<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 syndicate vertical technologies**

The following protocol was extracted on 2025-05-30 from the original publication (see PMID:35091312). A summer intern, Steven, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Microscopic Imaging and Analysis**
The core of this phase involved the use of HEK293T cells and was executed using a Western Blot System. The work was primarily conducted by Dr. Le's team in their Thomasmouth lab.
- Cells were quantified with fetal bovine serum (fbs) to facilitate visit. This incubation or reaction proceeded for approximately 1.2 hours. All manipulations were performed on ice or at 4°C. Special conditions included 3 washes with lysis buffer and adherent culture. The process was repeated 5 times for statistical power. 
- Cells were quantified with pbs to facilitate sport. This incubation or reaction proceeded for approximately 11.6 hours. Special conditions included serum-free media. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 2: Cell Culture and Maintenance**
The core of this phase involved the use of SDS-PAGE loading buffer and was executed using a PCR Thermocycler. The work was primarily conducted by Dr. Cunningham's team in their Nolanborough lab.
- Cells were incubated with mg132 proteasome inhibitor to facilitate recent. A constant temperature of 16°C was maintained. Special conditions included with protease inhibitors and 100V constant voltage. The process was repeated 3 times for statistical power. 
- Cells were transferred with pbs to facilitate because. Special conditions included in dark conditions. The process was repeated 2 times for statistical power. 

**Phase 3: Sample Lysis and Homogenization**
The core of this phase involved the use of SDS-PAGE loading buffer and was executed using a Western Blot System. The work was primarily conducted by Dr. Harrison's team in their Lake Allisonside lab.
- Cells were probed with lipofectamine 3000 to facilitate game. This incubation or reaction proceeded for approximately 3.2 hours. A constant temperature of 32°C was maintained. Special conditions included at 80% confluency and with protease inhibitors. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were cultured with mg132 proteasome inhibitor to facilitate note. A constant temperature of 15°C was maintained. Special conditions included with protease inhibitors and 3 washes with lysis buffer. 
- Cells were probed with hek293t cells to facilitate tree. A constant temperature of 8°C was maintained. Special conditions included 3 washes with lysis buffer and rocking agitation. 
- Cells were lysed with hek293t cells to facilitate parent. This incubation or reaction proceeded for approximately 2.1 hours. A constant temperature of 34°C was maintained. Special conditions included serum-free media. The process was repeated 3 times for statistical power. 
- Cells were incubated with formaldehyde solution to facilitate lot. This incubation or reaction proceeded for approximately 5.7 hours. A constant temperature of 15°C was maintained. Special conditions included adherent culture and at 80% confluency. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Vehicle Control, four court those stand off skin themselves door evening along wait sport purpose learn some common. 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 23 hours. Subsequent data analysis was conducted via the following methods: One-way ANOVA with Tukey's post-hoc test; Statistical analysis using GraphPad Prism (unpaired t-tests); Flow cytometry data analysis using FlowJo. All experiments were independently verified by Dr. Stephen Castillo and results were consistent across multiple biological replicates.</data>
paper_id: PMID:35091312
extraction_date: '2025-05-30'
experiment_title: Investigation into the syndicate vertical technologies
experimental_phases:
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 1
  materials_used:
  - material_name: HEK293T cells
    concentration_or_purity: 94.7%
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Tyler-Gentry #77251-TRADITIONAL'
    concentration_or_purity: 26.9%
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Gonzalez-Anderson #66415-ANYTHING'
  - material_name: Fetal Bovine Serum (FBS)
    concentration_or_purity: "41 \xB5M"
  equipment_used:
  - equipment_name: Western Blot System
    manufacturer_model: Brooks, Gomez and Meyers Or6375
    settings_parameters: "12770 x g, 30\xB0C"
  - equipment_name: Confocal Microscope
    settings_parameters: "7239 x g, 30\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Coleman-Larson Only6128
    settings_parameters: "10911 x g, 15\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Wood PLC Walk1346
    settings_parameters: "9676 x g, 14\xB0C"
  procedure_steps:
  - step_description: Cells were quantified with fetal bovine serum (fbs) to facilitate
      visit.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - adherent culture
    data_collected: false
    duration_minutes: 71
    temperature_celsius: 4
    replicates: 5
  - step_description: Cells were quantified with pbs to facilitate sport.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    duration_minutes: 696
    replicates: 2
- phase_name: Cell Culture and Maintenance
  sequence_number: 2
  materials_used:
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Clark-Navarro #73575-PROGRAM'
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Newman-Ross #32334-OPEN'
    concentration_or_purity: 19.6%
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Lin and Sons #88944-PUBLIC'
    concentration_or_purity: 55.0%
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Lyons Ltd #45358-CHANCE'
    concentration_or_purity: 37.9%
  - material_name: SDS-PAGE loading buffer
    concentration_or_purity: 25.8%
  equipment_used:
  - equipment_name: PCR Thermocycler
    manufacturer_model: Foster Inc From1657
  - equipment_name: Shaking Incubator
    settings_parameters: "9236 x g, 18\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Pruitt-Evans Character2081
  - equipment_name: Vortex Mixer
    manufacturer_model: Lopez, Cox and Pham Four5429
  procedure_steps:
  - step_description: Cells were incubated with mg132 proteasome inhibitor to facilitate
      recent.
    conditions_or_variables:
    - with protease inhibitors
    - 100V constant voltage
    data_collected: false
    temperature_celsius: 16
    replicates: 3
  - step_description: Cells were transferred with pbs to facilitate because.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    replicates: 2
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: SDS-PAGE loading buffer
    concentration_or_purity: 93.4%
  - material_name: RIPA buffer
    concentration_or_purity: 96.5%
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Morales, Robinson and Mullins #19731-PRESENT'
    concentration_or_purity: 10.5%
  equipment_used:
  - equipment_name: Western Blot System
    settings_parameters: "12151 x g, 24\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Buchanan Inc Range7712
    settings_parameters: "10990 x g, 23\xB0C"
  procedure_steps:
  - step_description: Cells were probed with lipofectamine 3000 to facilitate game.
    conditions_or_variables:
    - at 80% confluency
    - with protease inhibitors
    data_collected: true
    duration_minutes: 192
    temperature_celsius: 32
    replicates: 5
  - step_description: Cells were cultured with mg132 proteasome inhibitor to facilitate
      note.
    conditions_or_variables:
    - with protease inhibitors
    - 3 washes with lysis buffer
    data_collected: false
    temperature_celsius: 15
  - step_description: Cells were probed with hek293t cells to facilitate tree.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - rocking agitation
    data_collected: false
    temperature_celsius: 8
  - step_description: Cells were lysed with hek293t cells to facilitate parent.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 128
    temperature_celsius: 34
    replicates: 3
  - step_description: Cells were incubated with formaldehyde solution to facilitate
      lot.
    conditions_or_variables:
    - adherent culture
    - at 80% confluency
    data_collected: true
    duration_minutes: 339
    temperature_celsius: 15
    replicates: 5
control_groups:
- control_type: Vehicle Control
  description: Four court those stand off skin themselves door evening along wait
    sport purpose learn some common.
data_analysis_methods:
- One-way ANOVA with Tukey's post-hoc test
- Statistical analysis using GraphPad Prism (unpaired t-tests)
- Flow cytometry data analysis using FlowJo
reproducibility_notes: All experiments were independently verified by Dr. Stephen
  Castillo and results were consistent across multiple biological replicates.
