Protocol Examples

This page provides simple, ready-made protocols for Flex and OT-2. Feel free to copy and modify these examples to create unique protocols that help automate your laboratory workflows. Also, experimenting with these protocols is another way to build upon the skills you’ve learned from working through the tutorial. Try adding different hardware, labware, and commands to a sample protocol and test its validity after importing it into the Opentrons App.

Using These Protocols

These sample protocols are designed for anyone using an Opentrons Flex or OT-2 liquid handling robot. For our users with little to no Python experience, we’ve taken some liberties with the syntax and structure of the code to make it easier to understand. For example, we’ve formatted the samples with line breaks to show method arguments clearly and to avoid horizontal scrolling. Additionally, the methods use named arguments instead of positional arguments. For example:

# This code uses named arguments
tiprack_1 = protocol.load_labware(
    load_name="opentrons_flex_96_tiprack_200ul",
    location="D2")

# This code uses positional arguments
tiprack_1 = protocol.load_labware("opentrons_flex_96_tiprack_200ul", "D2")

Both examples instantiate the variable tiprack_1 with a Flex tip rack, but the former is more explicit. It shows the parameter name and its value together (e.g. location="D2"), which may be helpful when you’re unsure about what’s going on in a protocol code sample.

Python developers with more experience should feel free to ignore the code styling used here and work with these examples as you like.

Instruments and Labware

The sample protocols all use the following pipettes:

• Flex 1-Channel Pipette (5–1000 µL). The API load name for this pipette is flex_1channel_1000.

• P300 Single-Channel GEN2 pipette for the OT-2. The API load name for this pipette is p300_single_gen2.

They also use the labware listed below:

Labware type	Labware name	API load name
Reservoir	USA Scientific 12-Well Reservoir 22 mL	usascientific_12_reservoir_22ml
Well plate	Corning 96-Well Plate 360 µL Flat	corning_96_wellplate_360ul_flat
Flex tip rack	Opentrons Flex 96 Tip Rack 200 µL	opentrons_flex_96_tiprack_200ul
OT-2 tip rack	Opentrons 96 Tip Rack 300 µL	opentrons_96_tiprack_300ul

Protocol Template

This code only loads the instruments and labware listed above, and performs no other actions. Many code snippets from elsewhere in the documentation will run without modification when added at the bottom of this template. You can also use it to start writing and testing your own code.

FlexOT-2

from opentrons import protocol_api

requirements = {"robotType": "Flex", "apiLevel": "2.17"}

def run(protocol: protocol_api.ProtocolContext):
    # load tip rack in deck slot D3
    tiprack = protocol.load_labware(
        load_name="opentrons_flex_96_tiprack_1000ul", location="D3"
    )
    # attach pipette to left mount
    pipette = protocol.load_instrument(
        instrument_name="flex_1channel_1000",
        mount="left",
        tip_racks=[tiprack]
    )
    # load well plate in deck slot D2
    plate = protocol.load_labware(
        load_name="corning_96_wellplate_360ul_flat", location="D2"
    )
    # load reservoir in deck slot D1
    reservoir = protocol.load_labware(
        load_name="usascientific_12_reservoir_22ml", location="D1"
    )
    # load trash bin in deck slot A3
    trash = protocol.load_trash_bin(location="A3")
    # Put protocol commands here

Transferring Liquids

These protocols demonstrate how to move 100 µL of liquid from one well to another.

Basic Method

This protocol uses some building block commands to tell the robot, explicitly, where to go to aspirate and dispense liquid. These commands include the pick_up_tip(), aspirate(), and dispense() methods.

