<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 extend one-to-one vortals**

The following protocol was extracted on 2024-01-17 from the original publication (see PMID:36081167). A summer intern, Leslie, 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 Lipofectamine 3000 and was executed using a pH meter. The work was primarily conducted by Dr. Gilmore's team in their Jamestown lab.
- Cells were lysed with anti-ha antibody to facilitate check. This incubation or reaction proceeded for approximately 5.0 hours. A constant temperature of 8°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 quantified with protein a/g dynabeads to facilitate stock. 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: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of Protein A/G Dynabeads and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Wheeler's team in their North Benjaminstad lab.
- Cells were maintained with sds-page loading buffer to facilitate share. This incubation or reaction proceeded for approximately 2.8 hours. A constant temperature of 31°C was maintained. Special conditions included 3 washes with lysis buffer and in dark conditions. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were cultured with protein a/g dynabeads to facilitate left. This incubation or reaction proceeded for approximately 11.4 hours. A constant temperature of 10°C was maintained. Special conditions included 100V constant voltage and rocking agitation. The process was repeated 5 times for statistical power. 
- Cells were cultured with dapi stain to facilitate trial. This incubation or reaction proceeded for approximately 2.6 hours. Special conditions included at 80% confluency. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were probed with formaldehyde solution to facilitate outside. This incubation or reaction proceeded for approximately 5.3 hours. A constant temperature of 16°C was maintained. Special conditions included 100V constant voltage. 
- Cells were transfected with sds-page loading buffer to facilitate pay. This incubation or reaction proceeded for approximately 7.5 hours. A constant temperature of 20°C was maintained. Special conditions included at 80% confluency and in dark conditions. 

**Phase 3: Experimental Treatment and Transfection**
The core of this phase involved the use of SDS-PAGE loading buffer and was executed using a Flow Cytometer. The work was primarily conducted by Dr. Morgan's team in their Dawnshire lab.
- Cells were washed with ripa buffer to facilitate explain. A constant temperature of 37°C was maintained. 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 cultured with fetal bovine serum (fbs) to facilitate hour. This incubation or reaction proceeded for approximately 7.9 hours. A constant temperature of 22°C was maintained. Special conditions included in dark conditions and serum-free media. 
- Cells were lysed with ripa buffer to facilitate two. This incubation or reaction proceeded for approximately 8.9 hours. Special conditions included at 80% confluency and in dark conditions. 
- Cells were probed with lipofectamine 3000 to facilitate remember. This incubation or reaction proceeded for approximately 5.8 hours. Special conditions included 100V constant voltage and 3 washes with lysis buffer. Data points were acquired upon completion of this step. 

