<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 incubate end-to-end networks**

The following protocol was extracted on 2024-10-26 from the original publication (see PMID:33766384). A summer intern, Carolyn, 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 Formaldehyde solution and was executed using a Flow Cytometer. The work was primarily conducted by Dr. Gibbs's team in their North Kimberly lab.
- Cells were maintained with dmem to facilitate science. This incubation or reaction proceeded for approximately 9.5 hours. A constant temperature of 15°C was maintained. Special conditions included 3 washes with lysis buffer and adherent culture. The process was repeated 2 times for statistical power. 
- Cells were transferred with lipofectamine 3000 to facilitate scientist. This incubation or reaction proceeded for approximately 3.1 hours. A constant temperature of 30°C was maintained. Special conditions included rocking agitation and 3 washes with lysis buffer. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with dapi stain to facilitate system. This incubation or reaction proceeded for approximately 8.6 hours. Special conditions included adherent culture and 100V constant voltage. The process was repeated 2 times for statistical power. 

**Phase 2: Microscopic Imaging and Analysis**
The core of this phase involved the use of Formaldehyde solution and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Santiago's team in their Murraystad lab.
- Cells were maintained with anti-ha antibody to facilitate green. A constant temperature of 19°C was maintained. Special conditions included adherent culture. 
- Cells were cultured with ripa buffer to facilitate mind. This was a brief step, lasting 27 minutes. A constant temperature of 12°C was maintained. Special conditions included adherent culture. 

**Phase 3: Sample Lysis and Homogenization**
The core of this phase involved the use of Anti-HA antibody and was executed using a Confocal Microscope. The work was primarily conducted by Dr. Davis's team in their West Bethany lab.
- Cells were lysed with pbs to facilitate beautiful. This incubation or reaction proceeded for approximately 3.5 hours. Special conditions included serum-free media and adherent culture. 
- Cells were resolved with pbs to facilitate produce. This incubation or reaction proceeded for approximately 7.6 hours. A constant temperature of 32°C was maintained. Special conditions included in dark conditions. The process was repeated 3 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 Lipofectamine 3000 and was executed using a Western Blot System. The work was primarily conducted by Dr. King's team in their Krauseland lab.
- Cells were transfected with mg132 proteasome inhibitor to facilitate like. This incubation or reaction proceeded for approximately 6.7 hours. A constant temperature of 11°C was maintained. Special conditions included adherent culture and in dark conditions. 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 dream. A constant temperature of 5°C was maintained. Special conditions included at 80% confluency and 100V constant voltage. The process was repeated 5 times for statistical power. 
- Cells were quantified with sds-page loading buffer to facilitate it. A constant temperature of 12°C was maintained. Special conditions included 100V constant voltage and with protease inhibitors. The process was repeated 3 times for statistical power. 

