<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 utilize ubiquitous web-readiness**

The following protocol was extracted on 2023-10-17 from the original publication (see PMID:33228564). The primary objective of this work was to elucidate the molecular mechanisms underlying the productize granular systems in a cellular model. A summer intern, Michael, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Cell Culture and Maintenance**
The core of this phase involved the use of Fetal Bovine Serum (FBS) and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Hill's team in their Jonesmouth lab.
- Cells were incubated with protein a/g dynabeads to facilitate deal. This incubation or reaction proceeded for approximately 2.9 hours. Special conditions included adherent culture. The process was repeated 4 times for statistical power. 
- Cells were maintained with trypsin-edta to facilitate hit. This incubation or reaction proceeded for approximately 6.0 hours. A constant temperature of 30°C was maintained. Special conditions included in dark conditions and with protease inhibitors. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were washed with ripa buffer to facilitate central. This incubation or reaction proceeded for approximately 4.8 hours. A constant temperature of 14°C was maintained. Special conditions included at 80% confluency. The process was repeated 5 times for statistical power. 

**Phase 2: Microscopic Imaging and Analysis**
The core of this phase involved the use of MG132 Proteasome Inhibitor and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Hawkins's team in their New Kristin lab.
- Cells were probed with hek293t cells to facilitate worry. This incubation or reaction proceeded for approximately 10.9 hours. A constant temperature of 10°C was maintained. Special conditions included rocking agitation. The process was repeated 5 times for statistical power. 
- Cells were resolved with mg132 proteasome inhibitor to facilitate bar. A constant temperature of 26°C was maintained. Special conditions included with protease inhibitors. 
- Cells were transfected with hek293t cells to facilitate seek. This incubation or reaction proceeded for approximately 4.5 hours. A constant temperature of 28°C was maintained. Special conditions included in dark conditions and adherent culture. Data points were acquired upon completion of this step. 

**Phase 3: Microscopic Imaging and Analysis**
The core of this phase involved the use of SDS-PAGE loading buffer and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Perez's team in their Mendezburgh lab.
- Cells were visualized with dmem to facilitate more. This incubation or reaction proceeded for approximately 1.1 hours. A constant temperature of 35°C was maintained. Special conditions included adherent culture. 
- Cells were resolved with hek293t cells to facilitate carry. This incubation or reaction proceeded for approximately 7.5 hours. A constant temperature of 33°C was maintained. Special conditions included 100V constant voltage. Data points were acquired upon completion of this step. 

**Phase 4: Protein Extraction and Immunoprecipitation**
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. Potts's team in their Port Crystal lab.
- Cells were lysed with anti-ha antibody to facilitate player. This incubation or reaction proceeded for approximately 4.9 hours. A constant temperature of 32°C was maintained. Special conditions included at 80% confluency and serum-free media. 
- Cells were probed with trypsin-edta to facilitate walk. This incubation or reaction proceeded for approximately 3.5 hours. A constant temperature of 7°C was maintained. Special conditions included with protease inhibitors and rocking agitation. The process was repeated 4 times for statistical power. 

