def generate_steps_1_2_3(treatment, predicted_IC50=None):
return_val = "PASS"
# * Program variables
blowoff_volume = 10
num_mixes = 3
current_media_reservoir_volume = media_reservoir_volume = 7000
reservoir_z_shift = 0.5 # z shift for deep blocks (Deck Positions 3 and 5)
lambda6_path = "/lambda_stor/data/hudson/instructions/"
# Step 1 variables
culture_plate_column_num = 7 # Changed to column 7 for test on 06/15/21
media_transfer_volume_s1 = 60
culture_transfer_volume_s1 = 30
# dilution_media_volume = 198
half_dilution_media_volume = 99
dilution_culture_volume = 22
culture_plate_mix_volume_s1 = (
30 # best to mix with same volume transferred, ok for min volume
)
growth_plate_mix_volume_s1 = 40
# Step 2 variables
media_transfer_volume_s2 = (
108 # two times = 216 uL (will add 24 uL antibiotic for 1:10 dilution)
)
first_column_transfer_volume_s2 = (
120 # two times = 240uL (to equal volume in 1:10 dilution wells)
)
serial_antibiotic_transfer_volume_s2 = 24
serial_source_mixing_volume_s2 = 80
serial_source_num_mixes_s2 = 8
serial_destination_mixing_volume_s2 = 100
# Step 3 variables
antibiotic_transfer_volume_s3 = 90
antibiotic_mix_volume_s3 = 90
destination_mix_volume_s3 = 100
# * Get location of treatment
try:
treatment_plate_loc, treatment_column = find_treatment_loc(treatment)
except Error as e:
print(f"Unale to locate treatment {treatment}")
raise # need to know locaton of treatment, rest of protocol useless if not specified
# * TODO: handle predicted IC50
# * Create folder to store all instruction files
project = "Campaign1"
project_desc = "col" + str(culture_plate_column_num)
version_num = "v1"
timestamp = str(time.time()).split(".")[0]
directory_name = f"{project}-{project_desc}-{version_num}-{timestamp}"
directory_path = os.path.join(
os.path.realpath(os.path.dirname(lambda6_path)), directory_name)
print(f"Protocol directory created: {directory_path}")
# * create new directory to hold new instructions
try:
os.makedirs(directory_path, exist_ok=True)
except OSError as e:
print(e)
print(
f"failed to create new directory for instructions: {directory_path}"
)
"""
STEP 1: INNOCULATE GROWTH PLATE FROM SOURCE BACTERIA PLATE -----------------------------------------------------------------
"""
# * Initialize soloSoft (step 1)
step1_hso_filename = os.path.join(directory_path, "step_1.hso")
soloSoft = SoloSoft(
filename=step1_hso_filename,
plateList=[
"TipBox.180uL.Axygen-EVF-180-R-S.bluebox",
"Empty",
"DeepBlock.96.VWR-75870-792.sterile",
"Plate.96.Corning-3635.ClearUVAssay",
"DeepBlock.96.VWR-75870-792.sterile",
"Plate.96.Corning-3635.ClearUVAssay",
"Plate.96.Corning-3635.ClearUVAssay",
"Empty",
],
)
# * Fill all columns of empty 96 well plate (corning 3383 or Falcon - ref 353916) with fresh lb media (12 channel in Position 3, column 1)
soloSoft.getTip()
j = 1
for i in range(1, 7): # first half plate = media from column 1
soloSoft.aspirate(
position="Position3",
aspirate_volumes=Reservoir_12col_Agilent_201256_100_BATSgroup().
setColumn(1, media_transfer_volume_s1),
aspirate_shift=[0, 0, reservoir_z_shift],
)
soloSoft.dispense(
position="Position4",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, media_transfer_volume_s1),
dispense_shift=[0, 0, 2],
)
for i in range(7, 13): # second half plate = media from column 2
soloSoft.aspirate(
position="Position3",
aspirate_volumes=Reservoir_12col_Agilent_201256_100_BATSgroup().
setColumn(2, media_transfer_volume_s1),
aspirate_shift=[0, 0, reservoir_z_shift],
)
soloSoft.dispense(
position="Position4",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, media_transfer_volume_s1),
dispense_shift=[0, 0, 2],
)
# * Fill first two columns of culture 10 fold dilution plate with fresh lb media (do in two steps due to 180uL filter tips)
for i in range(2): # first column
soloSoft.aspirate(
position="Position3",
aspirate_volumes=Reservoir_12col_Agilent_201256_100_BATSgroup().
