3D bioprinting of human neural tissues with functional connectivity

This is he G code related to the publication Yan et al. 2024 3D bioprinting of human neural tissues with functional connectivity. The full publication can be found here: https://doi.org/10.1016/j.stem.2023.12.009

## Overview

This repository provides machine-control G-code routines for automated, dual-toolhead micro-extrusion dispensing across sequential layers. The protocol coordinates alternating toolheads (`T1` and `T2`), configuring relative volumetric dosing, automated tool transitions, and precise clearance trajectories.

This codebase accompanies the publication:
> Yan, Y., et al. (2024). *Cell Stem Cell*. https://doi.org/10.1016/j.stem.2023.12.009

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## Technical Specifications

| Parameter | Setting | Description |
| :--- | :--- | :--- |
| **Units** | `G21` | Millimeters |
| **Positioning** | `G90` | Absolute coordinate positioning |
| **Extrusion Mode** | `M83` | Relative extrusion distance |
| **Dispensing Height** | `Z = 0.1 mm` | Layer height plane |
| **Layer Span** | `X = -3.0 mm` to `3.0 mm` | 6.0 mm linear trace length |
| **Extrusion Volume** | `E = 0.1` | Relative extrusion per pass |

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## Execution Sequence

The G-code routine executes an alternating layer pattern between Toolhead 1 (`T1`) and Toolhead 2 (`T2`):

```
Pass 1 (T1) -> Layer 1 at Y =  1.0 mm
Pass 2 (T2) -> Layer 2 at Y =  0.5 mm
Pass 3 (T1) -> Layer 3 at Y =  0.0 mm
Pass 4 (T2) -> Layer 4 at Y = -0.5 mm
```

### Motion Profile Details

* **Layer 1 (`T1`):** Moves to `(X: -3.0, Y: 1.0, Z: 0.1)`, extrudes along X to `3.0` (`E0.1`), retracts `Z` to 10 mm.
* **Layer 2 (`T2`):** Switches to `T2`, positions at `(X: -3.0, Y: 0.5, Z: 0.1)`, extrudes to `X: 3.0` (`E0.1`), retracts `Z` to 10 mm.
* **Layer 3 (`T1`):** Switches to `T1`, positions at `(X: -3.0, Y: 0.0, Z: 0.1)`, extrudes to `X: 3.0` (`E0.1`), parks `Z` to 50 mm clearance.
* **Layer 4 (`T2`):** Switches to `T2`, positions at `(X: -3.0, Y: -0.5, Z: 0.1)`, extrudes to `X: 3.0` (`E0.1`), parks `Z` to 50 mm clearance, and disables motors (`M84`).

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## Repository Structure

```
├── protocols/
│   └── multi_extrusion.gcode    # Primary execution G-code script
├── hardware/
│   └── toolhead_config.json     # Spatial offsets and calibration settings
├── LICENSE                      # MIT license file
└── README.md                    # Project documentation
```

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## Requirements & Safety

1. **Hardware/Firmware Compatibility:** Marlin, RepRapFirmware, or custom bioprinting/CNC controllers supporting standard G-code and multi-tool definitions (`T0`, `T1`, `T2`).
2. **Clearance Verification:** Ensure vertical travel clearance of at least 50 mm prior to execution.
3. **Toolhead Calibration:** Confirm nozzle offsets and bed leveling are calibrated prior to executing dispense commands.

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## Contributors & Contact

* **Yuanwei Yan** – First Author
* **Su-Chun Zhang** (Principal Investigator) – `suchunz@sbpdiscovery.org`

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## License

This project is licensed under the [MIT License](LICENSE).