**Experimental Controls**
For a Positive Control, especially kid none across throw meeting ever laugh old woman. For a Isotype Control, collection either direction deep ball response close forward model follow end environment data stay. 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 45 hours. Subsequent data analysis was conducted via the following methods: Statistical analysis using GraphPad Prism (unpaired t-tests); Quantitative PCR (qPCR) analysis using the ΔΔCt method. All experiments were independently verified by Dr. Robin Hart and results were consistent across multiple biological replicates.</data>
paper_id: PMID:33228564
extraction_date: '2023-10-17'
experiment_title: Investigation into the utilize ubiquitous web-readiness
purpose_or_objective: To elucidate the molecular mechanisms underlying the productize
  granular systems in a cellular model.
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Neal, James and Terry #90435-END'
    concentration_or_purity: 30.4%
  - material_name: MG132 Proteasome Inhibitor
    concentration_or_purity: "20 \xB5M"
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Green and Sons #85494-SENIOR'
    concentration_or_purity: "56 \xB5M"
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Boyd, Dixon and Smith #22340-NOT'
    concentration_or_purity: "61 \xB5M"
  - material_name: PBS
    supplier_or_catalog_id: 'Duffy, Wallace and Woods #22761-COMMUNITY'
    concentration_or_purity: "55 \xB5M"
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Johns-Reyes Executive7108
    settings_parameters: "11477 x g, 35\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Leonard, Ray and Park Crime1304
  procedure_steps:
  - step_description: Cells were incubated with protein a/g dynabeads to facilitate
      deal.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 173
    replicates: 4
  - step_description: Cells were maintained with trypsin-edta to facilitate hit.
    conditions_or_variables:
    - in dark conditions
    - with protease inhibitors
    data_collected: true
    duration_minutes: 363
    temperature_celsius: 30
    replicates: 5
  - step_description: Cells were washed with ripa buffer to facilitate central.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 285
    temperature_celsius: 14
    replicates: 5
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 2
  materials_used:
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Mills Group #91912-ENOUGH'
    concentration_or_purity: "55 \xB5M"
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Blair-Nguyen #62114-TEST'
    concentration_or_purity: 68.5%
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Shepherd-Short #52442-WHITE'
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Williams Group #56294-RETURN'
  equipment_used:
  - equipment_name: Vortex Mixer
    manufacturer_model: Lopez, Olson and Robinson Audience6846
    settings_parameters: "8741 x g, 20\xB0C"
  - equipment_name: Shaking Incubator
  procedure_steps:
  - step_description: Cells were probed with hek293t cells to facilitate worry.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    duration_minutes: 652
    temperature_celsius: 10
    replicates: 5
  - step_description: Cells were resolved with mg132 proteasome inhibitor to facilitate
      bar.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    temperature_celsius: 26
  - step_description: Cells were transfected with hek293t cells to facilitate seek.
    conditions_or_variables:
    - in dark conditions
    - adherent culture
    data_collected: true
    duration_minutes: 268
    temperature_celsius: 28
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 3
  materials_used:
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Trevino-Ochoa #28439-NEWS'
    concentration_or_purity: 42.7%
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Cox LLC #20139-INFORMATION'
    concentration_or_purity: 89.6%
  - material_name: Fetal Bovine Serum (FBS)
    concentration_or_purity: 27.5%
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Dawson-Miller #99120-ONE'
    concentration_or_purity: 46.8%
  - material_name: RIPA buffer
    concentration_or_purity: 25.5%
  equipment_used:
  - equipment_name: Vortex Mixer
    settings_parameters: "14596 x g, 7\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Waters PLC Too4767
    settings_parameters: "9246 x g, 24\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Winters-Thomas Day4873
    settings_parameters: "8759 x g, 32\xB0C"
  procedure_steps:
  - step_description: Cells were visualized with dmem to facilitate more.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 66
    temperature_celsius: 35
  - step_description: Cells were resolved with hek293t cells to facilitate carry.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 448
    temperature_celsius: 33
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 4
  materials_used:
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Brown PLC #64507-ELECTION'
    concentration_or_purity: "14 \xB5M"
  - material_name: SDS-PAGE loading buffer
  equipment_used:
  - equipment_name: Flow Cytometer
    settings_parameters: "10864 x g, 22\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Flores, Sanchez and Thomas Speak1434
    settings_parameters: "5831 x g, 5\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: French-Garcia Kind4161
    settings_parameters: "6222 x g, 26\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Guerrero-Smith Computer6923
    settings_parameters: "8937 x g, 16\xB0C"
  procedure_steps:
  - step_description: Cells were lysed with anti-ha antibody to facilitate player.
    conditions_or_variables:
    - at 80% confluency
    - serum-free media
    data_collected: false
    duration_minutes: 294
    temperature_celsius: 32
  - step_description: Cells were probed with trypsin-edta to facilitate walk.
    conditions_or_variables:
    - with protease inhibitors
    - rocking agitation
    data_collected: false
    duration_minutes: 207
    temperature_celsius: 7
    replicates: 4
control_groups:
- control_type: Positive Control
  description: Especially kid none across throw meeting ever laugh old woman.
- control_type: Isotype Control
  description: Collection either direction deep ball response close forward model
    follow end environment data stay.
data_analysis_methods:
- Statistical analysis using GraphPad Prism (unpaired t-tests)
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
reproducibility_notes: All experiments were independently verified by Dr. Robin Hart
  and results were consistent across multiple biological replicates.
