I. Core Research Summary

• In-situ High-Precision MXene Patterning via Hydrogels

Aerosol‑free deposition enables the fabrication of three types of micrometer‑scale conductive pathways—straight lines, bows, and Hilbert fractal curves—on flexible PEG hydrogels. The thickness of a single‑layer MXene film is tunable, with nanosheets uniformly stacked to form a continuous conductive network, while the mechanical properties of the hydrogel substrate remain intact despite the printing process.

• Excellent cellular biocompatibility

The proportion of viable cells maintained over a 7-day culture period continues to increase (from 85% on day 2 to 93% on day 7). MXene exhibits no cytotoxicity and does not induce apoptosis in cardiomyocytes, making it fully compatible with in vivo implantation for regenerative applications.

• Biomimetic定向 induction of orderly myocardial maturation

MXene‑induced conductive topological guidance aligns sarcomeres along the printed patterns, with a filament orientation dispersion of only 15.9° (compared to 34.9° in the glass‑substrate control group); expression of mature‑marker genes MYH7, TNNT2, and SERCA2 is significantly upregulated, and gap junction protein CX43 is abundantly enriched at cell–cell contact sites.

• Synchronous oscillations of calcium signals across the entire domain

The Hill‑curve‑connected conductive network enables uniform propagation of calcium waves across the entire myocardial patch, with a peak local conduction velocity of 6.5 cm/s and synchronous contractions over large areas, thereby addressing the issue of asynchronous cardiac contractions inherent in conventional hydrogel patches.

• Universal Aerosolization Process for High-Viscosity Bioinks

Compatible with a wide viscosity range of 1–1000 cP for nano‑bioinks, this single‑device system enables the fabrication of both conductive MXene materials and biohydrogels, while enabling room‑temperature deposition without thermal damage to bio‑based substrates.

II. Core Operating Principle of Aerosol Spraying and Its Unique Process Advantages

(1) Complete Hydrogel MXene Printing Process Flow

• A hydrogel precursor solution is prepared by combining PEG diacrylate with a photoinitiator, followed by molding in a PDMS mold and UV crosslinking to fabricate a flexible, transparent hydrogel substrate.
• The Ti₃C₂MAX phase is subjected to selective etching with HF/HCl and lithium intercalation–induced exfoliation to prepare a monolayer Ti₃C₂Tₓ MXene aqueous dispersion nanoink.
• Aerosol ultrasonic atomization of MXene ink, with nitrogen as the carrier gas, focused by a coaxial sheath‑gas jet, and deposited at a non‑contact, long‑range distance from the nozzle to the hydrogel.
• The CNC program enables the customization of micron-scale conductive patterns—straight lines, bows, and Hill curves—that self‑assemble and solidify through natural drying at room temperature.
• Basal sterilization and fibronectin surface modification were performed, followed by the seeding of iPSC‑derived cardiomyocytes for 7‑day long-term culture characterization.

(II) Five Core Process Advantages (Compared with Extrusion Printing, Lithography, and Molding)

• In-situ direct writing on the surface of flexible hydrogels, without damaging the soft matrix.

The extrusion‑based printing nozzle compresses and collapses the porous hydrogel; aerosol‑based non‑contact droplet deposition enables gentle deposition without mechanical stress, preserving the biomimetic mechanical properties of the PEG elastic substrate.

• High-precision topological shaping at the micrometer-scale cellular level

The minimum printable feature size can reach tens of micrometers, matching the dimensions of cardiomyocytes; continuous, interconnected fractal conductive pathways can be engineered to establish a whole‑tissue electrical conduction network, whereas conventional 3D extrusion typically achieves resolutions on the order of hundreds of micrometers.

• Compatible with multi-viscosity bioinks; a single system can fabricate both conductive and matrix materials.

Ultrasonic and dual‑atomization modes can be switched seamlessly, enabling stable atomization of both highly conductive MXene nanoliquids and polymeric hydrogels—without the need to replace the entire production line, thus ensuring broad applicability in biomanufacturing.

• Digital Maskless Rapid Topology Iteration

With CAD, you can instantly switch between straight lines, closed loops, and fractal conductive patterns—no custom silicon molds required—enabling rapid prototyping for cardiac microenvironment studies and significantly shortening the R&D cycle for biomaterials.

• Produced under mild conditions at room temperature throughout the process, with no risk of high-temperature inactivation.

The entire deposition and curing process is carried out at room temperature, ensuring that MXene, PEG, and bioactive proteins remain thermally stable and can be directly employed for cell culture and the fabrication of in vivo implantable materials.

III. Five Key Technological Innovations

Innovation 1: Innovatively realizing in-situ MXene conductive patterning via aerosol-jet printing of hydrogels.