setColumn(1, half_dilution_media_volume),
aspirate_shift=[0, 0, reservoir_z_shift],
)
soloSoft.dispense(
position="Position7",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
1, half_dilution_media_volume),
dispense_shift=[0, 0, 2],
)
for i in range(2): # second column
soloSoft.aspirate(
position="Position3",
aspirate_volumes=Reservoir_12col_Agilent_201256_100_BATSgroup().
setColumn(2, half_dilution_media_volume),
aspirate_shift=[0, 0, reservoir_z_shift],
)
soloSoft.dispense(
position="Position7",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
2, half_dilution_media_volume),
dispense_shift=[0, 0, 2],
)
# * Make culture 10 fold dilution (one column for each half of plate)
for i in range(1, 3):
soloSoft.aspirate(
position="Position5",
aspirate_volumes=DeepBlock_96VWR_75870_792_sterile().setColumn(
culture_plate_column_num, dilution_culture_volume),
aspirate_shift=[0, 0, 2],
mix_at_start=True,
mix_cycles=num_mixes,
mix_volume=culture_plate_mix_volume_s1,
dispense_height=2,
# pre_aspirate=blowoff_volume,
syringe_speed=25,
)
soloSoft.dispense(
position="Position7",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, dilution_culture_volume),
dispense_shift=[0, 0, 2],
mix_at_finish=True,
mix_cycles=num_mixes,
mix_volume=culture_plate_mix_volume_s1,
aspirate_height=2,
syringe_speed=25,
# blowoff=blowoff_volume,
)
# * Add bacteria from 10 fold diluted culture plate (Position 7, column 1 and 2) to growth plate with fresh media (both halves)
for i in range(1, 7): # first half growth plate
soloSoft.aspirate( # already mixed the cells, no need to do it before every transfer
position="Position7",
aspirate_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
1, culture_transfer_volume_s1),
aspirate_shift=[
0,
0,
2,
], # prevents 50 uL tips from going too deep in 96 deep well plate
syringe_speed=25,
)
soloSoft.dispense( # do need to mix at end of transfer
position="Position4",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, culture_transfer_volume_s1),
mix_at_finish=True,
mix_cycles=num_mixes,
mix_volume=growth_plate_mix_volume_s1,
aspirate_height=2,
dispense_shift=[0, 0, 2],
syringe_speed=25,
)
for i in range(7, 13): # second half growth plate
soloSoft.aspirate( # already mixed the cells, no need to do it before every transfer
position="Position7",
aspirate_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
2, culture_transfer_volume_s1),
aspirate_shift=[
0,
0,
2,
], # prevents 50 uL tips from going too deep in 96 deep well plate
syringe_speed=25,
)
soloSoft.dispense( # do need to mix at end of transfer
position="Position4",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, culture_transfer_volume_s1),
mix_at_finish=True,
mix_cycles=num_mixes,
mix_volume=growth_plate_mix_volume_s1,
aspirate_height=2,
dispense_shift=[0, 0, 2],
syringe_speed=25,
)
soloSoft.shuckTip()
soloSoft.savePipeline()
"""
STEP 2: PERFORM SERIAL DILUTIONS ON TREATMENT -------------------------------------------------------------------------------
"""
# * Initialize soloSoft (step 2)
step2_hso_filename = os.path.join(directory_path, "step_2.hso")
soloSoft = SoloSoft(
filename=step2_hso_filename,
plateList=[
"TipBox.180uL.Axygen-EVF-180-R-S.bluebox",
"Empty",
"DeepBlock.96.VWR-75870-792.sterile",
"Plate.96.Corning-3635.ClearUVAssay",
"DeepBlock.96.VWR-75870-792.sterile",
"Plate.96.Corning-3635.ClearUVAssay",
"Plate.96.Corning-3635.ClearUVAssay",
"Empty",
],
)
# * Fill colums 2-6 of generic 96 well plate with lb media (will use for both halves of plate)
soloSoft.getTip()
for i in range(2, 7):
# draws from both lb media wells to prevent running out of media -> TODO: volume management
soloSoft.aspirate( # first lb media well
position="Position3",
aspirate_volumes=Reservoir_12col_Agilent_201256_100_BATSgroup().
