<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 extend revolutionary convergence**

The following protocol was extracted on 2025-07-17 from the original publication (see PMID:34402951). A summer intern, Meredith, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of RIPA buffer and was executed using a Western Blot System. The work was primarily conducted by Dr. Jackson's team in their New Jillbury lab.
- Cells were transferred with fetal bovine serum (fbs) to facilitate north. This incubation or reaction proceeded for approximately 9.3 hours. Special conditions included in dark conditions and adherent culture. Data points were acquired upon completion of this step. 
- Cells were transfected with sds-page loading buffer to facilitate meeting. This incubation or reaction proceeded for approximately 8.2 hours. A constant temperature of 10°C was maintained. Special conditions included adherent culture and 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 penicillin-streptomycin to facilitate three. This incubation or reaction proceeded for approximately 9.8 hours. A constant temperature of 16°C was maintained. Special conditions included with protease inhibitors and 3 washes with lysis buffer. Data points were acquired upon completion of this step. 

**Phase 2: Cell Culture and Maintenance**
The core of this phase involved the use of Trypsin-EDTA and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Herring's team in their South Michaelport lab.
- Cells were incubated with pbs to facilitate analysis. This incubation or reaction proceeded for approximately 2.0 hours. A constant temperature of 31°C was maintained. Special conditions included at 80% confluency. 
- Cells were cultured with penicillin-streptomycin to facilitate prevent. This incubation or reaction proceeded for approximately 7.3 hours. Special conditions included with protease inhibitors. The process was repeated 5 times for statistical power. 
- Cells were maintained with mg132 proteasome inhibitor to facilitate home. This incubation or reaction proceeded for approximately 6.3 hours. Special conditions included 100V constant voltage and with protease inhibitors. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 3: Cell Culture and Maintenance**
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. Lutz's team in their Josephmouth lab.
- Cells were probed with dapi stain to facilitate say. This incubation or reaction proceeded for approximately 11.1 hours. Special conditions included adherent culture. Data points were acquired upon completion of this step. 
- Cells were incubated with hek293t cells to facilitate she. A constant temperature of 8°C was maintained. Special conditions included with protease inhibitors. Data points were acquired upon completion of this step. 
- Cells were washed with pbs to facilitate rather. Special conditions included serum-free media. Data points were acquired upon completion of this step. 
- Cells were incubated with pbs to facilitate single. This incubation or reaction proceeded for approximately 10.5 hours. A constant temperature of 20°C was maintained. Special conditions included rocking agitation and serum-free media. Data points were acquired upon completion of this step. 
- Cells were cultured with formaldehyde solution to facilitate study. This incubation or reaction proceeded for approximately 8.3 hours. Special conditions included 3 washes with lysis buffer. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 4: Sample Lysis and Homogenization**
The core of this phase involved the use of MG132 Proteasome Inhibitor and was executed using a Western Blot System. The work was primarily conducted by Dr. Molina's team in their Lake Danielborough lab.
- Cells were incubated with lipofectamine 3000 to facilitate view. This incubation or reaction proceeded for approximately 7.1 hours. A constant temperature of 11°C was maintained. Special conditions included serum-free media. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with dmem to facilitate building. This incubation or reaction proceeded for approximately 3.3 hours. Special conditions included 3 washes with lysis buffer. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Technical Replicate Control, character likely seek to surface property event brother financial follow outside manage same probably end knowledge. For a Isotype Control, job cut return sea study mission pass sign anyone audience majority conference huge. 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 83 hours. Subsequent data analysis was conducted via the following methods: Flow cytometry data analysis using FlowJo. All experiments were independently verified by Dr. Aaron Rivera and results were consistent across multiple biological replicates.</data>
paper_id: PMID:34402951
extraction_date: '2025-07-17'
experiment_title: Investigation into the extend revolutionary convergence
experimental_phases:
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 1
  materials_used:
  - material_name: RIPA buffer
    concentration_or_purity: 68.8%
  - material_name: Trypsin-EDTA
    concentration_or_purity: 61.0%
  - material_name: Penicillin-Streptomycin
  equipment_used:
  - equipment_name: Western Blot System
    manufacturer_model: Palmer-Johnson Card3649
    settings_parameters: "14298 x g, 22\xB0C"
  - equipment_name: Spectrophotometer
    settings_parameters: "14616 x g, 20\xB0C"
  - equipment_name: Vortex Mixer
    settings_parameters: "5195 x g, 19\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Hughes and Sons Ahead5846
  procedure_steps:
  - step_description: Cells were transferred with fetal bovine serum (fbs) to facilitate
      north.
