Embedded Files
Our team selected to modify an Opentrons OT-2 robot to be capable of NMR sample preparation
Opentrons is a company that develops affordable liquid-handling robots commonly used in small research laboratories.
OT-2 Pipettes
OT-2 Pipettes
Pipettes for the OT-2 come in single and multichannel, both function like standard manual pipettes
Our team developed a physical prototype by adapting a stainless steel pipette tip for the OT-2
Stainless steel needles are compatible with inorganic solvents and are thinner than NMR tubes, allowing solutions to be dispensed closer to the bottom.
Stainless steel needles are compatible with inorganic solvents and are thinner than NMR tubes, allowing solutions to be dispensed closer to the bottom.
The final product has a fully adapted OT-2 with a stainless steel pipette tip instead of the standard polypropylene.
The final product has a fully adapted OT-2 with a stainless steel pipette tip instead of the standard polypropylene.
User-input Code Script
OT-2 protocols are written in python, with functions that utilize polypropylene tips. Reworking the tips into stainless steel required the following code adaptions:
Adjusting Volume Calibrations
Improving Precision
Tweaking Pipette Height Values
Other Considerations
The NMR tubes on the sample rack occupy significant Z-axis space, which may prevent a standard stainless steel pipette from reaching over them. To address this, the prototype incorporates a sleeve with an arm to attach the needle that increases the height that the needle sits at.
Design Results
The FillBot Sleeve
The selection of a sleeve support system to hold all necessary components on the Opentrons pipette casing allows for the necessary support of the needle and tubing to successfully fill NMR tubes with the necessary level of precision through the addition of tubing and needle supports, preventing any shaking or movement of the needle that may cause a shift in positioning. This sleeve also allows for our extension mechanism to slide up and down with the movement of the Opentrons pipette.
Bottom Plate
Bottom Plate
The inclusion of a bottom plate on the FillBot sleeve allows for the downward movement of the system by the Opentrons pipette pushing on the column of the bottom plate, causing the sleeve to slide down at the same rate as the Opentrons pipette. This design also prevents any obstruction of the second pipette found in the Opentrons casing.
Tubing Plate
Tubing Plate
Separate from the FillBot Sleeve, our design features a tubing plate attached to the PVC tubing that connects to the tip of the Opentrons pipette tip, allowing for the upward movement of our sleeve by this plate applying pressure to the sleeve as the Opentrons moves upward, dragging the sleeve in the same direction. The addition of this part gives our sleeve a full range of motion when installed to the Opentrons machine.
Modified Pipette Tip
Modified Pipette Tip
To reduce the chance for collisions, we modified a standard 1000 uL pipette tip. It is bent to be horizontal so that it does not extend downwards towards lab wear on the bottom of the Opentrons. All suction from the pipette was maintained even with the modification. The extra clearance gained allows for safer and less complicated movement throughout the Opentrons lab bench as it does not need to be programmed to move around taller labware.
pipette movement.movHow it all fits together:
How it all fits together:
The sleeve slides on to the metal gantry that contains the pipettes and the needle is attached to the pipette with PVC tubing and the modified pipette tip. It is able to move up and down with the robot.
The FillBot Software
The FillBot Software
The FillBot software utilizes a CSV file input. This CSV file includes information such as sample ID, volume of reagents, locations of reagents, and total volume of solution to dispense into the NMR tubes. This CSV is inputted into the developed FillBot User Interface (UI) . A visual 96-well plate is outputted to allow the user to verify their samples have been inputted correctly. Within the FillBot UI, the user always is able to adjust the labware specification for the protein solution, giving the user control over whether eppendorf tubes, falcon tubes, reagent reservoirs, etc. are used. The user is then able to select whether to generate the OT-2 run script in one or two parts, allowing extra flexibility for labs with access to two OT-2s. A run script with the user-inputted variables is outputted, which can then be uploaded.
Run File Process Flow
Run File Process Flow
The OT-2 run file begins with sample preparation. In this stage, protein target, small molecules in DMSO, and aqueous buffers are mixed using in a 96-well plate using the standard OT-2 pipetting capabilities. Once complete, the run is paused and the FillBot sleeve is added. With the stainless steel needle attached, the system can now aspirate out of the 96-well plate and dispense in the bottom of the NMR tubes. In between each sample, the needle is rinsed with water and acid to ensure no sample overlap.
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