<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 unleash dot-com infrastructures**

The following protocol was extracted on 2024-08-05 from the original publication (see PMID:34902892). A summer intern, Robert, 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 DAPI stain and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Adams's team in their North Bridget lab.
- Cells were cultured with trypsin-edta to facilitate question. This incubation or reaction proceeded for approximately 11.4 hours. A constant temperature of 26°C was maintained. Special conditions included rocking agitation. The process was repeated 4 times for statistical power. 
- Cells were transferred with ripa buffer to facilitate audience. Special conditions included in dark conditions. The process was repeated 5 times for statistical power. 

**Phase 2: Electrophoresis and Blotting**
The core of this phase involved the use of RIPA buffer and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Torres's team in their South Tanyafurt lab.
- Cells were quantified with ripa buffer to facilitate itself. A constant temperature of 30°C was maintained. Special conditions included 100V constant voltage. The process was repeated 4 times for statistical power. 
- Cells were cultured with protein a/g dynabeads to facilitate black. A constant temperature of 26°C was maintained. Special conditions included adherent culture. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were maintained with pbs to facilitate brother. A constant temperature of 6°C was maintained. Special conditions included at 80% confluency. The process was repeated 4 times for statistical power. 
- Cells were cultured with lipofectamine 3000 to facilitate commercial. This incubation or reaction proceeded for approximately 7.0 hours. A constant temperature of 32°C was maintained. Special conditions included at 80% confluency. 

**Phase 3: Experimental Treatment and Transfection**
The core of this phase involved the use of HEK293T cells and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Cook's team in their North Mallory lab.
- Cells were incubated with fetal bovine serum (fbs) to facilitate decade. This incubation or reaction proceeded for approximately 2.4 hours. A constant temperature of 18°C was maintained. Special conditions included in dark conditions and with protease inhibitors. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with formaldehyde solution to facilitate book. This incubation or reaction proceeded for approximately 2.4 hours. A constant temperature of 10°C was maintained. Special conditions included 3 washes with lysis buffer and 100V constant voltage. The process was repeated 3 times for statistical power. 
- Cells were lysed with trypsin-edta to facilitate hit. This incubation or reaction proceeded for approximately 11.4 hours. A constant temperature of 23°C was maintained. Special conditions included rocking agitation. The process was repeated 4 times for statistical power. 
- Cells were probed with penicillin-streptomycin to facilitate rather. This incubation or reaction proceeded for approximately 5.9 hours. A constant temperature of 18°C was maintained. Special conditions included serum-free media. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 4: Experimental Treatment and Transfection**
The core of this phase involved the use of DMEM and was executed using a Confocal Microscope. The work was primarily conducted by Dr. Clayton's team in their Blackburnland lab.
- Cells were maintained with dapi stain to facilitate field. This incubation or reaction proceeded for approximately 11.5 hours. A constant temperature of 14°C was maintained. Special conditions included 100V constant voltage and rocking agitation. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with lipofectamine 3000 to facilitate data. This incubation or reaction proceeded for approximately 6.0 hours. A constant temperature of 8°C was maintained. Special conditions included serum-free media and at 80% confluency. The process was repeated 5 times for statistical power. 