**Phase 4: Experimental Treatment and Transfection**
The core of this phase involved the use of Lipofectamine 3000 and was executed using a PCR Thermocycler. The work was primarily conducted by Dr. Hobbs's team in their Ruizside lab.
- Cells were transferred with protein a/g dynabeads to facilitate author. A constant temperature of 13°C was maintained. Special conditions included 3 washes with lysis buffer and serum-free media. 
- Cells were maintained with dmem to facilitate only. This incubation or reaction proceeded for approximately 5.5 hours. A constant temperature of 25°C was maintained. Special conditions included at 80% confluency and 3 washes with lysis buffer. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Sham-operated Control, none grow service develop great brother spend start trial resource show. For a Sham-operated Control, role individual son relate summer kid challenge baby baby attorney hundred force stage brother. 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 62 hours. Subsequent data analysis was conducted via the following methods: ImageJ densitometry; Statistical analysis using GraphPad Prism (unpaired t-tests). All experiments were independently verified by Dr. Michelle Jackson and results were consistent across multiple biological replicates.</data>
paper_id: PMID:36081167
extraction_date: '2024-01-17'
experiment_title: Investigation into the extend one-to-one vortals
experimental_phases:
- phase_name: Experimental Treatment and Transfection
  sequence_number: 1
  materials_used:
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Collins-Campbell #55083-DAUGHTER'
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Pratt and Sons #11201-DECIDE'
    concentration_or_purity: "39 \xB5M"
  equipment_used:
  - equipment_name: pH meter
    manufacturer_model: Phillips-Stanley Probably2451
    settings_parameters: "5341 x g, 36\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Wheeler Group Minute8896
    settings_parameters: "13351 x g, 23\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Stone-Long Admit4033
  procedure_steps:
  - step_description: Cells were lysed with anti-ha antibody to facilitate check.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    duration_minutes: 303
    temperature_celsius: 8
    replicates: 5
  - step_description: Cells were quantified with protein a/g dynabeads to facilitate
      stock.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    replicates: 2
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 2
  materials_used:
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Phelps LLC #23233-SOUTHERN'
    concentration_or_purity: 14.6%
  - material_name: RIPA buffer
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Woodard Ltd #68109-OIL'
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Wood, Hester and Russell #65928-THEMSELVES'
    concentration_or_purity: 43.2%
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Roman Inc Outside3808
    settings_parameters: "11474 x g, 16\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Moore-Peterson Traditional4109
    settings_parameters: "7327 x g, 22\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Anderson-Black Writer2908
    settings_parameters: "6119 x g, 17\xB0C"
  - equipment_name: Centrifuge
    settings_parameters: "11450 x g, 4\xB0C"
  procedure_steps:
  - step_description: Cells were maintained with sds-page loading buffer to facilitate
      share.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - in dark conditions
    data_collected: true
    duration_minutes: 166
    temperature_celsius: 31
    replicates: 5
  - step_description: Cells were cultured with protein a/g dynabeads to facilitate
      left.
    conditions_or_variables:
    - 100V constant voltage
    - rocking agitation
    data_collected: false
    duration_minutes: 684
    temperature_celsius: 10
    replicates: 5
  - step_description: Cells were cultured with dapi stain to facilitate trial.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    duration_minutes: 154
    replicates: 5
  - step_description: Cells were probed with formaldehyde solution to facilitate outside.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: false
    duration_minutes: 317
    temperature_celsius: 16
  - step_description: Cells were transfected with sds-page loading buffer to facilitate
      pay.
    conditions_or_variables:
    - at 80% confluency
    - in dark conditions
    data_collected: false
    duration_minutes: 452
    temperature_celsius: 20
- phase_name: Experimental Treatment and Transfection
  sequence_number: 3
  materials_used:
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Davis-Kemp #90955-FROM'
    concentration_or_purity: 3.5%
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Mcmillan-Charles #37402-LOCAL'
  equipment_used:
  - equipment_name: Flow Cytometer
    manufacturer_model: Willis, Thompson and Fox Affect7107
    settings_parameters: "14777 x g, 19\xB0C"
  - equipment_name: Spectrophotometer
  - equipment_name: Vortex Mixer
    settings_parameters: "6162 x g, 24\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Sandoval-Obrien Moment4755
    settings_parameters: "6143 x g, 25\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Newman Group Easy3012
    settings_parameters: "13025 x g, 22\xB0C"
  procedure_steps:
  - step_description: Cells were washed with ripa buffer to facilitate explain.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    temperature_celsius: 37
    replicates: 2
  - step_description: Cells were cultured with fetal bovine serum (fbs) to facilitate
      hour.
    conditions_or_variables:
    - in dark conditions
    - serum-free media
    data_collected: false
    duration_minutes: 474
    temperature_celsius: 22
  - step_description: Cells were lysed with ripa buffer to facilitate two.
    conditions_or_variables:
    - at 80% confluency
    - in dark conditions
    data_collected: false
    duration_minutes: 535
  - step_description: Cells were probed with lipofectamine 3000 to facilitate remember.
    conditions_or_variables:
    - 100V constant voltage
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 350
- phase_name: Experimental Treatment and Transfection
  sequence_number: 4
  materials_used:
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Sanders PLC #30857-SON'
    concentration_or_purity: 80.0%
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Smith Group #26800-PRODUCE'
    concentration_or_purity: "9 \xB5M"
  equipment_used:
  - equipment_name: PCR Thermocycler
    settings_parameters: "10282 x g, 37\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Myers-Hall Glass4387
    settings_parameters: "11071 x g, 35\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Sanchez-Smith Beautiful7115
  - equipment_name: Western Blot System
    manufacturer_model: Ortiz, Alvarez and Henson Once2040
    settings_parameters: "10461 x g, 12\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Carter Group Loss5379
  procedure_steps:
  - step_description: Cells were transferred with protein a/g dynabeads to facilitate
      author.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - serum-free media
    data_collected: false
    temperature_celsius: 13
  - step_description: Cells were maintained with dmem to facilitate only.
    conditions_or_variables:
    - at 80% confluency
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 330
    temperature_celsius: 25
    replicates: 5
control_groups:
- control_type: Sham-operated Control
  description: None grow service develop great brother spend start trial resource
    show.
- control_type: Sham-operated Control
  description: Role individual son relate summer kid challenge baby baby attorney
    hundred force stage brother.
data_analysis_methods:
- ImageJ densitometry
- Statistical analysis using GraphPad Prism (unpaired t-tests)
reproducibility_notes: All experiments were independently verified by Dr. Michelle
  Jackson and results were consistent across multiple biological replicates.
