1) was applied and incubated for 15 minutes.
- Electrochemical Readout: Differential Pulse Voltammetry (DPV) was used with 0.01 M K3[Fe(CN)6] in 0.1 M KCl as the redox probe to measure the current suppression resulting from the antibody-antigen binding event.
6CCVD Solutions & Capabilities
Section titled â6CCVD Solutions & CapabilitiesâThe successful development of this highly specific, label-free biosensor platform is fundamentally dependent on the quality and consistency of the Boron-Doped Diamond (BDD) electrode. 6CCVD is uniquely positioned to supply the advanced MPCVD diamond materials required to replicate, scale, and commercialize this technology for food authentication and Halal verification.
Applicable Materials
Section titled âApplicable MaterialsâTo replicate or extend this research, high-quality, electrochemically stable BDD is essential.
- Material Recommendation: Heavy Boron-Doped Diamond (BDD) Plates/Wafers.
- Justification: The BDD material must possess high conductivity and a wide potential window to support the DPV measurements and the robust electrografting process. 6CCVD provides BDD with precisely controlled doping levels, ensuring optimal electrochemical performance and low background current, which is critical for achieving the reported LoD of 142.15 pg mL-1.
- Alternative Material for High-Volume: Polycrystalline Diamond (PCD) Substrates.
- Justification: For large-scale manufacturing of biosensors, 6CCVD can supply PCD plates up to 125mm in diameter, which can be heavily boron-doped, offering a cost-effective and scalable alternative to smaller electrodes.
Customization Potential
Section titled âCustomization PotentialâThe stability and performance of the diazonium electrografting layer are highly dependent on the diamond surface quality.
- Polishing and Surface Preparation: 6CCVD offers ultra-smooth polishing (Ra < 5nm for inch-size PCD) to ensure uniform surface coverage during the diazonium electrografting step, leading to reproducible sensor performance and minimizing variability.
- Custom Dimensions: We supply BDD substrates in custom dimensions and thicknesses (up to 10mm substrates, 0.1”m - 500”m active layer thickness) tailored for specific device geometries, whether for laboratory research or integrated point-of-care devices.
- Metalization Services: While this paper focused on the BDD surface, future integration into microfluidic or array systems may require contact pads. 6CCVD offers in-house metalization capabilities (Au, Pt, Ti, W, Cu) for creating stable electrical contacts directly on the diamond substrate, streamlining device fabrication.
Engineering Support
Section titled âEngineering Supportâ6CCVD provides specialized technical consultation to accelerate biosensor development.
- Surface Chemistry Expertise: 6CCVDâs in-house PhD team can assist with material selection and surface preparation protocols for similar electrochemical immunosensor projects, ensuring the BDD surface is optimized for robust covalent modification techniques like diazonium electrografting.
- Application Focus: We support engineers and scientists developing platforms for Food Authentication, Halal Verification, and Clinical Diagnostics, leveraging our expertise in diamondâs unique electrochemical stability and biocompatibility.
- Global Supply Chain: We ensure reliable, global shipping (DDU default, DDP available) of custom diamond materials, supporting international research and commercialization efforts.
For custom specifications or material consultation, visit 6ccvd.com or contact our engineering team directly.
View Original Abstract
Porcine gelatin is widely used in the food and pharmaceutical industries due to its favorable functional properties and low cost. However, its presence in consumer products raises serious concerns for individuals with dietary restrictions based on religious, ethical, or health considerations. In this study, a label-free electrochemical immunosensor was developed using a boron-doped diamond electrode modified with aryl diazonium salt for the selective and sensitive detection of porcine gelatin. The diazonium electrografting enabled stable covalent immobilization of anti-porcine gelatin antibodies via protein A, preserving antiÂbody orientation and activity. Experimental parameters were optimized using the Box-Behnken design, yielding ideal conditions of 500Ă antibody concentration, 60 min antibody incubation, and 15 min gelatin incubation. Detection was performed using differential pulse voltammetry with [Fe(CN)â]3-/4- as a redox probe, allowing label-free monitoring of antiÂbody-antigen interactions based on changes in current. The immunosensor demonstrated excellent analytical performance, with a detection limit of 142.15 pg mL-1. Specificity testing confirmed that the sensor responds exclusively to porcine gelatin, showing no cross-reactivity with bovine gelatin. These results demonstrate that the proposed immunosensor provides a rapid, highly sensitive, and specific platform for porcine gelatin detection, offering great potential for food authentication and halal verification.