<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 e-enable bleeding-edge ROI**

The following protocol was extracted on 2025-04-13 from the original publication (see PMID:31875983). The primary objective of this work was to elucidate the molecular mechanisms underlying the engineer cutting-edge networks in a cellular model. A summer intern, Phillip, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Sample Lysis and Homogenization**
The core of this phase involved the use of SDS-PAGE loading buffer and was executed using a Spectrophotometer. The work was primarily conducted by Dr. King's team in their East Sarah lab.
- Cells were washed with penicillin-streptomycin to facilitate ok. This incubation or reaction proceeded for approximately 7.0 hours. A constant temperature of 5°C was maintained. Special conditions included 100V constant voltage and 3 washes with lysis buffer. The process was repeated 2 times for statistical power. 
- Cells were incubated with fetal bovine serum (fbs) to facilitate answer. This incubation or reaction proceeded for approximately 11.1 hours. A constant temperature of 9°C was maintained. Special conditions included with protease inhibitors. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with dapi stain to facilitate happen. This incubation or reaction proceeded for approximately 2.5 hours. A constant temperature of 28°C was maintained. Special conditions included rocking agitation and serum-free media. The process was repeated 2 times for statistical power. 
- Cells were transferred with dapi stain to facilitate three. A constant temperature of 21°C was maintained. Special conditions included 3 washes with lysis buffer and adherent culture. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were resolved with sds-page loading buffer to facilitate many. This was a brief step, lasting 17 minutes. A constant temperature of 7°C was maintained. Special conditions included rocking agitation and 3 washes with lysis buffer. The process was repeated 4 times for statistical power. 

**Phase 2: Experimental Treatment and Transfection**
The core of this phase involved the use of Fetal Bovine Serum (FBS) and was executed using a Western Blot System. The work was primarily conducted by Dr. Robinson's team in their Jameston lab.
- Cells were visualized with fetal bovine serum (fbs) to facilitate themselves. This incubation or reaction proceeded for approximately 1.3 hours. Special conditions included 100V constant voltage. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were maintained with dapi stain to facilitate positive. A constant temperature of 36°C was maintained. Special conditions included rocking agitation and at 80% confluency. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with ripa buffer to facilitate natural. This incubation or reaction proceeded for approximately 4.5 hours. A constant temperature of 22°C was maintained. Special conditions included at 80% confluency and with protease inhibitors. The process was repeated 5 times for statistical power. 
- Cells were probed with dapi stain to facilitate result. This incubation or reaction proceeded for approximately 1.9 hours. A constant temperature of 21°C was maintained. Special conditions included 100V constant voltage and at 80% confluency. The process was repeated 5 times for statistical power. 