setColumn(1, media_transfer_volume_s2),
aspirate_shift=[0, 0, reservoir_z_shift],
# pre_aspirate=blowoff_volume,
)
soloSoft.dispense(
position="Position6",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, media_transfer_volume_s2),
dispense_shift=[0, 0, 2],
# blowoff=blowoff_volume,
)
soloSoft.aspirate( # second lb media well
position="Position3",
aspirate_volumes=Reservoir_12col_Agilent_201256_100_BATSgroup().
setColumn(2, media_transfer_volume_s2),
aspirate_shift=[0, 0, reservoir_z_shift],
# pre_aspirate=blowoff_volume,
)
soloSoft.dispense(
position="Position6",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, media_transfer_volume_s2),
dispense_shift=[0, 0, 2],
# blowoff=blowoff_volume,
)
# * Transfer undiluted treatment stock solution (12 channel in Position 3, 2rd row) into empty first row of serial dilution plate
for i in range(2):
soloSoft.aspirate(
position=treatment_plate_loc,
aspirate_volumes=Reservoir_12col_Agilent_201256_100_BATSgroup().
setColumn(treatment_column, first_column_transfer_volume_s2),
pre_aspirate=blowoff_volume,
mix_at_start=True,
mix_cycles=serial_source_num_mixes_s2,
mix_volume=serial_source_mixing_volume_s2,
aspirate_shift=[0, 0, reservoir_z_shift],
dispense_height=reservoir_z_shift,
)
soloSoft.dispense(
position="Position6",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
1, first_column_transfer_volume_s2),
dispense_shift=[0, 0, 2],
blowoff=blowoff_volume,
# mix_at_finish=True,
# mix_cycles=num_mixes,
# mix_volume=serial_destination_mixing_volume_s2,
aspirate_height=2,
)
# * Serial dilution within Generic 96 well plate (Corning or Falcon) - mix 5 times before and after transfer
for i in range(1, 6):
if i == 4: # switch tips half way through to reduce error
soloSoft.getTip()
soloSoft.aspirate(
position="Position6",
aspirate_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, serial_antibiotic_transfer_volume_s2),
aspirate_shift=[0, 0, 2],
pre_aspirate=blowoff_volume,
mix_at_start=True,
mix_cycles=num_mixes,
mix_volume=serial_destination_mixing_volume_s2,
dispense_height=2,
)
soloSoft.dispense(
position="Position6",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i + 1, serial_antibiotic_transfer_volume_s2),
dispense_shift=[0, 0, 2],
blowoff=blowoff_volume,
mix_at_finish=True,
mix_cycles=num_mixes,
mix_volume=serial_destination_mixing_volume_s2,
aspirate_height=2,
)
# no need to throw away excess volume from last column of serial dilution
soloSoft.shuckTip()
soloSoft.savePipeline()
"""
STEP 3: ADD ANTIBIOTIC TO CULTURE PLATES -------------------------------------------------------------------------------------
"""
# * Initialize soloSoft (step 3)
step3_hso_filename = os.path.join(directory_path, "step_3.hso")
soloSoft = SoloSoft(
filename=step3_hso_filename,
plateList=[
"TipBox.180uL.Axygen-EVF-180-R-S.bluebox",
"Empty",
"DeepBlock.96.VWR-75870-792.sterile",
"Plate.96.Corning-3635.ClearUVAssay",
"DeepBlock.96.VWR-75870-792.sterile",
"Plate.96.Corning-3635.ClearUVAssay",
"Plate.96.Corning-3635.ClearUVAssay",
"Empty",
],
)
soloSoft.getTip()
for i in range(6, 0, -1): # first half of plate
if i == 3: # switch tips half way through to reduce error
soloSoft.getTip()
soloSoft.aspirate(
position="Position6",
aspirate_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, antibiotic_transfer_volume_s3),
mix_at_start=True,
mix_cycles=num_mixes,
mix_volume=antibiotic_mix_volume_s3,
dispense_height=2,
aspirate_shift=[0, 0, 2],
)
soloSoft.dispense(
position="Position4",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, antibiotic_transfer_volume_s3),
mix_at_finish=True,
mix_cycles=num_mixes,
mix_volume=destination_mix_volume_s3,
aspirate_height=2,
dispense_shift=[0, 0, 2],
)
soloSoft.getTip()
for i in range(6, 0, -1): # second half of plate
if i == 3: # switch tips half way through to reduce error
