I. Core Research Summary

Micrometer-scale, high-precision aerosol graphene interdigital electrode

Single‑channel printing fabricates 40‑μm‑wide graphene electrode fingers with a finger pitch of 100 μm; the resulting thin film is only 25 nm thick, and its electrical conductivity reaches 1×10⁴ to 7×10⁴ S/m—significantly thinner than conventionally screen‑printed or inkjet‑printed thick films, thereby substantially reducing material consumption. The device incorporates a complete set of 50 interdigitated electrode pairs, providing ample effective sensing area and delivering an excellent electrochemical signal‑to‑noise ratio.

CO₂ Annealing-Induced Modification to Construct a Bioactive Graphene Interface

After annealing, the surface of graphene exhibits a 10% increase in carboxyl and carbonyl oxygen-containing functional groups, enabling covalent immobilization of antibodies via EDC/NHS chemistry without the need for complex chemical modifications. The underlying graphene sp² framework remains largely defect-free, simultaneously offering high electrical conductivity and robust bioconjugation capabilities.

Dual-cytokine assay with ultra-wide linear range and ultra-low detection limit

Artificial bovine serum matrix validation:

• IFN-γ linear range: 0.1–5 ng/mL; limit of detection: 25 pg/mL.
• The linear range of IL-10 is 0.1–2 ng/mL, with a limit of detection of 46 pg/mL.
• At the same IL‑6 concentration, with cross‑cytokine interference, signal fluctuations are less than 20, demonstrating excellent specificity; no sample labeling or pre‑concentration is required throughout the entire process.

Excellent flexible mechanical stability

After 250 cycles of bending with a radius of just 5 mm, the electrode resistance variation remains below 1%, and the electrochemical impedance shift is only 6%. Even after immersion in buffer solution, the device maintains stable sensing performance, making it suitable for subcutaneous implantation and wearable monitoring applications.

Rapid Testing Procedure

Sample incubation takes only 30 minutes to achieve quantitative impedance measurements, a significant reduction compared with the several hours required by conventional immunoassays. The maskless aerosol‑based fabrication process enables rapid iteration of multi‑marker sensing chips.

II. Complete Preparation Process Flow and Core Process Advantages of Aerosol Spraying

(1) Complete Graphene IDE Aerosol Printing Process

• Graphite and nitrocellulose are shear‑exfoliated to prepare a composite powder, which is then formulated into a butyl lactate‑based stable aerosol ink.
• Ultrasonic atomization of graphene ink, focusing with a nitrogen sheath gas jet, and uniform-speed printing of interdigitated electrode arrays on a 60°C substrate.
• Air pyrolysis at 350°C is used to remove the binder, yielding a continuous, highly conductive graphene film.
• At 400°C, CO₂ tube‑type annealing induces the formation of numerous carboxyl functional groups on the graphene surface.
• After EDC/NHS activation, IFN-γ/IL-10–specific antibodies are immobilized to block non-specific adsorption sites, thereby completing the sensor chip.

(II) Five Core Processes (in comparison with lithography, screen printing, and inkjet printing)

• Sub-50 μm ultra-high-resolution maskless direct writing

Screen printing achieves a minimum line width of 100 μm, while inkjet typically offers 60 μm lines; AJP, meanwhile, stably produces 40 μm‑scale fine interdigitated electrodes, with extremely small electrode gaps that greatly enhance the electrochemical interface and significantly boost impedance‑sensing sensitivity—without the need for photolithographic masks or cleanroom facilities.

• Ultra-thin, low-power graphene film formation

A single-layer printed conductive film is only 25 nm thick; for the same sensing area, graphene-based materials require far less material than those used in other printing processes. The resulting thin film is continuous with minimal porosity, and its conductive network remains intact and stable.

• Compatible with wide-viscosity nano-ink; less prone to clogging.

Graphene‑based composite dispersions can maintain stable atomization over extended periods, addressing the issue of nozzle clogging in piezoelectric inkjet printers when using high‑solid carbon inks, and enabling highly consistent continuous batch printing.

• Gentle forming on flexible substrates, compatible with PI films.

The printing process employs a low-temperature substrate, eliminating high-temperature baking that could degrade the polyimide base material. The resulting devices exhibit exceptional flexibility, enabling substantial bending and making them well-suited for wearable applications and subcutaneous flexible sensing platforms.

• Digital rapid-iteration sensor deployment

CAD allows free modification of the number of interdigitated fingers, finger width, and array configuration, enabling rapid prototyping for various cytokines and multiplex assays, thereby reducing the R&D cycle from several days to just a few hours.

III. Five Key Technological Innovations

Innovation 1: The world’s first fully printed graphene interdigitated immunosensing platform for aerosols.

Innovative New Process for Surface Activation of Graphene via CO₂-Atmosphere Annealing

Innovative 3 Ultra-Thin, High-Sensitivity Interdigital Impedance Sensing Architecture

Innovation 4: Bendable, Body-Fluid-Compatible Flexible Immunochip

Innovation 5: Standardized Manufacturing Protocol for Universal Label-Free Immunoassays

IV. Existing Process Challenges and Corresponding Solutions for Overcoming Them

Challenge 1: Traditional printed lines are thick, resulting in insufficient electrochemical sensitivity of the interdigitated electrodes.

