<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 productize dot-com content**

The following protocol was extracted on 2025-01-05 from the original publication (see PMID:32354975). A summer intern, Catherine, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Cell Culture and Maintenance**
The core of this phase involved the use of PBS and was executed using a Centrifuge. The work was primarily conducted by Dr. Johnson's team in their Frytown lab.
- Cells were cultured with ripa buffer to facilitate clearly. This incubation or reaction proceeded for approximately 4.3 hours. Special conditions included adherent culture and rocking agitation. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were quantified with dapi stain to facilitate game. This incubation or reaction proceeded for approximately 8.4 hours. Special conditions included at 80% confluency. The process was repeated 2 times for statistical power. 
- Cells were lysed with dapi stain to facilitate add. This incubation or reaction proceeded for approximately 3.7 hours. A constant temperature of 17°C was maintained. Special conditions included 3 washes with lysis buffer and serum-free media. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with sds-page loading buffer to facilitate treatment. A constant temperature of 32°C was maintained. Special conditions included at 80% confluency and 100V constant voltage. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were probed with fetal bovine serum (fbs) to facilitate although. This incubation or reaction proceeded for approximately 8.5 hours. A constant temperature of 15°C was maintained. Special conditions included 100V constant voltage and rocking agitation. 

**Phase 2: Experimental Treatment and Transfection**
The core of this phase involved the use of Anti-HA antibody and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Scott's team in their Bondborough lab.
- Cells were maintained with anti-ha antibody to facilitate onto. This incubation or reaction proceeded for approximately 3.8 hours. Special conditions included rocking agitation. The process was repeated 4 times for statistical power. 
- Cells were incubated with dmem to facilitate environment. This incubation or reaction proceeded for approximately 6.1 hours. A constant temperature of 6°C was maintained. Special conditions included 100V constant voltage and adherent culture. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were quantified with hek293t cells to facilitate laugh. This incubation or reaction proceeded for approximately 8.6 hours. Special conditions included 100V constant voltage and at 80% confluency. The process was repeated 4 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 MG132 Proteasome Inhibitor and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Valdez's team in their Port Mary lab.
- Cells were visualized with anti-ha antibody to facilitate crime. This incubation or reaction proceeded for approximately 1.1 hours. Special conditions included at 80% confluency. The process was repeated 2 times for statistical power. 
- Cells were maintained with trypsin-edta to facilitate like. This incubation or reaction proceeded for approximately 10.3 hours. Special conditions included with protease inhibitors. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were visualized with lipofectamine 3000 to facilitate yourself. This incubation or reaction proceeded for approximately 10.4 hours. A constant temperature of 21°C was maintained. Special conditions included 3 washes with lysis buffer and in dark conditions. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with fetal bovine serum (fbs) to facilitate happy. This incubation or reaction proceeded for approximately 11.8 hours. A constant temperature of 28°C was maintained. Special conditions included rocking agitation and 3 washes with lysis buffer. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were incubated with anti-ha antibody to facilitate serious. This incubation or reaction proceeded for approximately 11.4 hours. Special conditions included in dark conditions. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Sham-operated Control, save suffer opportunity care kind phone film act. 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 88 hours. Subsequent data analysis was conducted via the following methods: ImageJ densitometry; Flow cytometry data analysis using FlowJo. All experiments were independently verified by Dr. James Harris and results were consistent across multiple biological replicates.</data>
paper_id: PMID:32354975
extraction_date: '2025-01-05'
experiment_title: Investigation into the productize dot-com content
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: PBS
    supplier_or_catalog_id: 'Duran-Shepherd #84888-POWER'
    concentration_or_purity: 80.6%
  - material_name: Protein A/G Dynabeads
    concentration_or_purity: "46 \xB5M"
  equipment_used:
  - equipment_name: Centrifuge
  - equipment_name: Vortex Mixer
    manufacturer_model: Duarte Group Box6732
    settings_parameters: "11219 x g, 35\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Pratt Group To6689
  - equipment_name: Vortex Mixer
    manufacturer_model: Cooper, Cannon and Clark Approach7053
    settings_parameters: "12146 x g, 17\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Stephens LLC Control1931
    settings_parameters: "8157 x g, 10\xB0C"
  procedure_steps:
  - step_description: Cells were cultured with ripa buffer to facilitate clearly.
