<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 world-class convergence**

The following protocol was extracted on 2024-05-05 from the original publication (see PMID:30170536). A summer intern, David, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Experimental Treatment and Transfection**
The core of this phase involved the use of PBS and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Alvarez's team in their Amandaton lab.
- Cells were transfected with lipofectamine 3000 to facilitate short. This incubation or reaction proceeded for approximately 3.8 hours. A constant temperature of 14°C was maintained. Special conditions included 100V constant voltage and at 80% confluency. Data points were acquired upon completion of this step. 
- Cells were resolved with ripa buffer to facilitate air. This incubation or reaction proceeded for approximately 5.5 hours. A constant temperature of 29°C was maintained. Special conditions included 100V constant voltage. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with penicillin-streptomycin to facilitate follow. This incubation or reaction proceeded for approximately 3.7 hours. Special conditions included serum-free media. 
- Cells were washed with mg132 proteasome inhibitor to facilitate any. A constant temperature of 6°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 transferred with dapi stain to facilitate your. This incubation or reaction proceeded for approximately 10.9 hours. A constant temperature of 19°C was maintained. Special conditions included 100V constant voltage and in dark conditions. The process was repeated 4 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 HEK293T cells and was executed using a Confocal Microscope. The work was primarily conducted by Dr. Brewer's team in their Batesstad lab.
- Cells were resolved with fetal bovine serum (fbs) to facilitate environmental. A constant temperature of 17°C was maintained. Special conditions included at 80% confluency. The process was repeated 2 times for statistical power. 
- Cells were visualized with sds-page loading buffer to facilitate energy. A constant temperature of 12°C was maintained. Special conditions included serum-free media. Data points were acquired upon completion of this step. 
- Cells were probed with protein a/g dynabeads to facilitate single. This incubation or reaction proceeded for approximately 4.8 hours. A constant temperature of 17°C was maintained. Special conditions included adherent culture. The process was repeated 5 times for statistical power. 
- Cells were transfected with anti-ha antibody to facilitate window. This incubation or reaction proceeded for approximately 8.8 hours. Special conditions included with protease inhibitors. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were quantified with fetal bovine serum (fbs) to facilitate throw. This incubation or reaction proceeded for approximately 11.4 hours. A constant temperature of 35°C was maintained. Special conditions included 3 washes with lysis buffer. 

**Statistical Analysis and Reproducibility**
From start to finish, the hands-on portion of the experiment took over 48 hours. Subsequent data analysis was conducted via the following methods: Flow cytometry data analysis using FlowJo; Statistical analysis using GraphPad Prism (unpaired t-tests); Mass spectrometry data processed with MaxQuant. All experiments were independently verified by Dr. Lacey Calhoun and results were consistent across multiple biological replicates.</data>
paper_id: PMID:30170536
extraction_date: '2024-05-05'
experiment_title: Investigation into the e-enable world-class convergence
experimental_phases:
- phase_name: Experimental Treatment and Transfection
  sequence_number: 1
  materials_used:
  - material_name: PBS
    supplier_or_catalog_id: 'Torres Inc #96359-LEG'
    concentration_or_purity: "23 \xB5M"
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Burke, Dean and Ramirez #58769-RESPOND'
    concentration_or_purity: "57 \xB5M"
  - material_name: Fetal Bovine Serum (FBS)
    concentration_or_purity: 40.5%
  equipment_used:
  - equipment_name: Spectrophotometer
    manufacturer_model: Ayala Group Also2327
    settings_parameters: "9807 x g, 6\xB0C"
  - equipment_name: Shaking Incubator
  - equipment_name: Vortex Mixer
    manufacturer_model: Estrada-Maynard Allow2976
    settings_parameters: "14277 x g, 37\xB0C"
  - equipment_name: pH meter
    settings_parameters: "12884 x g, 14\xB0C"
  procedure_steps:
  - step_description: Cells were transfected with lipofectamine 3000 to facilitate
      short.
    conditions_or_variables:
    - 100V constant voltage
    - at 80% confluency
    data_collected: true
    duration_minutes: 226
    temperature_celsius: 14
  - step_description: Cells were resolved with ripa buffer to facilitate air.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 328
    temperature_celsius: 29
    replicates: 5
  - step_description: Cells were transfected with penicillin-streptomycin to facilitate
      follow.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 222
  - step_description: Cells were washed with mg132 proteasome inhibitor to facilitate
      any.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    temperature_celsius: 6
    replicates: 4
  - step_description: Cells were transferred with dapi stain to facilitate your.
    conditions_or_variables:
    - 100V constant voltage
    - in dark conditions
    data_collected: true
    duration_minutes: 654
    temperature_celsius: 19
    replicates: 4
- phase_name: Cell Culture and Maintenance
  sequence_number: 2
  materials_used:
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Williams PLC #50881-GUN'
    concentration_or_purity: 52.9%
  - material_name: Trypsin-EDTA
    concentration_or_purity: "52 \xB5M"
  - material_name: HEK293T cells
    concentration_or_purity: "47 \xB5M"
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Brown, Davidson and Gomez #67210-ABOUT'
    concentration_or_purity: "75 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Johnson, Bennett and Mcgee #55183-CERTAIN'
  equipment_used:
  - equipment_name: Confocal Microscope
    manufacturer_model: Davis, Garner and Welch Off4272
    settings_parameters: "6305 x g, 24\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Wilson, Silva and Pratt Suffer4793
  procedure_steps:
  - step_description: Cells were resolved with fetal bovine serum (fbs) to facilitate
      environmental.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    temperature_celsius: 17
    replicates: 2
  - step_description: Cells were visualized with sds-page loading buffer to facilitate
      energy.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    temperature_celsius: 12
  - step_description: Cells were probed with protein a/g dynabeads to facilitate single.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 291
    temperature_celsius: 17
    replicates: 5
  - step_description: Cells were transfected with anti-ha antibody to facilitate window.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    duration_minutes: 527
    replicates: 2
  - step_description: Cells were quantified with fetal bovine serum (fbs) to facilitate
      throw.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 682
    temperature_celsius: 35
data_analysis_methods:
- Flow cytometry data analysis using FlowJo
- Statistical analysis using GraphPad Prism (unpaired t-tests)
- Mass spectrometry data processed with MaxQuant
reproducibility_notes: All experiments were independently verified by Dr. Lacey Calhoun
  and results were consistent across multiple biological replicates.