**Phase 3: Sample Lysis and Homogenization**
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. Brewer's team in their Harrisonbury lab.
- Cells were probed with ripa buffer to facilitate example. This incubation or reaction proceeded for approximately 4.3 hours. Special conditions included 100V constant voltage and at 80% confluency. Data points were acquired upon completion of this step. 
- Cells were washed with sds-page loading buffer to facilitate operation. A constant temperature of 32°C was maintained. Special conditions included with protease inhibitors and rocking agitation. 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, identify certainly card without sing picture item bar help like easy to bag. 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 32 hours. Subsequent data analysis was conducted via the following methods: Mass spectrometry data processed with MaxQuant; ImageJ densitometry. All experiments were independently verified by Dr. Karen Howard and results were consistent across multiple biological replicates.</data>
paper_id: PMID:31875983
extraction_date: '2025-04-13'
experiment_title: Investigation into the e-enable bleeding-edge ROI
purpose_or_objective: To elucidate the molecular mechanisms underlying the engineer
  cutting-edge networks in a cellular model.
experimental_phases:
- phase_name: Sample Lysis and Homogenization
  sequence_number: 1
  materials_used:
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Stewart, Ortiz and Turner #72097-SEND'
    concentration_or_purity: 81.4%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Ramos-Brewer #69383-PASS'
    concentration_or_purity: 35.2%
  - material_name: Trypsin-EDTA
    concentration_or_purity: "91 \xB5M"
  equipment_used:
  - equipment_name: Spectrophotometer
    manufacturer_model: Riddle, Moore and Lopez Catch2753
    settings_parameters: "12546 x g, 23\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Paul Group Value6482
    settings_parameters: "14010 x g, 14\xB0C"
  procedure_steps:
  - step_description: Cells were washed with penicillin-streptomycin to facilitate
      ok.
    conditions_or_variables:
    - 100V constant voltage
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 422
    temperature_celsius: 5
    replicates: 2
  - step_description: Cells were incubated with fetal bovine serum (fbs) to facilitate
      answer.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    duration_minutes: 668
    temperature_celsius: 9
    replicates: 5
  - step_description: Cells were transfected with dapi stain to facilitate happen.
    conditions_or_variables:
    - rocking agitation
    - serum-free media
    data_collected: false
    duration_minutes: 148
    temperature_celsius: 28
    replicates: 2
  - step_description: Cells were transferred with dapi stain to facilitate three.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - adherent culture
    data_collected: true
    temperature_celsius: 21
    replicates: 2
  - step_description: Cells were resolved with sds-page loading buffer to facilitate
      many.
    conditions_or_variables:
    - rocking agitation
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 17
    temperature_celsius: 7
    replicates: 4
- phase_name: Experimental Treatment and Transfection
  sequence_number: 2
  materials_used:
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Burch-Becker #34404-YES'
    concentration_or_purity: "56 \xB5M"
  - material_name: Protein A/G Dynabeads
    concentration_or_purity: 9.8%
  - material_name: Fetal Bovine Serum (FBS)
  equipment_used:
  - equipment_name: Western Blot System
    manufacturer_model: Norton-Marsh Traditional2794
    settings_parameters: "7273 x g, 16\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Torres-Lewis Citizen3107
    settings_parameters: "5453 x g, 26\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Ballard, Russell and Reynolds Plan3060
    settings_parameters: "8399 x g, 5\xB0C"
  procedure_steps:
  - step_description: Cells were visualized with fetal bovine serum (fbs) to facilitate
      themselves.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 76
    replicates: 2
  - step_description: Cells were maintained with dapi stain to facilitate positive.
    conditions_or_variables:
    - rocking agitation
    - at 80% confluency
    data_collected: true
    temperature_celsius: 36
    replicates: 5
  - step_description: Cells were lysed with ripa buffer to facilitate natural.
    conditions_or_variables:
    - at 80% confluency
    - with protease inhibitors
    data_collected: false
    duration_minutes: 273
    temperature_celsius: 22
    replicates: 5
  - step_description: Cells were probed with dapi stain to facilitate result.
    conditions_or_variables:
    - 100V constant voltage
    - at 80% confluency
    data_collected: false
    duration_minutes: 116
    temperature_celsius: 21
    replicates: 5
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Hampton Ltd #31285-OPPORTUNITY'
    concentration_or_purity: 59.7%
  - material_name: HEK293T cells
    concentration_or_purity: "31 \xB5M"
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Robinson Ltd #27693-ONE'
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Scott-Schultz #47118-AWAY'
    concentration_or_purity: "31 \xB5M"
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Wilson-Vaughn Only7490
  - equipment_name: Confocal Microscope
    settings_parameters: "8277 x g, 5\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Moore, Hopkins and Boyd Cover3541
  - equipment_name: Shaking Incubator
  procedure_steps:
  - step_description: Cells were probed with ripa buffer to facilitate example.
    conditions_or_variables:
    - 100V constant voltage
    - at 80% confluency
    data_collected: true
    duration_minutes: 256
  - step_description: Cells were washed with sds-page loading buffer to facilitate
      operation.
    conditions_or_variables:
    - with protease inhibitors
    - rocking agitation
    data_collected: true
    temperature_celsius: 32
    replicates: 2
control_groups:
- control_type: Sham-operated Control
  description: Identify certainly card without sing picture item bar help like easy
    to bag.
data_analysis_methods:
- Mass spectrometry data processed with MaxQuant
- ImageJ densitometry
reproducibility_notes: All experiments were independently verified by Dr. Karen Howard
  and results were consistent across multiple biological replicates.
