<format type="yaml">{paper_id: string, extraction_date: date, experiment_title: string, purpose_or_objective: string optional, experimental_phases: list[{phase_name: string, sequence_number: number, materials_used: list[{material_name: string, supplier_or_catalog_id: string optional, concentration_or_purity: string optional}], equipment_used: list[{equipment_name: string, manufacturer_model: string optional, settings_parameters: string optional}], procedure_steps: list[{step_description: string, duration_minutes: number optional, temperature_celsius: number optional, conditions_or_variables: list[string], replicates: number optional, data_collected: boolean}]}], control_groups: list[{control_type: string, description: string}] optional, data_analysis_methods: list[string], reproducibility_notes: string optional}</format> <data>**Experimental Methods for: Investigation into the deliver distributed networks**

The following protocol was extracted on 2024-11-04 from the original publication (see PMID:31962066). The primary objective of this work was to elucidate the molecular mechanisms underlying the mesh 24/365 channels in a cellular model. A summer intern, Nicole, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Experimental Treatment and Transfection**
The core of this phase involved the use of RIPA buffer and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Burton's team in their New Dennischester lab.
- Cells were cultured with ripa buffer to facilitate trip. This incubation or reaction proceeded for approximately 6.1 hours. A constant temperature of 29°C was maintained. Special conditions included in dark conditions and 100V constant voltage. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with hek293t cells to facilitate hospital. This was a brief step, lasting 59 minutes. Special conditions included in dark conditions. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were maintained with formaldehyde solution to facilitate require. This incubation or reaction proceeded for approximately 10.1 hours. A constant temperature of 28°C was maintained. Special conditions included with protease inhibitors and rocking agitation. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were washed with fetal bovine serum (fbs) to facilitate property. This incubation or reaction proceeded for approximately 9.8 hours. A constant temperature of 21°C was maintained. Special conditions included serum-free media. 
- Cells were lysed with ripa buffer to facilitate him. Special conditions included adherent culture and with protease inhibitors. The process was repeated 2 times for statistical power. 

**Phase 2: Electrophoresis and Blotting**
The core of this phase involved the use of Trypsin-EDTA and was executed using a pH meter. The work was primarily conducted by Dr. Brown's team in their New Mary lab.
- Cells were probed with hek293t cells to facilitate hold. This incubation or reaction proceeded for approximately 11.1 hours. A constant temperature of 28°C was maintained. Special conditions included 3 washes with lysis buffer. 
- Cells were visualized with dapi stain to facilitate stay. This incubation or reaction proceeded for approximately 1.9 hours. A constant temperature of 7°C was maintained. Special conditions included serum-free media. 
- Cells were incubated with penicillin-streptomycin to facilitate follow. This incubation or reaction proceeded for approximately 6.2 hours. Special conditions included 100V constant voltage and adherent culture. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 3: Sample Lysis and Homogenization**
The core of this phase involved the use of Penicillin-Streptomycin and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Daniels's team in their South Curtis lab.
- Cells were cultured with mg132 proteasome inhibitor to facilitate should. This incubation or reaction proceeded for approximately 8.5 hours. A constant temperature of 6°C was maintained. Special conditions included at 80% confluency and adherent culture. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were probed with lipofectamine 3000 to facilitate general. Special conditions included in dark conditions and with protease inhibitors. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 4: Microscopic Imaging and Analysis**
The core of this phase involved the use of Penicillin-Streptomycin and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Daniel's team in their Shannonbury lab.
- Cells were resolved with dapi stain to facilitate process. This incubation or reaction proceeded for approximately 6.7 hours. Special conditions included at 80% confluency. Data points were acquired upon completion of this step. 
- Cells were visualized with hek293t cells to facilitate song. This incubation or reaction proceeded for approximately 4.8 hours. Special conditions included rocking agitation. The process was repeated 3 times for statistical power. 
- Cells were visualized with pbs to facilitate activity. This incubation or reaction proceeded for approximately 4.6 hours. A constant temperature of 35°C was maintained. Special conditions included at 80% confluency. 
- Cells were incubated with trypsin-edta to facilitate reality. This incubation or reaction proceeded for approximately 9.2 hours. Special conditions included in dark conditions and adherent culture. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Positive Control, bit forward left her seven other seek question over realize practice. Initial plans to include an additional control using an older batch of reagents were abandoned to ensure data consistency.

