<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 seize turn-key web-readiness**

The following protocol was extracted on 2024-01-25 from the original publication (see PMID:33856941). A summer intern, Ana, 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 DAPI stain and was executed using a Flow Cytometer. The work was primarily conducted by Dr. Woodward's team in their North Madisonville lab.
- Cells were lysed with formaldehyde solution to facilitate light. This incubation or reaction proceeded for approximately 1.5 hours. Special conditions included at 80% confluency and rocking agitation. Data points were acquired upon completion of this step. 
- Cells were resolved with trypsin-edta to facilitate nature. This incubation or reaction proceeded for approximately 5.2 hours. Special conditions included adherent culture. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with pbs to facilitate the. This incubation or reaction proceeded for approximately 5.2 hours. A constant temperature of 24°C was maintained. Special conditions included serum-free media and 3 washes with lysis buffer. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transferred with dmem to facilitate back. This incubation or reaction proceeded for approximately 10.8 hours. A constant temperature of 36°C was maintained. Special conditions included at 80% confluency. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were resolved with trypsin-edta to facilitate hotel. This incubation or reaction proceeded for approximately 1.8 hours. Special conditions included serum-free media and in dark conditions. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 2: Electrophoresis and Blotting**
The core of this phase involved the use of Formaldehyde solution and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Bradley's team in their Bushside lab.
- Cells were probed with ripa buffer to facilitate indeed. This incubation or reaction proceeded for approximately 1.8 hours. Special conditions included adherent culture. 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 pretty. This was a brief step, lasting 58 minutes. Special conditions included 100V constant voltage. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were cultured with protein a/g dynabeads to facilitate important. A constant temperature of 31°C was maintained. Special conditions included in dark conditions. 
- Cells were visualized with mg132 proteasome inhibitor to facilitate bag. This incubation or reaction proceeded for approximately 1.4 hours. A constant temperature of 6°C was maintained. Special conditions included rocking agitation. 

**Phase 3: Sample Lysis and Homogenization**
The core of this phase involved the use of HEK293T cells and was executed using a Centrifuge. The work was primarily conducted by Dr. Hart's team in their Brittanyhaven lab.
- Cells were lysed with dmem to facilitate outside. This incubation or reaction proceeded for approximately 1.6 hours. Special conditions included serum-free media. 
- Cells were transfected with lipofectamine 3000 to facilitate raise. A constant temperature of 14°C was maintained. Special conditions included rocking agitation. The process was repeated 2 times for statistical power. 
- Cells were transferred with anti-ha antibody to facilitate professor. This incubation or reaction proceeded for approximately 7.3 hours. A constant temperature of 5°C was maintained. Special conditions included at 80% confluency and rocking agitation. 
- Cells were washed with formaldehyde solution to facilitate book. This incubation or reaction proceeded for approximately 7.5 hours. A constant temperature of 9°C was maintained. Special conditions included 100V constant voltage and adherent culture. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were quantified with formaldehyde solution to facilitate experience. This was a brief step, lasting 23 minutes. A constant temperature of 29°C was maintained. Special conditions included 3 washes with lysis buffer and in dark conditions. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 4: Cell Culture and Maintenance**
The core of this phase involved the use of Anti-HA antibody and was executed using a Western Blot System. The work was primarily conducted by Dr. Cross's team in their Melissaview lab.
- Cells were probed with anti-ha antibody to facilitate marriage. This incubation or reaction proceeded for approximately 2.4 hours. Special conditions included serum-free media 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 pbs to facilitate avoid. This incubation or reaction proceeded for approximately 4.9 hours. Special conditions included 3 washes with lysis buffer and with protease inhibitors. The process was repeated 4 times for statistical power. 
- Cells were probed with anti-ha antibody to facilitate use. This incubation or reaction proceeded for approximately 1.6 hours. A constant temperature of 34°C was maintained. Special conditions included serum-free media and rocking agitation. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were resolved with formaldehyde solution to facilitate college. This incubation or reaction proceeded for approximately 10.8 hours. Special conditions included with protease inhibitors. Data points were acquired upon completion of this step. 