FlexOT-2

from opentrons import protocol_api

requirements = {"robotType": "Flex", "apiLevel":"2.17"}

def run(protocol: protocol_api.ProtocolContext):
    plate = protocol.load_labware(
        load_name="corning_96_wellplate_360ul_flat",
        location="D1")
    tiprack_1 = protocol.load_labware(
        load_name="opentrons_flex_96_tiprack_200ul",
        location="D2")
    trash = protocol.load_trash_bin("A3")
    pipette = protocol.load_instrument(
        instrument_name="flex_1channel_1000",
        mount="left",
    tip_racks=[tiprack_1])

    pipette.pick_up_tip()
    pipette.aspirate(100, plate["A1"])
    pipette.dispense(100, plate["B1"])
    pipette.drop_tip()

Advanced Method

This protocol accomplishes the same thing as the previous example, but does it a little more efficiently. Notice how it uses the InstrumentContext.transfer() method to move liquid between well plates. The source and destination well arguments (e.g., plate["A1"], plate["B1"]) are part of transfer() method parameters. You don’t need separate calls to aspirate or dispense here.

FlexOT-2

from opentrons import protocol_api

requirements = {"robotType": "Flex", "apiLevel": "2.17"}

def run(protocol: protocol_api.ProtocolContext):
    plate = protocol.load_labware(
        load_name="corning_96_wellplate_360ul_flat",
        location="D1")
    tiprack_1 = protocol.load_labware(
        load_name="opentrons_flex_96_tiprack_200ul",
        location="D2")
    trash = protocol.load_trash_bin("A3")
    pipette = protocol.load_instrument(
        instrument_name="flex_1channel_1000",
        mount="left",
        tip_racks=[tiprack_1])
    # transfer 100 µL from well A1 to well B1
    pipette.transfer(100, plate["A1"], plate["B1"])

Loops

In Python, a loop is an instruction that keeps repeating an action until a specific condition is met.

When used in a protocol, loops automate repetitive steps such as aspirating and dispensing liquids from a reservoir to a a range of wells, or all the wells, in a well plate. For example, this code sample loops through the numbers 0 to 7, and uses the loop’s current value to transfer liquid from all the wells in a reservoir to all the wells in a 96-well plate.

FlexOT-2

from opentrons import protocol_api

requirements = {"robotType": "Flex", "apiLevel":"2.17"}

def run(protocol: protocol_api.ProtocolContext):
    plate = protocol.load_labware(
        load_name="corning_96_wellplate_360ul_flat",
        location="D1")
    tiprack_1 = protocol.load_labware(
        load_name="opentrons_flex_96_tiprack_200ul",
        location="D2")
    reservoir = protocol.load_labware(
        load_name="usascientific_12_reservoir_22ml",
        location="D3")
    trash = protocol.load_trash_bin("A3")
    pipette = protocol.load_instrument(
        instrument_name="flex_1channel_1000",
        mount="left",
        tip_racks=[tiprack_1])

    # distribute 20 µL from reservoir:A1 -> plate:row:1
    # distribute 20 µL from reservoir:A2 -> plate:row:2
    # etc...
    # range() starts at 0 and stops before 8, creating a range of 0-7
    for i in range(8):
        pipette.distribute(200, reservoir.wells()[i], plate.rows()[i])

Notice here how Python’s range class (e.g., range(8)) determines how many times the code loops. Also, in Python, a range of numbers is exclusive of the end value and counting starts at 0, not 1. For the Corning 96-well plate used here, this means well A1=0, B1=1, C1=2, and so on to the last well in the row, which is H1=7.

Multiple Air Gaps

Opentrons electronic pipettes can do some things that a human cannot do with a pipette, like accurately alternate between liquid and air aspirations that create gaps within the same tip. The protocol shown below shows you how to aspirate from the first five wells in the reservoir and create an air gap between each sample.

FlexOT-2

from opentrons import protocol_api

requirements = {"robotType": "Flex", "apiLevel":"2.17"}

def run(protocol: protocol_api.ProtocolContext):
    plate = protocol.load_labware(
        load_name="corning_96_wellplate_360ul_flat",
        location="D1")
    tiprack_1 = protocol.load_labware(
        load_name="opentrons_flex_96_tiprack_1000ul",
        location="D2")
    reservoir = protocol.load_labware(
        load_name="usascientific_12_reservoir_22ml",
        location="D3")
    trash = protocol.load_trash_bin("A3")
    pipette = protocol.load_instrument(
        instrument_name="flex_1channel_1000",
        mount="left",
        tip_racks=[tiprack_1])

    pipette.pick_up_tip()

    # aspirate from the first 5 wells
    for well in reservoir.wells()[:5]:
        pipette.aspirate(volume=35, location=well)
        pipette.air_gap(10)

    pipette.dispense(225, plate["A1"])

    pipette.return_tip()

Notice here how Python’s slice functionality (in the code sample as [:4]) lets us select the first five wells of the well plate only. Also, in Python, a range of numbers is exclusive of the end value and counting starts at 0, not 1. For the Corning 96-well plate used here, this means well A1=0, B1=1, C1=2, and so on to the last well used, which is E1=4. See also, the Commands section of the Tutorial.