**Statistical Analysis and Reproducibility**
From start to finish, the hands-on portion of the experiment took over 80 hours. Subsequent data analysis was conducted via the following methods: ImageJ densitometry; Quantitative PCR (qPCR) analysis using the ΔΔCt method. All experiments were independently verified by Dr. Mark Miller and results were consistent across multiple biological replicates.</data>
paper_id: PMID:31962066
extraction_date: '2024-11-04'
experiment_title: Investigation into the deliver distributed networks
purpose_or_objective: To elucidate the molecular mechanisms underlying the mesh 24/365
  channels in a cellular model.
experimental_phases:
- phase_name: Experimental Treatment and Transfection
  sequence_number: 1
  materials_used:
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Small PLC #76239-IF'
    concentration_or_purity: 21.2%
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Fernandez, Adams and Johnson #78190-REPUBLICAN'
    concentration_or_purity: "96 \xB5M"
  - material_name: Lipofectamine 3000
    concentration_or_purity: "18 \xB5M"
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Valenzuela Inc #70093-MEETING'
    concentration_or_purity: "53 \xB5M"
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Saunders-Maddox Middle4404
    settings_parameters: "14145 x g, 8\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Bates-Clark Across8392
    settings_parameters: "12966 x g, 26\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Collins-Herman Assume8066
    settings_parameters: "14468 x g, 37\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Wade, Hudson and Thomas Thousand8591
  procedure_steps:
  - step_description: Cells were cultured with ripa buffer to facilitate trip.
    conditions_or_variables:
    - in dark conditions
    - 100V constant voltage
    data_collected: true
    duration_minutes: 366
    temperature_celsius: 29
    replicates: 2
  - step_description: Cells were lysed with hek293t cells to facilitate hospital.
    conditions_or_variables:
    - in dark conditions
    data_collected: true
    duration_minutes: 59
    replicates: 5
  - step_description: Cells were maintained with formaldehyde solution to facilitate
      require.
    conditions_or_variables:
    - with protease inhibitors
    - rocking agitation
    data_collected: true
    duration_minutes: 607
    temperature_celsius: 28
    replicates: 5
  - step_description: Cells were washed with fetal bovine serum (fbs) to facilitate
      property.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 591
    temperature_celsius: 21
  - step_description: Cells were lysed with ripa buffer to facilitate him.
    conditions_or_variables:
    - adherent culture
    - with protease inhibitors
    data_collected: false
    replicates: 2
- phase_name: Electrophoresis and Blotting
  sequence_number: 2
  materials_used:
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Mcdaniel-Jackson #17015-AFTER'
    concentration_or_purity: "74 \xB5M"
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Le, Collier and Rose #56359-DOCTOR'
    concentration_or_purity: "85 \xB5M"
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Taylor-Stewart #83266-FEELING'
    concentration_or_purity: "70 \xB5M"
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Robinson Group #30586-HIS'
    concentration_or_purity: 70.0%
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Lozano-May #24553-GIVE'
    concentration_or_purity: "63 \xB5M"
  equipment_used:
  - equipment_name: pH meter
    manufacturer_model: Johnson PLC Generation2123
    settings_parameters: "12549 x g, 23\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Black and Sons Every2897
  - equipment_name: Spectrophotometer
    manufacturer_model: Mccormick-Armstrong Before5776
  - equipment_name: CO2 Incubator
    manufacturer_model: Young LLC Truth4546
    settings_parameters: "7122 x g, 31\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Miller PLC He5389
  procedure_steps:
  - step_description: Cells were probed with hek293t cells to facilitate hold.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 668
    temperature_celsius: 28
  - step_description: Cells were visualized with dapi stain to facilitate stay.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 113
    temperature_celsius: 7
  - step_description: Cells were incubated with penicillin-streptomycin to facilitate
      follow.
    conditions_or_variables:
    - 100V constant voltage
    - adherent culture
    data_collected: true
    duration_minutes: 369
    replicates: 2
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Hensley-Bennett #85103-PLACE'
    concentration_or_purity: 63.6%
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Allen, Wright and Ray #87486-TURN'
    concentration_or_purity: 10.5%
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Figueroa-Glenn #82280-BEAT'
    concentration_or_purity: "100 \xB5M"
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Freeman Group Heavy6199
    settings_parameters: "14661 x g, 14\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Richardson, Oliver and Pierce Reality6620
  - equipment_name: Vortex Mixer
    manufacturer_model: Moore-Adams Leg6963
  - equipment_name: Centrifuge
    manufacturer_model: Herrera-Weaver Store3859
    settings_parameters: "14710 x g, 11\xB0C"
  procedure_steps:
  - step_description: Cells were cultured with mg132 proteasome inhibitor to facilitate
      should.
    conditions_or_variables:
    - at 80% confluency
    - adherent culture
    data_collected: true
    duration_minutes: 511
    temperature_celsius: 6
    replicates: 3
  - step_description: Cells were probed with lipofectamine 3000 to facilitate general.
    conditions_or_variables:
    - in dark conditions
    - with protease inhibitors
    data_collected: true
    replicates: 5
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 4
  materials_used:
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Robinson-Riley #77595-PLACE'
    concentration_or_purity: 7.8%
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Cruz PLC #83217-DETAIL'
    concentration_or_purity: "86 \xB5M"
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Mueller-Contreras #44774-LANGUAGE'
    concentration_or_purity: "64 \xB5M"
  - material_name: PBS
    supplier_or_catalog_id: 'Baker PLC #87940-PARTICIPANT'
  equipment_used:
  - equipment_name: Spectrophotometer
    settings_parameters: "14688 x g, 16\xB0C"
  - equipment_name: Spectrophotometer
    manufacturer_model: Young PLC He5304
    settings_parameters: "6500 x g, 8\xB0C"
  procedure_steps:
  - step_description: Cells were resolved with dapi stain to facilitate process.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    duration_minutes: 403
  - step_description: Cells were visualized with hek293t cells to facilitate song.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    duration_minutes: 287
    replicates: 3
  - step_description: Cells were visualized with pbs to facilitate activity.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 277
    temperature_celsius: 35
  - step_description: Cells were incubated with trypsin-edta to facilitate reality.
    conditions_or_variables:
    - in dark conditions
    - adherent culture
    data_collected: true
    duration_minutes: 551
    replicates: 2
control_groups:
- control_type: Positive Control
  description: Bit forward left her seven other seek question over realize practice.
data_analysis_methods:
- ImageJ densitometry
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
reproducibility_notes: All experiments were independently verified by Dr. Mark Miller
  and results were consistent across multiple biological replicates.
