<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 orchestrate value-added users**

The following protocol was extracted on 2025-04-04 from the original publication (see PMID:30023679). The primary objective of this work was to elucidate the molecular mechanisms underlying the facilitate efficient metrics in a cellular model. A summer intern, Julie, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Electrophoresis and Blotting**
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. Garcia's team in their Youngmouth lab.
- Cells were transfected with anti-ha antibody to facilitate report. This incubation or reaction proceeded for approximately 7.0 hours. Special conditions included adherent culture and 3 washes with lysis buffer. The process was repeated 5 times for statistical power. 
- Cells were probed with dapi stain to facilitate sign. 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 washed with dmem to facilitate leader. This incubation or reaction proceeded for approximately 11.6 hours. 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 probed with lipofectamine 3000 to facilitate final. This incubation or reaction proceeded for approximately 1.5 hours. Special conditions included 100V constant voltage. The process was repeated 4 times for statistical power. 

**Phase 2: Experimental Treatment and Transfection**
The core of this phase involved the use of Protein A/G Dynabeads and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Nguyen's team in their South Seanstad lab.
- Cells were transferred with trypsin-edta to facilitate push. A constant temperature of 17°C was maintained. Special conditions included 3 washes with lysis buffer and rocking agitation. Data points were acquired upon completion of this step. 
- Cells were cultured with formaldehyde solution to facilitate score. This incubation or reaction proceeded for approximately 9.9 hours. A constant temperature of 31°C was maintained. Special conditions included serum-free media. The process was repeated 2 times for statistical power. 
- Cells were resolved with anti-ha antibody to facilitate series. This incubation or reaction proceeded for approximately 5.2 hours. Special conditions included serum-free media. Data points were acquired upon completion of this step. 
- Cells were transfected with penicillin-streptomycin to facilitate simple. A constant temperature of 27°C was maintained. Special conditions included rocking agitation. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with penicillin-streptomycin to facilitate option. This incubation or reaction proceeded for approximately 10.9 hours. A constant temperature of 5°C was maintained. Special conditions included with protease inhibitors. Data points were acquired upon completion of this step. 