Challenge 2: Graphene films lack active functional groups, causing antibodies to desorb easily through physical adsorption.

Challenge 3: Carbon-based inks tend to clog the print head, making continuous batch printing impossible.

Challenge 4: Most carbon sensing membranes are thick and have high material costs.

Challenge 5: Rigid lithographic electrodes cannot be adapted for wearable or subcutaneous flexible monitoring.

V. Future Multi-Industry Application Scenarios Scenery

• Rapid On-Site Screening for Zoonotic Diseases

• Human autoimmune and tuberculosis home-monitoring patch

• Hospital Multiplex Cytokine Diagnostic Chip

• In vitro high-throughput screening of immunological efficacy for new drugs

• Implantable Subcutaneous Long-Acting Immune Monitoring Device

Figure 1. Schematic illustration of the fabrication and biofunctionalization of AJP‑fabricated graphene IDEs. (a) Schematic showing the formulation of the graphene ink used for aerosol printing. (b) The aerosol jet printing mechanism for graphene ink, illustrating how the sheath gas surrounding the aerosolized graphene‑ink particles focuses the ink to the desired diameter. (c) An AJP‑fabricated graphene IDE deposited on a polyimide (Kapton) substrate. (d) Using the N‑(3‑dimethylaminopropyl)‑N′‑ethylcarbodiimide/N‑hydroxysuccinimide (EDC/NHS) chemistry, selective antibodies targeting IL‑10 or IFN‑γ are immobilized onto the carboxyl‑functionalized graphene surface. (e) The remaining exposed surfaces of the graphene sensor are coated with a blocking agent (a mixture of bovine serum albumin, fish gelatin, and Tween‑20) and incubated together with the antigen.

Figure 2. Optical characterization of the AJP graphene IDE. (a) Optical micrograph showing line resolution; (b) SEM micrograph revealing the surface morphology of a graphene sheet on a polyimide substrate (magnification: 75,000×); (c) Top-view SEM micrograph of an ultrathin film (thickness < 100 nm) printed on silicon dioxide, acquired at a 45° tilt (magnification: 30,000×); (d) Two‑dimensional and three‑dimensional height images illustrating the surface irregularities and roughness of an untreated Kapton film; (e) AFM image of the finger-like structures of the graphene IDE; (f) AFM cross-sectional profile showing that the printed graphene has an average thickness of 25 nm (data extracted from the region within the box in panel e).

Figure 3. Spectral analysis of graphene IDEs. (a) XPS spectrum of the graphene IDE before thermal annealing in carbon dioxide (CO₂). (b) XPS spectrum of the graphene IDE after thermal annealing in carbon dioxide (CO₂). (c) Raman spectra of the printed graphene IDE before and after thermal annealing in carbon dioxide (CO₂), showing the D peak, G peak, and 2D peak.

Figure 4. (a) Analysis of multiple printed lines at a sheath flow rate of 55 sccm, a carrier gas flow rate of 35 sccm, and various printing speeds reveals that both line thickness and line width vary with printing time (s/mm), which is inversely proportional to the printing speed (mm/s). (b) The relationship between the electrical conductivity of graphene lines printed on a Kapton substrate and the maximum film thickness shows that conductivity increases with increasing film thickness, but begins to decline beyond approximately 200 nm. (c) A plot of resistance multiplied by cross-sectional area versus the dimensions of the printed graphene lines (resembling an IDE finger‑like structure) for different numbers of printing passes (labeled 1, 2, 3, 4, 5, 10, 15, 20, 25, and 30 in the legend). The slope of this curve represents the resistivity of the printed graphene. Electrochemical characterization of the graphene IDE includes: (d) cyclic voltammetry (CV) curves obtained at scan rates of 5, 10, 25, 50, and 100 mV/s, before and after CO₂ annealing; (e) Nyquist plots derived from electrochemical impedance spectroscopy (EIS) measurements taken before and after thermal annealing in CO₂; and (f) an enlarged view of the Nyquist plot for the annealed IDE shown in the lower left corner of panel (e), exhibiting the characteristic semicircular Nyquist curve.

Figure 5. (Top) Detection of IFN‑γ and IL‑10 using a graphene‑based IDE sensor. Percentage changes in the charge-transfer resistance (Rct) as a function of (a) IFN‑γ and (b) IL‑10 concentration. Error bars represent the standard deviation of the mean values obtained from three independent biofunctionalized electrodes, each coated with the corresponding bovine monoclonal antibody. (Bottom) (c) Selectivity testing of the IFN‑γ antibody toward IL‑10 and IL‑6 antigens, and (d) selectivity testing of the IL‑10 antibody toward IFN‑γ and IL‑6 antigens.

Figure 6. Bending‑test experiments conducted using the AJP graphene IDE. (a) Photograph of the testing apparatus, showing the IL‑10‑functionalized biosensor being bent around a suspended rod within a 3D‑printed channel filled with PBS solution. (b) Variation of the electrical resistance of the AJP graphene IDE with the number of bending cycles when no biological functionalization has been performed. (c) Relationship between the number of bends of the biosensor around a rod (diameter 5 mm) and the corresponding change in electrical resistance.