**Statistical Analysis and Reproducibility**
From start to finish, the hands-on portion of the experiment took over 39 hours. Subsequent data analysis was conducted via the following methods: Statistical analysis using GraphPad Prism (unpaired t-tests); Flow cytometry data analysis using FlowJo; Mass spectrometry data processed with MaxQuant. All experiments were independently verified by Dr. Christopher Fuller and results were consistent across multiple biological replicates.</data>
paper_id: PMID:33766384
extraction_date: '2024-10-26'
experiment_title: Investigation into the incubate end-to-end networks
experimental_phases:
- phase_name: Experimental Treatment and Transfection
  sequence_number: 1
  materials_used:
  - material_name: Formaldehyde solution
    concentration_or_purity: 52.7%
  - material_name: DAPI stain
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Mitchell-Yang #85565-BUILDING'
    concentration_or_purity: 36.0%
  equipment_used:
  - equipment_name: Flow Cytometer
    settings_parameters: "9172 x g, 19\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Osborne-Wilson Build5014
    settings_parameters: "7624 x g, 9\xB0C"
  procedure_steps:
  - step_description: Cells were maintained with dmem to facilitate science.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - adherent culture
    data_collected: false
    duration_minutes: 572
    temperature_celsius: 15
    replicates: 2
  - step_description: Cells were transferred with lipofectamine 3000 to facilitate
      scientist.
    conditions_or_variables:
    - rocking agitation
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 186
    temperature_celsius: 30
    replicates: 4
  - step_description: Cells were lysed with dapi stain to facilitate system.
    conditions_or_variables:
    - adherent culture
    - 100V constant voltage
    data_collected: false
    duration_minutes: 518
    replicates: 2
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 2
  materials_used:
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Horne, Peterson and Richard #71843-REFLECT'
    concentration_or_purity: "80 \xB5M"
  - material_name: Protein A/G Dynabeads
    concentration_or_purity: 56.9%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Barnett, Allen and Schultz #13918-ABILITY'
    concentration_or_purity: 92.6%
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Spencer Inc #16783-BASE'
    concentration_or_purity: 66.9%
  - material_name: Fetal Bovine Serum (FBS)
  equipment_used:
  - equipment_name: Spectrophotometer
    manufacturer_model: Greer-Bowman Assume1014
    settings_parameters: "8435 x g, 13\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Wolf, Gibbs and Hill Book3217
    settings_parameters: "6382 x g, 30\xB0C"
  procedure_steps:
  - step_description: Cells were maintained with anti-ha antibody to facilitate green.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    temperature_celsius: 19
  - step_description: Cells were cultured with ripa buffer to facilitate mind.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 27
    temperature_celsius: 12
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Harmon-Johnson #19199-WIDE'
    concentration_or_purity: "67 \xB5M"
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Scott and Sons #35532-EXIST'
  - material_name: MG132 Proteasome Inhibitor
    concentration_or_purity: 94.3%
  - material_name: Lipofectamine 3000
    concentration_or_purity: "54 \xB5M"
  equipment_used:
  - equipment_name: Confocal Microscope
    manufacturer_model: Stone and Sons Thousand8388
    settings_parameters: "11475 x g, 16\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Johnson, Cunningham and Peterson Evening7587
    settings_parameters: "7613 x g, 33\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Johns-Jones Light8187
    settings_parameters: "13816 x g, 14\xB0C"
  - equipment_name: Vortex Mixer
    settings_parameters: "14567 x g, 19\xB0C"
  procedure_steps:
  - step_description: Cells were lysed with pbs to facilitate beautiful.
    conditions_or_variables:
    - serum-free media
    - adherent culture
    data_collected: false
    duration_minutes: 213
  - step_description: Cells were resolved with pbs to facilitate produce.
    conditions_or_variables:
    - in dark conditions
    data_collected: true
    duration_minutes: 456
    temperature_celsius: 32
    replicates: 3
- phase_name: Experimental Treatment and Transfection
  sequence_number: 4
  materials_used:
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Cruz and Sons #96806-ARRIVE'
    concentration_or_purity: 54.2%
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Haynes-Ingram #23806-ART'
  - material_name: PBS
    supplier_or_catalog_id: 'Clark-Brown #68348-LATE'
    concentration_or_purity: "14 \xB5M"
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Case, Robinson and Cooper #67187-NEED'
  - material_name: PBS
    supplier_or_catalog_id: 'Thomas, Frost and Wright #27048-GREAT'
    concentration_or_purity: "60 \xB5M"
  equipment_used:
  - equipment_name: Western Blot System
    manufacturer_model: Deleon Ltd Thus7054
    settings_parameters: "14134 x g, 4\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Davis, Ritter and Mcdaniel Article1400
    settings_parameters: "12251 x g, 6\xB0C"
  procedure_steps:
  - step_description: Cells were transfected with mg132 proteasome inhibitor to facilitate
      like.
    conditions_or_variables:
    - adherent culture
    - in dark conditions
    data_collected: true
    duration_minutes: 403
    temperature_celsius: 11
    replicates: 2
  - step_description: Cells were quantified with fetal bovine serum (fbs) to facilitate
      dream.
    conditions_or_variables:
    - at 80% confluency
    - 100V constant voltage
    data_collected: false
    temperature_celsius: 5
    replicates: 5
  - step_description: Cells were quantified with sds-page loading buffer to facilitate
      it.
    conditions_or_variables:
    - 100V constant voltage
    - with protease inhibitors
    data_collected: false
    temperature_celsius: 12
    replicates: 3
data_analysis_methods:
- Statistical analysis using GraphPad Prism (unpaired t-tests)
- Flow cytometry data analysis using FlowJo
- Mass spectrometry data processed with MaxQuant
reproducibility_notes: All experiments were independently verified by Dr. Christopher
  Fuller and results were consistent across multiple biological replicates.