Dilution

This protocol dispenses diluent to all wells of a Corning 96-well plate. Next, it dilutes 8 samples from the reservoir across all 8 columns of the plate.

FlexOT-2

from opentrons import protocol_api

requirements = {"robotType": "Flex", "apiLevel": "2.17"}

def run(protocol: protocol_api.ProtocolContext):
    plate = protocol.load_labware(
        load_name="corning_96_wellplate_360ul_flat",
        location="D1")
    tiprack_1 = protocol.load_labware(
        load_name="opentrons_flex_96_tiprack_200ul",
        location="D2")
    tiprack_2 = protocol.load_labware(
        load_name="opentrons_flex_96_tiprack_200ul",
        location="D3")
    reservoir = protocol.load_labware(
        load_name="usascientific_12_reservoir_22ml",
        location="C1")
    trash = protocol.load_trash_bin("A3")
    pipette = protocol.load_instrument(
        instrument_name="flex_1channel_1000",
        mount="left",
        tip_racks=[tiprack_1, tiprack_2])
    # Dispense diluent
    pipette.distribute(50, reservoir["A12"], plate.wells())

    # loop through each row
    for i in range(8):
        # save the source well and destination column to variables
        source = reservoir.wells()[i]
        row = plate.rows()[i]

    # transfer 30 µL of source to first well in column
    pipette.transfer(30, source, row[0], mix_after=(3, 25))

    # dilute the sample down the column
    pipette.transfer(
        30, row[:11], row[1:],
        mix_after=(3, 25))

Notice here how the code sample loops through the rows and uses slicing to distribute the diluent. For information about these features, see the Loops and Air Gaps examples above. See also, the Commands section of the Tutorial.

Plate Mapping

This protocol dispenses different volumes of liquids to a well plate and automatically refills the pipette when empty.

FlexOT-2

from opentrons import protocol_api

requirements = {"robotType": "Flex", "apiLevel": "2.17"}

def run(protocol: protocol_api.ProtocolContext):
    plate = protocol.load_labware(
        load_name="corning_96_wellplate_360ul_flat",
        location="D1")
    tiprack_1 = protocol.load_labware(
        load_name="opentrons_flex_96_tiprack_200ul",
        location="D2")
    tiprack_2 = protocol.load_labware(
        load_name="opentrons_flex_96_tiprack_200ul",
        location="D3")
    reservoir = protocol.load_labware(
        load_name="usascientific_12_reservoir_22ml",
        location="C1")
    trash = protocol.load_trash_bin("A3")
    pipette = protocol.load_instrument(
        instrument_name="flex_1channel_1000",
        mount="right",
    tip_racks=[tiprack_1, tiprack_2])

    # Volume amounts are for demonstration purposes only
    water_volumes = [
        1,  2,  3,  4,  5,  6,  7,  8,
        9,  10, 11, 12, 13, 14, 15, 16,
        17, 18, 19, 20, 21, 22, 23, 24,
        25, 26, 27, 28, 29, 30, 31, 32,
        33, 34, 35, 36, 37, 38, 39, 40,
        41, 42, 43, 44, 45, 46, 47, 48,
        49, 50, 51, 52, 53, 54, 55, 56,
        57, 58, 59, 60, 61, 62, 63, 64,
        65, 66, 67, 68, 69, 70, 71, 72,
        73, 74, 75, 76, 77, 78, 79, 80,
        81, 82, 83, 84, 85, 86, 87, 88,
        89, 90, 91, 92, 93, 94, 95, 96
        ]

    pipette.distribute(water_volumes, reservoir["A12"], plate.wells())