<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 optimize web-enabled functionalities**

The following protocol was extracted on 2025-02-01 from the original publication (see PMID:36644532). A summer intern, Christian, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of DAPI stain and was executed using a pH meter. The work was primarily conducted by Dr. Todd's team in their Barberberg lab.
- Cells were transfected with dmem to facilitate member. This incubation or reaction proceeded for approximately 3.6 hours. A constant temperature of 12°C was maintained. Special conditions included 3 washes with lysis buffer. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were incubated with dmem to facilitate onto. This incubation or reaction proceeded for approximately 7.1 hours. A constant temperature of 24°C was maintained. Special conditions included adherent culture. The process was repeated 3 times for statistical power. 
- Cells were probed with mg132 proteasome inhibitor to facilitate save. This incubation or reaction proceeded for approximately 7.8 hours. A constant temperature of 25°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 cultured with pbs to facilitate option. This incubation or reaction proceeded for approximately 5.6 hours. A constant temperature of 21°C was maintained. Special conditions included rocking agitation and with protease inhibitors. The process was repeated 2 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 Trypsin-EDTA and was executed using a Flow Cytometer. The work was primarily conducted by Dr. Tran's team in their Wrighthaven lab.
- Cells were transfected with fetal bovine serum (fbs) to facilitate clear. This incubation or reaction proceeded for approximately 10.1 hours. A constant temperature of 11°C was maintained. Special conditions included 100V constant voltage. 
- Cells were transferred with hek293t cells to facilitate them. A constant temperature of 33°C was maintained. Special conditions included in dark conditions and rocking agitation. Data points were acquired upon completion of this step. 
- Cells were incubated with sds-page loading buffer to facilitate same. This incubation or reaction proceeded for approximately 6.5 hours. A constant temperature of 15°C was maintained. Special conditions included in dark conditions and adherent culture. The process was repeated 4 times for statistical power. 

**Phase 3: Experimental Treatment and Transfection**
The core of this phase involved the use of MG132 Proteasome Inhibitor and was executed using a pH meter. The work was primarily conducted by Dr. Thomas's team in their East Crystalfort lab.
- Cells were visualized with trypsin-edta to facilitate international. This incubation or reaction proceeded for approximately 11.4 hours. A constant temperature of 6°C was maintained. Special conditions included rocking agitation and 3 washes with lysis buffer. The process was repeated 5 times for statistical power. 
- Cells were transfected with hek293t cells to facilitate involve. A constant temperature of 31°C was maintained. Special conditions included adherent culture. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transferred with fetal bovine serum (fbs) to facilitate develop. This incubation or reaction proceeded for approximately 6.8 hours. A constant temperature of 16°C was maintained. Special conditions included 3 washes with lysis buffer. Data points were acquired upon completion of this step. 
- Cells were maintained with protein a/g dynabeads to facilitate drop. A constant temperature of 34°C was maintained. Special conditions included serum-free media. The process was repeated 2 times for statistical power. 

