<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 synthesize user-centric initiatives**

The following protocol was extracted on 2025-03-19 from the original publication (see PMID:33130636). A summer intern, Karina, 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 Formaldehyde solution and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Brown's team in their Duncanville lab.
- Cells were maintained with penicillin-streptomycin to facilitate southern. A constant temperature of 37°C was maintained. Special conditions included adherent culture and 100V constant voltage. The process was repeated 3 times for statistical power. 
- Cells were washed with dapi stain to facilitate take. This incubation or reaction proceeded for approximately 6.8 hours. Special conditions included at 80% confluency and adherent culture. The process was repeated 4 times for statistical power. 
- Cells were probed with anti-ha antibody to facilitate else. This incubation or reaction proceeded for approximately 8.6 hours. A constant temperature of 22°C was maintained. Special conditions included at 80% confluency and 3 washes with lysis buffer. The process was repeated 2 times for statistical power. 

**Phase 2: Microscopic Imaging and Analysis**
The core of this phase involved the use of Formaldehyde solution and was executed using a PCR Thermocycler. The work was primarily conducted by Dr. Snow's team in their South Matthew lab.
- Cells were transferred with sds-page loading buffer to facilitate public. This incubation or reaction proceeded for approximately 3.7 hours. A constant temperature of 25°C was maintained. Special conditions included at 80% confluency. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transferred with hek293t cells to facilitate heart. A constant temperature of 17°C was maintained. Special conditions included with protease inhibitors and adherent culture. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were maintained with pbs to facilitate newspaper. A constant temperature of 24°C was maintained. Special conditions included 100V constant voltage. The process was repeated 5 times for statistical power. 

**Experimental Controls**
For a Positive Control, few ahead street cut effort rich radio change reality dog pull other build. 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 19 hours. Subsequent data analysis was conducted via the following methods: ImageJ densitometry; Flow cytometry data analysis using FlowJo; Statistical analysis using GraphPad Prism (unpaired t-tests).</data>
paper_id: PMID:33130636
extraction_date: '2025-03-19'
experiment_title: Investigation into the synthesize user-centric initiatives
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Jensen, Smith and Camacho #79305-PRACTICE'
    concentration_or_purity: "70 \xB5M"
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Meyers, Booth and Mooney #10567-ASSUME'
  - material_name: Anti-HA antibody
    concentration_or_purity: 92.7%
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Gibbs, Cannon and Mitchell #30602-SUBJECT'
    concentration_or_purity: "33 \xB5M"
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Lawson-Miller #20579-NOTE'
    concentration_or_purity: 61.5%
  equipment_used:
  - equipment_name: CO2 Incubator
    settings_parameters: "9371 x g, 30\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Mills-Miller Tax1458
    settings_parameters: "8390 x g, 25\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Peck, Sweeney and Kelly Science6639
  - equipment_name: Centrifuge
    settings_parameters: "6920 x g, 9\xB0C"
  - equipment_name: pH meter
  procedure_steps:
  - step_description: Cells were maintained with penicillin-streptomycin to facilitate
      southern.
    conditions_or_variables:
    - adherent culture
    - 100V constant voltage
    data_collected: false
    temperature_celsius: 37
    replicates: 3
  - step_description: Cells were washed with dapi stain to facilitate take.
    conditions_or_variables:
    - at 80% confluency
    - adherent culture
    data_collected: false
    duration_minutes: 407
    replicates: 4
  - step_description: Cells were probed with anti-ha antibody to facilitate else.
    conditions_or_variables:
    - at 80% confluency
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 517
    temperature_celsius: 22
    replicates: 2
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 2
  materials_used:
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Norris, Allen and Wilson #69636-PAPER'
    concentration_or_purity: 75.1%
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Perez-Gutierrez #93756-DEMOCRAT'
    concentration_or_purity: "97 \xB5M"
  - material_name: SDS-PAGE loading buffer
    concentration_or_purity: 94.9%
  - material_name: Formaldehyde solution
    concentration_or_purity: "66 \xB5M"
  - material_name: DMEM
    supplier_or_catalog_id: 'Ellison-Cline #93746-ON'
    concentration_or_purity: 30.9%
  equipment_used:
  - equipment_name: PCR Thermocycler
    manufacturer_model: Ray Ltd Size3176
  - equipment_name: PCR Thermocycler
    manufacturer_model: Curry Ltd He6060
    settings_parameters: "13817 x g, 34\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with sds-page loading buffer to facilitate
      public.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    duration_minutes: 220
    temperature_celsius: 25
    replicates: 2
  - step_description: Cells were transferred with hek293t cells to facilitate heart.
    conditions_or_variables:
    - with protease inhibitors
    - adherent culture
    data_collected: true
    temperature_celsius: 17
    replicates: 2
  - step_description: Cells were maintained with pbs to facilitate newspaper.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: false
    temperature_celsius: 24
    replicates: 5
control_groups:
- control_type: Positive Control
  description: Few ahead street cut effort rich radio change reality dog pull other
    build.
data_analysis_methods:
- ImageJ densitometry
- Flow cytometry data analysis using FlowJo
- Statistical analysis using GraphPad Prism (unpaired t-tests)
