Hi everyone, I'm Ethan, the head of technical support at Opentrons. This video is the 7th in our series of videos from unboxing to protocol running. In the previous video, I took you through the tip length and pipette offset calibration. Next, I will explain how to upload and run the protocol. To do this I need to prepare all the consumables that I will use in the protocol run, I have tube racks, reservoirs and 96 well plates, I also need boxes of tips and pipettes, I have a P1000 single on the right Channel pipette, with a P20 on the left 8-channel pipette, and finally, before running the protocol, I'm going to put the wastebasket in place, the wastebasket will protect the metal plate from any liquid that falls in the tip, in order to run on the 0T-2 protocol, I needed to hook up my pipette to the OT-2 and complete the deck, tip length, and offset calibration of the pipette. Before we start, I need a protocol. I have created a protocol for this video using the Python API. You can find it in our online protocol library. The link to the protocol can be found in the video description. This is a very simple protocol. The running content is to draw liquid from the reservoir and distribute it into the well plate. Let's go into the application and import a protocol. I will go into Opentrons application, navigate to the "Protocols" tab, this step takes us to the protocol import page, I can import the protocol by dragging and dropping the file into the application or selecting file upload, I choose to select the file upload option and a file preview window pops up, I can select a protocol that I've downloaded, and once I import the protocol, the application will run it through protocol analysis, which is a process of locally simulating the protocol in order to obtain specific information related to the metadata and experiment deck layout. information.
Once the protocol is imported I can start the run, to do this I need to be in the protocol tab and then I can select the 3 dot menu and when clicked it shows a drop down menu from which to select the run option so there is only one in this list , if I have multiple 0T-2, I select this device and click "Proceed to Setup", and then enter the protocol setting page, which provides information about the protocol I imported. If you've followed me in the previous videos, you know that we completed the calibration of the equipment required to run it. So there is no need to redo the first step. If the calibration is not completed, there will be a yellow warning that calibration is required. Let me briefly introduce step 2, labware setup. Opentrons recommends this labware setup as part of the protocol setup. , this step primarily involves using labware positions to check the application or creating labware offsets, which are small adjustments that are used to position the pipette in the correct position for a specific labware and a specific experiment plate .
Since I have not performed an alignment check on any labware before, I can move on to step 2, which just gives me the opportunity to show you what a labware alignment check is and how to perform it. Let's move on to the labware positioning check. As soon as I click to start the labware positioning check, OT-2 will move the pipette to the A1 tip in plate position 6. I need to position all the nozzles with the first column. Alignment, when the 0T2 is moved to the tip rack, I need to make sure all the nozzles are in the correct position and aligned. If the channels aren't perfectly aligned, I can click on the trim controls. I'm going to start adjusting now because the pipetting channel is slightly off. Now that the fine-tuning controls will show up, I'm going to quickly adjust my pipette to align it.
Now I want to confirm the position, the pipette will move to plate position 5 and the other pipette will come down to tip A1. Now I need to make sure that the nozzle of the P1000 pipette is centered on tip A1, with the nozzle somewhat offset from the top of the tip. So I'm going to go to the fine-tune control just like before, move it a little bit, and once it's aligned, I'm going to click confirm positioning, pick up the tip OT-2 will try to pick up the tip and the app will ask me if the pipette was successful Picked up the tip, it had been picked up, so I clicked OK. It will move to plate position 1, the tip should be in the center of well A1, and the bottom of the tip should be flush with the top of the well. I'm going to fine-tune it because it's a little bit off. So, again I need to fine-tune the tip positioning to the center of the A1 hole, with the bottom of the tip flush with the top of the hole slot. I want to confirm the position, and the pipette will move to plate position 2, it will move to the first column of the reservoir, the pipette will move to the A1 column of the reservoir 2, and now I need to move the tip Align the slots in column A1 and make sure the bottom of the tip is flush with the top of the slot. It was a little on the low side, so I had to fine-tune the control and adjust it a little so that the tip was centered in the well A1 slot and the bottom of the tip was flush with the top of the slot.
So I will confirm the position and move to plate position 3, just like the previous two steps, it will move to the A1 well, now I need to make sure that the tip is aligned with the A1 hole, and the bottom of the tip is aligned with the top of the well flat. It's pretty good, but I think it could be better, so I'm going to fine-tune the controls and tweak it a little bit. Now this calibration is done very well. At this point I click to confirm the positioning and the pipette will place the tip back on the tip rack in position 5. It will lower the needle and the OT-2 will return to its original position. OT-2 is reset, indicating that the experimental deck positioning check is completed. Because my deck map all has green checkmarks. Now the offset for each labware I run is shown here as well. So I'm going to close the screen and apply the labware offset data, and on this DEC map you can see the labware offset data that I just calibrated, and now I go back to the protocol settings tab and all the calibration is done. I can start running and we can click start. Run to start! This video ends here, let OT-2 continue to execute the protocol calmly. Thank you so much for watching the final video in this series. This video summarizes all the steps required to get the OT-2 to perform the protocol, thank you for watching, and see you soon!
