LabPilot

Overview

In previous experiments, Lambert iGEM experienced firsthand difficulties of tedious pipetting and inaccurate results. Last year, Rolling Circle Amplification (RCA) reactions we ran took the majority of our time. This took important research time away, limiting us from our full potential. Issues like these inspired us to develop LabPilot, a frugal automated liquid handler that allows researchers to focus time elsewhere from running lengthy reactions like RCA. Pipetting is a fundamental laboratory technique used to transfer precise volumes of liquid from one container to another. Consistent pipetting is essential for achieving accurate results, maintaining data integrity, and facilitating advancements in synthetic biology research. However, manual pipetting, the primary method utilized in most labs, limits throughput (the amount of liquid passing through a pipette) and is easily susceptible to human error. To resolve these issues, automated liquid handlers are designed to automate and streamline the process of pipetting liquids in laboratory settings. Liquid handlers can be programmed to perform a wide range of pipetting tasks with high precision, accuracy, and efficiency, enabling high throughput experiments (Liquid Handling Automation Benefits). However, current automated liquid handlers costing between $5000 and $300,000 are difficult to use (Retisoft, 2021). To provide underfunded labs access to liquid handlers, Lambert iGEM developed LabPilot, a frugal liquid handler made with 3D printable and accessible parts that can accurately pipette based on direct user input (Hentz & Knaide, 2014). Therefore, it enables research to be conducted in an efficient manner, filling an essential gap in lab settings.

Parts list

Design

LabPilot is an automated liquid handler compatible with micropipettes. It utilizes an XYZ layout for movement, with most of its structural parts designed on Fusion 360, a computer-aided design (CAD) application. LabPilot’s design can be split into the frame, detachable molds for the bed, and XYZ planes with motors.

Frame

Our team designed Labpilot’s frame to be small enough to fit inside a flow hood and large enough to fit standard laboratory containers on its bed (e.g. microcentrifuge tube racks, micropipette tip boxes, and well plates). We designed the frame by creating a linear rail extrusion, as shown in Figure 1, typically found on commercial 3D printers. We used linear extrusions to make the cage-like design of LabPilot, inspired by Ender 3D printers like the Ender 5 (see Table 1) (Ender 5 S1 3D Printer). The finalized XYZ dimensions of LabPilot are 366mm x 296mm x 356mm (see Fig. 2). To provide stability to LabPilot’s frame, we added angle brackets that connect two of the extrusions that run perpendicular to each other at all 16 corners of the frame. We spoke to Priya Soneji, an undergraduate researcher at the Georgia Institute of Technology, who is a part of the Bhamla Lab. For the future, she recommended the switch to metal frames if the 3D-printed parts were imprecise. She confirmed the validity of our overall plan for controlling each motor axis on the XYZ plane and methods for motor movement.

Figure 1. CAD model of linear rail extrusion used to make the structural frame.

Figure 2. CAD model of LabPilot frame.

Molds

Lambert iGEM created molds that snap onto the bed plate of LabPilot so that the user can attach their desired components to the bed. The molds have pegs (see Fig. 3) that can be placed inside engraved indentations on the bed plate (see Fig. 4), which fixes the molds in place and enables the LabPilot app to calculate their position. LabPilot has a mold that fits every micropipette tip box, beaker, microcentrifuge tube rack, PCR tube rack, and well plate.

Figure 3. Pegs placed on the bottom of the molds to fit into the bed plate.

Figure 4. Indentations on the bed plate for the molds to rest on.