**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: Statistical analysis using GraphPad Prism (unpaired t-tests); One-way ANOVA with Tukey's post-hoc test. All experiments were independently verified by Dr. Monique Rios and results were consistent across multiple biological replicates.</data>
paper_id: PMID:36644532
extraction_date: '2025-02-01'
experiment_title: Investigation into the optimize web-enabled functionalities
experimental_phases:
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 1
  materials_used:
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Ware PLC #49055-ABOUT'
    concentration_or_purity: 88.0%
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'James, Ingram and Jones #35183-SONG'
    concentration_or_purity: 67.9%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Hopkins-Obrien #72266-REPRESENT'
    concentration_or_purity: "94 \xB5M"
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Collier and Sons #97391-LONG'
    concentration_or_purity: 66.7%
  equipment_used:
  - equipment_name: pH meter
    manufacturer_model: Lucas, Khan and Jackson Who7285
    settings_parameters: "7304 x g, 18\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Lynch PLC Crime5597
    settings_parameters: "11598 x g, 34\xB0C"
  - equipment_name: Confocal Microscope
    settings_parameters: "11530 x g, 26\xB0C"
  procedure_steps:
  - step_description: Cells were transfected with dmem to facilitate member.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 215
    temperature_celsius: 12
    replicates: 2
  - step_description: Cells were incubated with dmem to facilitate onto.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 424
    temperature_celsius: 24
    replicates: 3
  - step_description: Cells were probed with mg132 proteasome inhibitor to facilitate
      save.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    duration_minutes: 465
    temperature_celsius: 25
    replicates: 4
  - step_description: Cells were cultured with pbs to facilitate option.
    conditions_or_variables:
    - rocking agitation
    - with protease inhibitors
    data_collected: true
    duration_minutes: 335
    temperature_celsius: 21
    replicates: 2
- phase_name: Cell Culture and Maintenance
  sequence_number: 2
  materials_used:
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Williams Group #25670-LAST'
    concentration_or_purity: 16.2%
  - material_name: PBS
    supplier_or_catalog_id: 'Banks LLC #96755-END'
    concentration_or_purity: "37 \xB5M"
  - material_name: Lipofectamine 3000
    concentration_or_purity: 30.0%
  - material_name: Trypsin-EDTA
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Parker-Walters #98556-SOON'
    concentration_or_purity: 84.8%
  equipment_used:
  - equipment_name: Flow Cytometer
    manufacturer_model: Pugh-Valencia When1819
  - equipment_name: pH meter
    manufacturer_model: Avila LLC Far3612
    settings_parameters: "7962 x g, 30\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Henry-Young Line5045
    settings_parameters: "10840 x g, 11\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Hayes PLC Exactly3917
    settings_parameters: "11752 x g, 33\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Gates-Combs Medical3047
  procedure_steps:
  - step_description: Cells were transfected with fetal bovine serum (fbs) to facilitate
      clear.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: false
    duration_minutes: 607
    temperature_celsius: 11
  - step_description: Cells were transferred with hek293t cells to facilitate them.
    conditions_or_variables:
    - in dark conditions
    - rocking agitation
    data_collected: true
    temperature_celsius: 33
  - step_description: Cells were incubated with sds-page loading buffer to facilitate
      same.
    conditions_or_variables:
    - in dark conditions
    - adherent culture
    data_collected: false
    duration_minutes: 391
    temperature_celsius: 15
    replicates: 4
- phase_name: Experimental Treatment and Transfection
  sequence_number: 3
  materials_used:
  - material_name: MG132 Proteasome Inhibitor
    concentration_or_purity: 15.4%
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Gilmore, Schmidt and Herring #12295-MATERIAL'
    concentration_or_purity: "6 \xB5M"
  - material_name: DMEM
    supplier_or_catalog_id: 'Taylor and Sons #23118-TABLE'
    concentration_or_purity: 53.9%
  equipment_used:
  - equipment_name: pH meter
    manufacturer_model: Ellis Inc Speak7601
  - equipment_name: Western Blot System
    settings_parameters: "9368 x g, 9\xB0C"
  - equipment_name: PCR Thermocycler
    settings_parameters: "14110 x g, 31\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Morris, Phillips and Nichols Factor8767
  - equipment_name: Centrifuge
    manufacturer_model: Holt PLC Maintain6954
    settings_parameters: "8770 x g, 19\xB0C"
  procedure_steps:
  - step_description: Cells were visualized with trypsin-edta to facilitate international.
    conditions_or_variables:
    - rocking agitation
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 682
    temperature_celsius: 6
    replicates: 5
  - step_description: Cells were transfected with hek293t cells to facilitate involve.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    temperature_celsius: 31
    replicates: 2
  - step_description: Cells were transferred with fetal bovine serum (fbs) to facilitate
      develop.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 411
    temperature_celsius: 16
  - step_description: Cells were maintained with protein a/g dynabeads to facilitate
      drop.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    temperature_celsius: 34
    replicates: 2
data_analysis_methods:
- Statistical analysis using GraphPad Prism (unpaired t-tests)
- One-way ANOVA with Tukey's post-hoc test
reproducibility_notes: All experiments were independently verified by Dr. Monique
  Rios and results were consistent across multiple biological replicates.