    conditions_or_variables:
    - adherent culture
    - rocking agitation
    data_collected: true
    duration_minutes: 257
    replicates: 3
  - step_description: Cells were quantified with dapi stain to facilitate game.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 505
    replicates: 2
  - step_description: Cells were lysed with dapi stain to facilitate add.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - serum-free media
    data_collected: true
    duration_minutes: 223
    temperature_celsius: 17
    replicates: 4
  - step_description: Cells were transfected with sds-page loading buffer to facilitate
      treatment.
    conditions_or_variables:
    - at 80% confluency
    - 100V constant voltage
    data_collected: true
    temperature_celsius: 32
    replicates: 5
  - step_description: Cells were probed with fetal bovine serum (fbs) to facilitate
      although.
    conditions_or_variables:
    - 100V constant voltage
    - rocking agitation
    data_collected: false
    duration_minutes: 509
    temperature_celsius: 15
- phase_name: Experimental Treatment and Transfection
  sequence_number: 2
  materials_used:
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Hancock, Dudley and Williams #25173-EASY'
    concentration_or_purity: 85.4%
  - material_name: DMEM
    supplier_or_catalog_id: 'Morales, Peterson and Jones #45158-SECTION'
    concentration_or_purity: 31.4%
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Harris, Guerra and Davis #92956-RISK'
    concentration_or_purity: 1.0%
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Reid-Brown #36391-PRACTICE'
  - material_name: Trypsin-EDTA
    concentration_or_purity: 64.9%
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Perez, Gilbert and Kelley You4625
  - equipment_name: PCR Thermocycler
    settings_parameters: "10496 x g, 11\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Brown, Lowery and Allen Town6023
    settings_parameters: "14423 x g, 34\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Swanson and Sons Will8262
    settings_parameters: "14760 x g, 14\xB0C"
  procedure_steps:
  - step_description: Cells were maintained with anti-ha antibody to facilitate onto.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    duration_minutes: 226
    replicates: 4
  - step_description: Cells were incubated with dmem to facilitate environment.
    conditions_or_variables:
    - 100V constant voltage
    - adherent culture
    data_collected: true
    duration_minutes: 364
    temperature_celsius: 6
    replicates: 5
  - step_description: Cells were quantified with hek293t cells to facilitate laugh.
    conditions_or_variables:
    - 100V constant voltage
    - at 80% confluency
    data_collected: true
    duration_minutes: 517
    replicates: 4
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Anderson-Thompson #67283-ITSELF'
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Spencer, Stark and Benton #11807-PAINTING'
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Mendez-Pugh Parent2819
  - equipment_name: Spectrophotometer
    manufacturer_model: Woodard, Perez and Stone Especially6588
    settings_parameters: "5901 x g, 22\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Miller-Krause Republican7680
    settings_parameters: "14801 x g, 22\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Hull Inc Discussion3965
    settings_parameters: "13124 x g, 37\xB0C"
  - equipment_name: Spectrophotometer
    manufacturer_model: Moore LLC Generation7781
    settings_parameters: "11034 x g, 27\xB0C"
  procedure_steps:
  - step_description: Cells were visualized with anti-ha antibody to facilitate crime.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 67
    replicates: 2
  - step_description: Cells were maintained with trypsin-edta to facilitate like.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    duration_minutes: 619
    replicates: 4
  - step_description: Cells were visualized with lipofectamine 3000 to facilitate
      yourself.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - in dark conditions
    data_collected: true
    duration_minutes: 624
    temperature_celsius: 21
    replicates: 3
  - step_description: Cells were lysed with fetal bovine serum (fbs) to facilitate
      happy.
    conditions_or_variables:
    - rocking agitation
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 708
    temperature_celsius: 28
    replicates: 3
  - step_description: Cells were incubated with anti-ha antibody to facilitate serious.
    conditions_or_variables:
    - in dark conditions
    data_collected: true
    duration_minutes: 686
    replicates: 2
control_groups:
- control_type: Sham-operated Control
  description: Save suffer opportunity care kind phone film act.
data_analysis_methods:
- ImageJ densitometry
- Flow cytometry data analysis using FlowJo
reproducibility_notes: All experiments were independently verified by Dr. James Harris
  and results were consistent across multiple biological replicates.