XYZ Motor Axis

LabPilot utilizes five Nema 17 motors (Nema 17 Stepper Motor) (see Table 1) to control the three axes of movement needed for the pipetting mechanism. Nema 17 motors have a micro-stepping feature, enabling small and precise movements. Each motor is controlled by an Arduino Mega 2560 with a Ramps 1.4 CNC shield containing five A4988 motor drivers (see Table 1). The CNC shield uses two motors to control the vertical movement of the bed plate, as it is the heaviest moving part. The left/right movement and the forward/backward movement of the micropipette are controlled by one motor each. The last motor controls the pipetting mechanism (see Fig. 5) and is placed above the micropipette. Additionally, we placed limit switches for each axis to prevent over-rotation of the motors and calibration to home coordinates similar to 3D printers. Each axis has linear steel rods that act as supports, with bearings in place to ensure the pipette mechanism slides smoothly across the rods. We attached timing belts (thin rubber belts) with teeth (see Table 1) to the motor to carry the pipette left, right, forwards, and backwards across the X and Y axes. In addition to the linearly moving axes, LabPilot requires a motor to press the micropipette to the first and second stops. The pipetting mechanism motor converts the rotational motion of the motor into a linear force that can push down on the micropipette button. The motor applies a fine-tuned force to achieve first and second stops. Currently, LabPilot is designed to be used with Eppendorf micropipettes to ensure everything fits correctly. We attached a stationary arm to the bed plate to eject the pipette tips. The micropipette moves into the arm while sequentially traveling upwards. This way, the tip is pulled off the micropipette and discarded underneath.

Figure 5. Pipette mechanism

App

To make LabPilot easier for researchers, Lambert iGEM designed a web app to control the liquid handler. Unlike the complex software that operates commercial liquid handlers such as Opentrons, the LabPilot app has a simple, human-centric user interface designed to be used immediately with simple and intuitive commands. This design enables users to quickly start using LabPilot and efficiently simplify their tasks without any difficulties.

How it Works

To intelligently control its actions, LabPilot utilizes an Arduino microcontroller connected to a laptop running the LabPilot web app via a USB cable. The microcontroller is in charge of controlling the movement of the pipette mechanism and the action of pipetting. The microcontroller moves the micropipette to a specific location by using a virtual coordinate system, which defines different areas of the surface with numbers. The LabPilot app knows what actions to take in order to move the desired axes to their coordinate destination.

Figure 6. shows how the user input on the app controls the microcontroller.

The app provides instructions to the microcontroller via the Arduino serial USB connection (see Fig. 6). By utilizing a system where the app executes high-level operations by compiling pipette actions into low-level instructions for the microcontroller, users have complete control over their LabPilot device. They can stop or pause it at any time.

Figure 7. Shows how the user input controls LabPilot.

How to use

To use LabPilot, users must connect a laptop to LabPilot via a USB cable and open its web dashboard. Once the dashboard is open, there is a simple setup process where users can drag and drop components from the sidebar to recreate the physical layout of their LabPilot device, as seen in Figure 8. After setup, the app digitally recreates the layout of this device from data gathered in the setup process, enabling adequate simulation and control. Then, the users can queue and perform pipetting actions.

Figure 8. LabPilot’s setup interface.

All pipetting actions consist of a reagent source and a dispensing source. Users can click on any well or beaker to select a reagent source. After choosing a reagent source, users can select a dispensing source by clicking on any well/beaker or multiple dispensing sources (by clicking and dragging or holding “shift” while they select multiple sources). Afterward, users will select the amount they want to dispense to each dispensing source and can add it to the queue (see Fig. 7). All pipetting actions in the queue are performed in the order they were assigned but can be dragged around to reorder in the queue.

Conclusion and Future Directions

Researchers can leverage LabPilot’s capabilities to streamline other critical processes such as DNA sequencing, sample preparation, high-throughput screening, and assay development. By automating these tasks, LabPilot enables scientists to focus on higher-level analysis and interpretation of results, accelerating the pace of scientific discoveries. Next year, Lambert iGEM hopes to switch to an aluminum frame for its cost-effective, rigid nature and increase compatibility with various laboratory equipment. Additionally, we plan to test with polymerase chain reaction (PCR) as a proof of concept experiment. A successful PCR will have LabPilot demonstrate accuracy in pipetting volumes, with substantial precision and a significant reduction in PCR time. Once we can prove that LabPilot upholds the accuracy and efficiency of a human through a proof of concept, we plan on using LabPilot to automate a portion of our rolling circle amplification biosensors to improve efficiency and reduce the potential for human error. Our end goal is to develop a kit available for purchase and make LabPilot commercially available. LabPilot’s cost-effectiveness, user-friendly interface, and open-source nature make it accessible and adaptable to all laboratory settings.

References