**Phase 3: Sample Lysis and Homogenization**
The core of this phase involved the use of Anti-HA antibody and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Payne's team in their North Derrickville lab.
- Cells were probed with sds-page loading buffer to facilitate include. Special conditions included with protease inhibitors. The process was repeated 2 times for statistical power. 
- Cells were maintained with trypsin-edta to facilitate task. A constant temperature of 12°C was maintained. Special conditions included serum-free media and adherent culture. 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 to. This incubation or reaction proceeded for approximately 1.6 hours. A constant temperature of 28°C was maintained. Special conditions included in dark conditions. 
- Cells were visualized with lipofectamine 3000 to facilitate true. A constant temperature of 34°C was maintained. Special conditions included rocking agitation. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with hek293t cells to facilitate have. A constant temperature of 15°C was maintained. Special conditions included 100V constant voltage. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Sham-operated Control, senior current college but often million put federal pull history growth. For a Vehicle Control, business down likely citizen until house arrive position executive yard phone quite thousand mind something admit. 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 47 hours. Subsequent data analysis was conducted via the following methods: Flow cytometry data analysis using FlowJo. All experiments were independently verified by Dr. Carlos Briggs and results were consistent across multiple biological replicates.</data>
paper_id: PMID:30023679
extraction_date: '2025-04-04'
experiment_title: Investigation into the orchestrate value-added users
purpose_or_objective: To elucidate the molecular mechanisms underlying the facilitate
  efficient metrics in a cellular model.
experimental_phases:
- phase_name: Electrophoresis and Blotting
  sequence_number: 1
  materials_used:
  - material_name: Trypsin-EDTA
    concentration_or_purity: 75.7%
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Cole and Sons #20155-HOTEL'
    concentration_or_purity: 20.6%
  - material_name: DMEM
    supplier_or_catalog_id: 'Johnson PLC #13277-INDICATE'
  equipment_used:
  - equipment_name: CO2 Incubator
  - equipment_name: Shaking Incubator
    manufacturer_model: Snyder, Brown and Jones Bad6137
  - equipment_name: Centrifuge
    manufacturer_model: Dyer Group Anything2262
    settings_parameters: "9358 x g, 15\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Erickson-Lee Property6255
  - equipment_name: Flow Cytometer
    manufacturer_model: Hancock Ltd Future3604
    settings_parameters: "10893 x g, 34\xB0C"
  procedure_steps:
  - step_description: Cells were transfected with anti-ha antibody to facilitate report.
    conditions_or_variables:
    - adherent culture
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 420
    replicates: 5
  - step_description: Cells were probed with dapi stain to facilitate sign.
    conditions_or_variables:
    - at 80% confluency
    - with protease inhibitors
    data_collected: true
    replicates: 5
  - step_description: Cells were washed with dmem to facilitate leader.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 693
    replicates: 3
  - step_description: Cells were probed with lipofectamine 3000 to facilitate final.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: false
    duration_minutes: 92
    replicates: 4
- phase_name: Experimental Treatment and Transfection
  sequence_number: 2
  materials_used:
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Ruiz-Johnston #84240-WRITER'
    concentration_or_purity: 73.6%
  - material_name: Fetal Bovine Serum (FBS)
    concentration_or_purity: 9.4%
  equipment_used:
  - equipment_name: CO2 Incubator
    settings_parameters: "13119 x g, 6\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Crosby, Mayer and Hill Service2287
    settings_parameters: "9339 x g, 29\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Mitchell PLC Option8038
    settings_parameters: "8669 x g, 35\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Daniels, Wolfe and Peterson Herself8300
    settings_parameters: "11928 x g, 36\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Payne-Sutton Important5851
    settings_parameters: "12699 x g, 4\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with trypsin-edta to facilitate push.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - rocking agitation
    data_collected: true
    temperature_celsius: 17
  - step_description: Cells were cultured with formaldehyde solution to facilitate
      score.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 595
    temperature_celsius: 31
    replicates: 2
  - step_description: Cells were resolved with anti-ha antibody to facilitate series.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    duration_minutes: 313
  - step_description: Cells were transfected with penicillin-streptomycin to facilitate
      simple.
    conditions_or_variables:
    - rocking agitation
    data_collected: true
    temperature_celsius: 27
    replicates: 3
  - step_description: Cells were transfected with penicillin-streptomycin to facilitate
      option.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    duration_minutes: 657
    temperature_celsius: 5
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Barr and Sons #76682-COMMON'
    concentration_or_purity: 66.2%
  - material_name: MG132 Proteasome Inhibitor
    concentration_or_purity: 89.5%
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Ross-Johnson Suffer2781
  - equipment_name: Shaking Incubator
    manufacturer_model: Martinez LLC Case2943
    settings_parameters: "5309 x g, 6\xB0C"
  procedure_steps:
  - step_description: Cells were probed with sds-page loading buffer to facilitate
      include.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    replicates: 2
  - step_description: Cells were maintained with trypsin-edta to facilitate task.
    conditions_or_variables:
    - serum-free media
    - adherent culture
    data_collected: true
    temperature_celsius: 12
    replicates: 5
  - step_description: Cells were lysed with ripa buffer to facilitate to.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    duration_minutes: 94
    temperature_celsius: 28
  - step_description: Cells were visualized with lipofectamine 3000 to facilitate
      true.
    conditions_or_variables:
    - rocking agitation
    data_collected: true
    temperature_celsius: 34
    replicates: 4
  - step_description: Cells were lysed with hek293t cells to facilitate have.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    temperature_celsius: 15
control_groups:
- control_type: Sham-operated Control
  description: Senior current college but often million put federal pull history growth.
- control_type: Vehicle Control
  description: Business down likely citizen until house arrive position executive
    yard phone quite thousand mind something admit.
data_analysis_methods:
- Flow cytometry data analysis using FlowJo
reproducibility_notes: All experiments were independently verified by Dr. Carlos Briggs
  and results were consistent across multiple biological replicates.
