Neuronal growth on high-aspect-ratio diamond nanopillar arrays for biosensing applications
At a Glance
Section titled âAt a Glanceâ| Metadata | Details |
|---|---|
| Publication Date | 2023-04-11 |
| Journal | Scientific Reports |
| Authors | Elena Losero, Somanath Jagannath, Maurizio Pezzoli, Valentin Goblot, Hossein Babashah |
| Institutions | Ăcole Polytechnique FĂ©dĂ©rale de Lausanne, The University of Sydney |
| Citations | 29 |
| Analysis | Full AI Review Included |
Executive Summary
Section titled âExecutive SummaryâThis research successfully validates a nanostructured single-crystal diamond platform for high-resolution, label-free quantum biosensing of neuronal activity, addressing the challenge of low signal sensitivity in mammalian neurons.
- Platform Development: Large-scale (up to 2 mm x 2 mm) arrays of high-aspect-ratio (HAR) nanopillars (d: 100-500 nm; h: ~1 ”m; AR > 10:1) were reliably fabricated on single-crystal CVD diamond.
- Quantum Sensing Enhancement: The nanopillars demonstrated a waveguiding effect, increasing Photoluminescence (PL) collection efficiency by 15% compared to flat surfaces, crucial for boosting the sensitivity of Nitrogen-Vacancy (NV) centers.
- Biocompatibility Confirmed: Primary mouse hippocampal neurons were successfully grown on the nanostructured arrays, maintaining full viability and functional electrophysiological properties (e.g., typical resting potentials and action potential firing).
- Optimal Interface Achieved: Scanning Electron Microscopy (SEM) confirmed close physical contact between neurites and the pillar apices, validating the geometry required for near-surface NV centers to detect local electric fields without ionic screening.
- Neuronal Guidance: The nanopillar grid induced preferential neuronal growth along the array axes, demonstrating a method to tailor network architecture for targeted sensing applications.
- Functional Milestone: This work establishes a critical milestone toward realizing an NV-based quantum sensing platform capable of wide-field, sub-cellular resolution recording of living neuronal networks.
Technical Specifications
Section titled âTechnical Specificationsâ| Parameter | Value | Unit | Context |
|---|---|---|---|
| Substrate Material | Single-crystal CVD Diamond (100) | - | Optical grade, Element6 |
| Substrate Dimensions | 3 x 3 x 0.25 | mm | Chip size |
| Array Size (Max) | 2 x 2 | mm2 | Uniform coverage area |
| Nanopillar Diameter (d) | 100 to 500 | nm | Range tested |
| Nanopillar Height (h) | ~1 (up to 2) | ”m | Target height |
| Aspect Ratio (h:d) | > 10:1 | - | Achieved verticality |
| Nanopillar Pitch (p) | 1 to 10 | ”m | Distance between pillars |
| NV Center Concentration | 1.4 | ppb | Uniform density in optical grade diamond |
| Surface Roughness (RMS) | < 2 | nm | After non-contact polishing |
| Diamond Etch Rate (O2 Plasma) | 100 | nm/min | Highly directional O2-plasma etch |
| PL Enhancement | +15 | % | Compared to flat surface (d=500 nm, p=1 ”m) |
| Neuronal Resting Potential | -63 to -49 | mV | Measured in functional hippocampal neurons |
| Action Potential Threshold | ~30 | pA | Current injection required to elicit firing |
| EPSP Time Decay Constant (Ï) | ~25 | ms | Excitatory postsynaptic potential fit |
| EBL Exposure Time | Less than 20 | minutes | For a 2 mm x 2 mm array |
Key Methodologies
Section titled âKey MethodologiesâThe fabrication of the high-aspect-ratio diamond nanopillar arrays followed a multi-step process utilizing electron beam lithography (EBL) and highly directional dry etching.
- Substrate Preparation:
- Commercial (100) CVD single-crystal diamond was cleaned (acetone, piranha solution: 3:1 H2SO4:H2O2).
- A preliminary non-contact polishing step (physical bombardment with accelerated inert gas ions) was applied to remove mechanical defects and achieve a root mean square (rms) roughness of less than 2 nm.
- Hard Mask Deposition:
- A 200 nm Titanium (Ti) layer was sputtered onto the diamond surface to serve as the hard mask.
- Resist Patterning (EBL):
- A ~150 nm layer of Hydrogen Silsesquioxane (HSQ XR-1541-006) negative resist was spin-coated.
- Pillars were patterned using Electron Beam Lithography (EBL), followed by development in Tetramethylammonium hydroxide (TMAH 25%).
- Mask Transfer (Ti Etch):
- The HSQ pattern was transferred to the Ti hard mask using a Cl2-based Reactive Ion Etching (RIE) process (STS Multiplex ICP: 800 W ICP power, 150 W bias power, 10 sccm Cl2, 10 sccm BCl3, 3 mTorr).
- Diamond Etch (HAR):
- A highly directional O2-plasma etch was used to create the nanopillars (STS Multiplex ICP: 400 W ICP power, 200 W bias power, 30 sccm O2, 15 mTorr), achieving vertical sidewalls and high aspect ratios.
- Mask Removal:
- The remaining HSQ and Ti masks were stripped using diluted Hydrofluoric Acid (HF, 1% concentration).
- Cell Culture Preparation:
- Sterile diamond chips were coated sequentially with Poly-L-lysine (1 hour) and Laminin (30 minutes at 37 °C) to facilitate neuronal adhesion and growth.
- Neuronal Plating:
- Primary mouse hippocampal neurons (P0/P1 pups) were plated at a density of 150,000 cells/ml and maintained for 10-14 days in vitro (DIV) before imaging or electrophysiology.
Commercial Applications
Section titled âCommercial ApplicationsâThe developed nanostructured diamond platform is highly relevant for advanced biosensing and quantum technology sectors:
- Quantum Biosensing Platforms: Realization of nanophotonic quantum sensing devices utilizing near-surface NV centers for wide-field, label-free recording of biological signals.
- Neuroscience Research Tools: Development of high-resolution neural interfaces capable of sub-cellular spatial and sub-millisecond temporal resolution for analyzing neuronal network dynamics and information processing.
- Biophysics Modeling: Providing high-sensitivity measurements of electric and magnetic fields generated by neuronal activity, enabling better modeling of neuronal biophysics.
- Neurodegenerative Diagnostics: Potential application in identifying early stages of brain disorders (e.g., Parkinsonâs and Alzheimerâs) by sensitively measuring subtle changes in neuronal electric/magnetic fields.
- Diamond Photonics: The robust, scalable fabrication process for HAR diamond structures is applicable to creating other integrated photonic devices, such as waveguides and single-photon sources, particularly when using electronic grade diamond substrates with shallow NV implantation.
- Biocompatible Interfaces: Use of nanostructured diamond as a highly biocompatible neural interface for electrodes and microelectrode arrays.