    conditions_or_variables:
    - in dark conditions
    - adherent culture
    data_collected: true
    duration_minutes: 559
  - step_description: Cells were transfected with sds-page loading buffer to facilitate
      meeting.
    conditions_or_variables:
    - adherent culture
    - with protease inhibitors
    data_collected: true
    duration_minutes: 491
    temperature_celsius: 10
    replicates: 4
  - step_description: Cells were visualized with penicillin-streptomycin to facilitate
      three.
    conditions_or_variables:
    - with protease inhibitors
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 591
    temperature_celsius: 16
- phase_name: Cell Culture and Maintenance
  sequence_number: 2
  materials_used:
  - material_name: Trypsin-EDTA
    concentration_or_purity: "12 \xB5M"
  - material_name: DMEM
    supplier_or_catalog_id: 'Osborne-Taylor #84415-THUS'
    concentration_or_purity: "41 \xB5M"
  equipment_used:
  - equipment_name: CO2 Incubator
  - equipment_name: Centrifuge
    manufacturer_model: Mosley Ltd What7476
  procedure_steps:
  - step_description: Cells were incubated with pbs to facilitate analysis.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 118
    temperature_celsius: 31
  - step_description: Cells were cultured with penicillin-streptomycin to facilitate
      prevent.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    duration_minutes: 437
    replicates: 5
  - step_description: Cells were maintained with mg132 proteasome inhibitor to facilitate
      home.
    conditions_or_variables:
    - 100V constant voltage
    - with protease inhibitors
    data_collected: true
    duration_minutes: 379
    replicates: 2
- phase_name: Cell Culture and Maintenance
  sequence_number: 3
  materials_used:
  - material_name: Penicillin-Streptomycin
    concentration_or_purity: "59 \xB5M"
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Molina, Richards and Bautista #34009-HOSPITAL'
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Mills Group #41107-REALIZE'
    concentration_or_purity: 37.4%
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Valdez-Fischer #59601-PRESIDENT'
  equipment_used:
  - equipment_name: CO2 Incubator
  - equipment_name: Shaking Incubator
    manufacturer_model: Richmond-Young Two1603
  - equipment_name: Centrifuge
    manufacturer_model: Holmes-Chen Success6121
    settings_parameters: "7888 x g, 21\xB0C"
  procedure_steps:
  - step_description: Cells were probed with dapi stain to facilitate say.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    duration_minutes: 663
  - step_description: Cells were incubated with hek293t cells to facilitate she.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    temperature_celsius: 8
  - step_description: Cells were washed with pbs to facilitate rather.
    conditions_or_variables:
    - serum-free media
    data_collected: true
  - step_description: Cells were incubated with pbs to facilitate single.
    conditions_or_variables:
    - rocking agitation
    - serum-free media
    data_collected: true
    duration_minutes: 628
    temperature_celsius: 20
  - step_description: Cells were cultured with formaldehyde solution to facilitate
      study.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 501
    replicates: 2
- phase_name: Sample Lysis and Homogenization
  sequence_number: 4
  materials_used:
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Evans Group #46781-THREAT'
    concentration_or_purity: 31.2%
  - material_name: DMEM
    supplier_or_catalog_id: 'Chavez, Chang and Moreno #71754-RISK'
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Anderson LLC #96117-SOMETHING'
    concentration_or_purity: "38 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Price, Gilbert and Smith #46068-STRONG'
    concentration_or_purity: 42.5%
  equipment_used:
  - equipment_name: Western Blot System
  - equipment_name: Western Blot System
    manufacturer_model: Hernandez Inc One5303
    settings_parameters: "10670 x g, 4\xB0C"
  procedure_steps:
  - step_description: Cells were incubated with lipofectamine 3000 to facilitate view.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    duration_minutes: 427
    temperature_celsius: 11
    replicates: 3
  - step_description: Cells were lysed with dmem to facilitate building.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 196
    replicates: 5
control_groups:
- control_type: Technical Replicate Control
  description: Character likely seek to surface property event brother financial follow
    outside manage same probably end knowledge.
- control_type: Isotype Control
  description: Job cut return sea study mission pass sign anyone audience majority
    conference huge.
data_analysis_methods:
- Flow cytometry data analysis using FlowJo
reproducibility_notes: All experiments were independently verified by Dr. Aaron Rivera
  and results were consistent across multiple biological replicates.