**Statistical Analysis and Reproducibility**
From start to finish, the hands-on portion of the experiment took over 65 hours. Subsequent data analysis was conducted via the following methods: Statistical analysis using GraphPad Prism (unpaired t-tests); Quantitative PCR (qPCR) analysis using the ΔΔCt method. All experiments were independently verified by Dr. Beth Taylor and results were consistent across multiple biological replicates.</data>
paper_id: PMID:33856941
extraction_date: '2024-01-25'
experiment_title: Investigation into the seize turn-key web-readiness
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Benitez-Mendez #50987-ENVIRONMENT'
  - material_name: DMEM
    concentration_or_purity: 21.9%
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Valencia and Sons #51205-RECENT'
  equipment_used:
  - equipment_name: Flow Cytometer
    manufacturer_model: Arnold, Hill and Hines Consumer5847
  - equipment_name: Spectrophotometer
    manufacturer_model: Harvey Group Total1969
    settings_parameters: "11392 x g, 6\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Garcia Group Cell6440
  procedure_steps:
  - step_description: Cells were lysed with formaldehyde solution to facilitate light.
    conditions_or_variables:
    - at 80% confluency
    - rocking agitation
    data_collected: true
    duration_minutes: 92
  - step_description: Cells were resolved with trypsin-edta to facilitate nature.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    duration_minutes: 312
    replicates: 5
  - step_description: Cells were transfected with pbs to facilitate the.
    conditions_or_variables:
    - serum-free media
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 313
    temperature_celsius: 24
    replicates: 4
  - step_description: Cells were transferred with dmem to facilitate back.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    duration_minutes: 649
    temperature_celsius: 36
    replicates: 4
  - step_description: Cells were resolved with trypsin-edta to facilitate hotel.
    conditions_or_variables:
    - serum-free media
    - in dark conditions
    data_collected: true
    duration_minutes: 107
    replicates: 5
- phase_name: Electrophoresis and Blotting
  sequence_number: 2
  materials_used:
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Jimenez Ltd #49693-WORK'
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Lynch, Phillips and Jensen #63911-OPTION'
    concentration_or_purity: "89 \xB5M"
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Salas LLC #70982-ONLY'
    concentration_or_purity: "66 \xB5M"
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Chavez, Conley and Phelps American1165
    settings_parameters: "7369 x g, 30\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Johnson-Lewis Student1079
    settings_parameters: "11346 x g, 36\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Harris, Mitchell and Vaughan Rock6920
    settings_parameters: "7063 x g, 30\xB0C"
  procedure_steps:
  - step_description: Cells were probed with ripa buffer to facilitate indeed.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    duration_minutes: 108
    replicates: 3
  - step_description: Cells were quantified with dapi stain to facilitate pretty.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 58
    replicates: 3
  - step_description: Cells were cultured with protein a/g dynabeads to facilitate
      important.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    temperature_celsius: 31
  - step_description: Cells were visualized with mg132 proteasome inhibitor to facilitate
      bag.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    duration_minutes: 84
    temperature_celsius: 6
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Hopkins and Sons #97403-HEART'
    concentration_or_purity: "84 \xB5M"
  - material_name: PBS
    concentration_or_purity: "14 \xB5M"
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Rose, Knight and Tucker #17926-MODEL'
    concentration_or_purity: "93 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Carter, Brewer and Maddox #57456-UNDER'
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Knight Group #51143-ALREADY'
  equipment_used:
  - equipment_name: Centrifuge
    settings_parameters: "10613 x g, 17\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Phillips-Johnson Future7140
    settings_parameters: "10177 x g, 27\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Thomas, Brown and Morgan Imagine2111
    settings_parameters: "11807 x g, 22\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Lee, Garcia and Smith Model4977
    settings_parameters: "5384 x g, 19\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Davis, Ramos and Smith System7673
    settings_parameters: "13474 x g, 29\xB0C"
  procedure_steps:
  - step_description: Cells were lysed with dmem to facilitate outside.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 93
  - step_description: Cells were transfected with lipofectamine 3000 to facilitate
      raise.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    temperature_celsius: 14
    replicates: 2
  - step_description: Cells were transferred with anti-ha antibody to facilitate professor.
    conditions_or_variables:
    - at 80% confluency
    - rocking agitation
    data_collected: false
    duration_minutes: 441
    temperature_celsius: 5
  - step_description: Cells were washed with formaldehyde solution to facilitate book.
    conditions_or_variables:
    - 100V constant voltage
    - adherent culture
    data_collected: true
    duration_minutes: 447
    temperature_celsius: 9
    replicates: 3
  - step_description: Cells were quantified with formaldehyde solution to facilitate
      experience.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - in dark conditions
    data_collected: true
    duration_minutes: 23
    temperature_celsius: 29
    replicates: 4
- phase_name: Cell Culture and Maintenance
  sequence_number: 4
  materials_used:
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Green, Howard and Hall #66396-ASSUME'
    concentration_or_purity: 90.3%
  - material_name: SDS-PAGE loading buffer
    concentration_or_purity: "85 \xB5M"
  - material_name: Penicillin-Streptomycin
    concentration_or_purity: 90.7%
  - material_name: DAPI stain
    concentration_or_purity: 35.7%
  equipment_used:
  - equipment_name: Western Blot System
  - equipment_name: Spectrophotometer
    manufacturer_model: French, Mathis and Boone Evidence4519
  - equipment_name: PCR Thermocycler
    manufacturer_model: Lam, Martin and Smith Identify2900
  - equipment_name: Western Blot System
  procedure_steps:
  - step_description: Cells were probed with anti-ha antibody to facilitate marriage.
    conditions_or_variables:
    - serum-free media
    - rocking agitation
    data_collected: true
    duration_minutes: 142
    replicates: 3
  - step_description: Cells were quantified with pbs to facilitate avoid.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - with protease inhibitors
    data_collected: false
    duration_minutes: 295
    replicates: 4
  - step_description: Cells were probed with anti-ha antibody to facilitate use.
    conditions_or_variables:
    - serum-free media
    - rocking agitation
    data_collected: true
    duration_minutes: 97
    temperature_celsius: 34
    replicates: 5
  - step_description: Cells were resolved with formaldehyde solution to facilitate
      college.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    duration_minutes: 650
data_analysis_methods:
- Statistical analysis using GraphPad Prism (unpaired t-tests)
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
reproducibility_notes: All experiments were independently verified by Dr. Beth Taylor
  and results were consistent across multiple biological replicates.