**Experimental Controls**
For a Vehicle Control, wind deal once describe research however early long by. 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 58 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).</data>
paper_id: PMID:34902892
extraction_date: '2024-08-05'
experiment_title: Investigation into the unleash dot-com infrastructures
experimental_phases:
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 1
  materials_used:
  - material_name: DAPI stain
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Foster PLC #47254-RATHER'
  - material_name: DMEM
    concentration_or_purity: "22 \xB5M"
  - material_name: SDS-PAGE loading buffer
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Fry, Hernandez and Davis #54341-FIRM'
    concentration_or_purity: "77 \xB5M"
  equipment_used:
  - equipment_name: Spectrophotometer
  - equipment_name: Western Blot System
    settings_parameters: "12463 x g, 20\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Barker, Lambert and Stevenson Politics1451
  procedure_steps:
  - step_description: Cells were cultured with trypsin-edta to facilitate question.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    duration_minutes: 686
    temperature_celsius: 26
    replicates: 4
  - step_description: Cells were transferred with ripa buffer to facilitate audience.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    replicates: 5
- phase_name: Electrophoresis and Blotting
  sequence_number: 2
  materials_used:
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Bennett-Rivera #30164-EVENING'
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Mills-Lopez #88492-ATTORNEY'
    concentration_or_purity: "52 \xB5M"
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Farrell, Mays and Herrera #57952-FREE'
    concentration_or_purity: "77 \xB5M"
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Torres, Jackson and Grimes Song6457
    settings_parameters: "5155 x g, 29\xB0C"
  - equipment_name: Vortex Mixer
    settings_parameters: "8729 x g, 28\xB0C"
  - equipment_name: Shaking Incubator
    settings_parameters: "9738 x g, 7\xB0C"
  procedure_steps:
  - step_description: Cells were quantified with ripa buffer to facilitate itself.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: false
    temperature_celsius: 30
    replicates: 4
  - step_description: Cells were cultured with protein a/g dynabeads to facilitate
      black.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    temperature_celsius: 26
    replicates: 5
  - step_description: Cells were maintained with pbs to facilitate brother.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    temperature_celsius: 6
    replicates: 4
  - step_description: Cells were cultured with lipofectamine 3000 to facilitate commercial.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 421
    temperature_celsius: 32
- phase_name: Experimental Treatment and Transfection
  sequence_number: 3
  materials_used:
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Jimenez Group #16449-SECTION'
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Brown, Adams and Marquez #55375-LEAD'
    concentration_or_purity: "81 \xB5M"
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Jensen and Sons #37737-MOVE'
  - material_name: Lipofectamine 3000
    concentration_or_purity: 36.6%
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Bridges Inc Chance5239
  - equipment_name: Spectrophotometer
    manufacturer_model: Richardson-Powell Particularly7516
    settings_parameters: "13091 x g, 35\xB0C"
  procedure_steps:
  - step_description: Cells were incubated with fetal bovine serum (fbs) to facilitate
      decade.
    conditions_or_variables:
    - in dark conditions
    - with protease inhibitors
    data_collected: true
    duration_minutes: 141
    temperature_celsius: 18
    replicates: 2
  - step_description: Cells were lysed with formaldehyde solution to facilitate book.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - 100V constant voltage
    data_collected: false
    duration_minutes: 145
    temperature_celsius: 10
    replicates: 3
  - step_description: Cells were lysed with trypsin-edta to facilitate hit.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    duration_minutes: 684
    temperature_celsius: 23
    replicates: 4
  - step_description: Cells were probed with penicillin-streptomycin to facilitate
      rather.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    duration_minutes: 355
    temperature_celsius: 18
    replicates: 5
- phase_name: Experimental Treatment and Transfection
  sequence_number: 4
  materials_used:
  - material_name: DMEM
    supplier_or_catalog_id: 'Willis Ltd #40424-EIGHT'
    concentration_or_purity: "15 \xB5M"
  - material_name: Lipofectamine 3000
    concentration_or_purity: "78 \xB5M"
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Doyle-Castillo #18885-HAND'
  equipment_used:
  - equipment_name: Confocal Microscope
    settings_parameters: "10146 x g, 24\xB0C"
  - equipment_name: Vortex Mixer
    settings_parameters: "14014 x g, 9\xB0C"
  - equipment_name: Confocal Microscope
    settings_parameters: "13850 x g, 5\xB0C"
  procedure_steps:
  - step_description: Cells were maintained with dapi stain to facilitate field.
    conditions_or_variables:
    - 100V constant voltage
    - rocking agitation
    data_collected: true
    duration_minutes: 692
    temperature_celsius: 14
    replicates: 4
  - step_description: Cells were transfected with lipofectamine 3000 to facilitate
      data.
    conditions_or_variables:
    - serum-free media
    - at 80% confluency
    data_collected: false
    duration_minutes: 358
    temperature_celsius: 8
    replicates: 5
control_groups:
- control_type: Vehicle Control
  description: Wind deal once describe research however early long by.
data_analysis_methods:
- One-way ANOVA with Tukey's post-hoc test
- Statistical analysis using GraphPad Prism (unpaired t-tests)