soloSoft.getTip()
soloSoft.aspirate(
position="Position6",
aspirate_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i, antibiotic_transfer_volume_s3),
mix_at_start=True,
mix_cycles=num_mixes,
mix_volume=antibiotic_mix_volume_s3,
dispense_height=2,
aspirate_shift=[0, 0, 2],
)
soloSoft.dispense(
position="Position4",
dispense_volumes=Plate_96_Corning_3635_ClearUVAssay().setColumn(
i + 6, antibiotic_transfer_volume_s3),
mix_at_finish=True,
mix_cycles=num_mixes,
mix_volume=destination_mix_volume_s3,
aspirate_height=2,
dispense_shift=[0, 0, 2],
)
soloSoft.shuckTip()
soloSoft.savePipeline()
"""
ADD ALL STEPS TO SOFTLINX PROTOCOL AND SEND TO HUDSON01 -----------------------------------------------------------------------
"""
# initialize softLinx
softLinx = SoftLinx("Steps_1_2_3",
os.path.join(directory_path, "steps_1_2_3.slvp"))
# define starting plate layout
softLinx.setPlates(
{"SoftLinx.PlateCrane.Stack5": "Plate.96.Corning-3635.ClearUVAssay"})
# restock growth assay plate before run
softLinx.plateCraneMovePlate(["SoftLinx.PlateCrane.Stack5"],
["SoftLinx.Solo.Position4"])
softLinx.plateCraneMoveCrane("SoftLinx.PlateCrane.Safe")
# run all three liquid handling steps (with paths to .hso files on hudson01)
softLinx.soloSoftResetTipCount(
1) # SoloSoft will reset to full tip box before run
softLinx.soloSoftRun("C:\\labautomation\\instructions\\" + directory_name +
"\\" + os.path.basename(step1_hso_filename))
softLinx.soloSoftRun("C:\\labautomation\\instructions\\" + directory_name +
"\\" + os.path.basename(step2_hso_filename))
softLinx.soloSoftRun("C:\\labautomation\\instructions\\" + directory_name +
"\\" + os.path.basename(step3_hso_filename))
# move growth plate to Hidex
softLinx.plateCraneMovePlate(
["SoftLinx.Solo.Position4"],
["SoftLinx.Hidex.Nest"]) # no need to open hidex
softLinx.hidexClose()
softLinx.plateCraneMoveCrane("SoftLinx.PlateCrane.Safe")
softLinx.hidexRun("Campaign1")
# save protocol to write instructions to .slvp file, create .txt manifest, and .ahk remote start file
softLinx.saveProtocol()
"""
SEND NEW INSTRUCTIONS TO WORK CELL (HUDSON01) ------------------------------------------------------------------
"""
try:
# TODO: change to full path on lambda6
child_message_sender = child_pid = Popen(
[
"python",
"../../zeromq/lambda6_send_instructions.py",
"-d",
directory_path,
],
start_new_session=True,
).pid
print("New instruction directory passed to lambda6_send_message.py")
except Error as e:
print(e)
print("Could not send new instructions to hudson01")
return return_val
),
dispense_shift=[0, 0, default_z_shift],
)
# * aspirate/dispense column 1, rows B D F H J L N P (even rows) -> start B1
# change value in first row to 0 -> shortcut to make soloSoft start at Row B
# TODO: Add an easier way to accomplish this
aspirate_volumes_startB = Plate_384_Corning_3540_BlackwClearBottomAssay().setColumn(
1, trans_volume
)
aspirate_volumes_startB[0][0] = 0
soloSoft.aspirate(
position="Position8",
aspirate_volumes=aspirate_volumes_startB,
aspirate_shift=[0, 0, default_z_shift],
)
dispense_volumes_startB = Plate_384_Corning_3540_BlackwClearBottomAssay().setColumn(
2, trans_volume
)
dispense_volumes_startB[0][1] = 0
soloSoft.dispense(
position="Position8",
dispense_volumes=dispense_volumes_startB,
dispense_shift=[0, 0, default_z_shift],
)
soloSoft.shuckTip()
soloSoft.savePipeline()
import sys
import os
from liquidhandling import SoloSoft
soloSoft = SoloSoft(
filename="example.hso",
plateList=[
"TipBox-Corning 200uL",
"Corning 3383",
"Corning 3383",
"Corning 3383",
"Corning 3383",
"Corning 3383",
"Corning 3383",
"Corning 3383",
],
)
soloSoft.getTip()
soloSoft.loop(1)
soloSoft.aspirate()
soloSoft.dispense()
soloSoft.endLoop()
soloSoft.moveArm()
# print(soloSoft.pipeline)
soloSoft.savePipeline(CRLF=True)