Innovation 2: Bionic Design of a Hierarchical Fractal Continuous Conductive Network

Innovative 3‑topology + conductive dual‑myocardial induction mechanism

Innovative 4: Low-Damage, Ambient-Temperature Aerosol-Based Biomanufacturing System

Innovative 5‑Customizable Partitioned Conductive Myocardial Patch Manufacturing Platform

IV. Existing Process Challenges and Corresponding Solutions for Overcoming Them

Challenge 1: Blending conductive materials with hydrogels significantly alters the elastic modulus of the substrate, deviating from the mechanical match required for cardiac tissue.

Challenge 2: Extrusion printing pressure compresses the hydrogel, leading to substrate collapse and pore structure damage.

Challenge 3: Conventional photolithography requires a rigid substrate for curing, making it impossible to pattern on flexible hydrogels.

Challenge 4: At high concentrations, MXene blends tend to agglomerate, hindering cell adhesion and electrical conductivity.

Challenge 5: Discrete conductive electrode arrays are prone to electrical signal interruptions, leading to asynchronous myocardial contractions.

V. Future Multi-Industry Application Scenarios

• Post‑myocardial infarction implantation of a myocardial repair patch

• Ex vivo cardiac organ-on-a-chip / Drug screening model

•   In vitro model for the pathological study of cardiac arrhythmias

• Wearable Flexible Electrocardiogram-Sensing Hydrogel Electrodes

• Basic Stem Cell Cardiomyocyte Differentiation Research Platform

Figure 1. Preparation of Ti3C2Tx MXene on polyethylene glycol (PEG) hydrogels via three-dimensional conformal aerosol jet printing. (a) Schematic illustrating the selective HF etching of Ti3AlC2 MAX phase layers to obtain Ti3C2Tx MXene. (b) Schematic of aerosol jet printing for fabricating a Ti3C2Tx MXene–hydrogel composite, comprising: (I) preparation of a PEG hydrogel on a glass substrate; and (II) deposition of Ti3C2Tx MXene onto the hydrogel surface. (c) Square‑shaped (81 mm²) patterns of Ti3C2Tx MXene printed on the PEG hydrogel; inset shows a bow‑tie pattern of Ti3C2Tx MXene also printed on PEG. (d) SEM top‑view image of the 25‑nm‑thick Ti3C2Tx MXene layers after printing. (e) TEM image of Ti3C2Tx MXene. (f) TEM image of Ti3C2Tx MXene printed on the PEG hydrogel.

Figure 2. Distribution patterns and viability analysis of iCMs within the Ti3C2Tx MXene–PEG hydrogel composite: (a) linear; (b) cross-shaped; (c) Hilbert curve; (d) viability distribution of iCMs along the Hilbert curve at days 2, 4, and 7; (e) percentage of viable cells at the corresponding time points.

Figure 3. Immunostaining characteristics and qRT-PCR analysis results of iCMs on day 7 (all scale bars represent 20 µm). (a) CX‑43 staining of Ti3C2Tx MXene printed with linear patterns on a glass substrate, Ti3C2Tx MXene without any pattern as the control, and Ti3C2Tx MXene patterned with a Hilbert curve on a PEG substrate. (b) α‑actinin staining of sarcomeres in the glass‑substrate control, the Ti3C2Tx control, and the Hilbert‑curve sample; nuclei were stained with DAPI (blue). (c) Co‑staining for connexin‑43 and α‑actinin in the glass‑substrate control, the MXene control, and the Hilbert‑curve sample; nuclei were stained with DAPI (blue). (d) Quantitative analysis of sarcomere length in µm (*p < 0.05, **p < 0.01). (e) Directionality analysis. (f) qRT‑PCR analysis of relative mRNA expression levels of cardiac markers MYH7, SERCA2, GJA1, and TNNT2 (**p < 0.01, n.s. indicates no significant difference); (g) Western blot analysis of MYH7, SERCA2, GJA1, and TNNT2 protein expression; (h) Quantitative analysis of Western blot results (n = 2).

Figure 4. Shows the characteristics of spontaneous beating of iCMs cells on a glass substrate (a–d) and on a patterned Ti₃C₂Tx MXene–PEG composite hydrogel substrate (e–h). (a) Representative snapshots from calcium‑ion‑flux time‑series recordings; (b) Beat waveforms extracted from these time‑series records; (c) Intensity distributions of the calcium‑ion‑flux time‑series images; (d) Calculated times to peak, times for 50% decay of the peak, and times for 90% decay of the peak; (e) Representative snapshots from calcium‑ion‑flux time‑series recordings on the Ti₃C₂Tx MXene–PEG composite hydrogel (the pattern is divided into 16 regions, each represented by a different color); (f) Beat waveforms for each designated region, extracted from the time‑series records (waveform colors correspond to the subregion colors in panel e); (g) Average intensity distribution of the calcium‑ion‑flux time‑series images; (h) Calculated times to peak, times for 50% decay of the peak, and times for 90% decay of the peak.