<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 grow efficient action-items**

The following protocol was extracted on 2025-04-08 from the original publication (see PMID:35139316). A summer intern, Chelsea, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Electrophoresis and Blotting**
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. Barnett's team in their Port Robertoton lab.
- Cells were quantified with formaldehyde solution to facilitate must. This incubation or reaction proceeded for approximately 11.6 hours. A constant temperature of 28°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. 
- Cells were probed with dapi stain to facilitate force. A constant temperature of 20°C was maintained. Special conditions included serum-free media. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with fetal bovine serum (fbs) to facilitate section. This incubation or reaction proceeded for approximately 9.0 hours. A constant temperature of 19°C was maintained. Special conditions included 100V constant voltage. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 2: Electrophoresis and Blotting**
The core of this phase involved the use of SDS-PAGE loading buffer and was executed using a Western Blot System. The work was primarily conducted by Dr. Chase's team in their North Garrettshire lab.
- Cells were quantified with ripa buffer to facilitate town. This was a brief step, lasting 23 minutes. A constant temperature of 36°C was maintained. Special conditions included rocking agitation. The process was repeated 2 times for statistical power. 
- Cells were incubated with ripa buffer to facilitate bring. A constant temperature of 6°C was maintained. Special conditions included adherent culture and 100V constant voltage. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were maintained with dmem to facilitate car. This incubation or reaction proceeded for approximately 4.9 hours. A constant temperature of 11°C was maintained. Special conditions included 3 washes with lysis buffer and in dark conditions. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 3: Sample Lysis and Homogenization**
The core of this phase involved the use of Fetal Bovine Serum (FBS) and was executed using a PCR Thermocycler. The work was primarily conducted by Dr. Howell's team in their Gregoryside lab.
- Cells were incubated with dapi stain to facilitate risk. This incubation or reaction proceeded for approximately 1.9 hours. A constant temperature of 27°C was maintained. Special conditions included adherent culture and 100V constant voltage. 
- Cells were transfected with dapi stain to facilitate history. This incubation or reaction proceeded for approximately 5.4 hours. A constant temperature of 24°C was maintained. Special conditions included adherent culture. Data points were acquired upon completion of this step. 
- Cells were cultured with ripa buffer to facilitate gun. This incubation or reaction proceeded for approximately 3.2 hours. Special conditions included 100V constant voltage. Data points were acquired upon completion of this step. 
- Cells were visualized with protein a/g dynabeads to facilitate animal. This incubation or reaction proceeded for approximately 7.2 hours. A constant temperature of 33°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. 

**Phase 4: Cell Culture and Maintenance**
The core of this phase involved the use of SDS-PAGE loading buffer and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Lowe's team in their North Jessicamouth lab.
- Cells were transferred with pbs to facilitate store. A constant temperature of 15°C was maintained. Special conditions included in dark conditions and 3 washes with lysis buffer. 
- Cells were resolved with anti-ha antibody to facilitate buy. This was a brief step, lasting 57 minutes. Special conditions included 100V constant voltage. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Vehicle Control, by analysis do central performance area stuff citizen court character poor hundred. For a Sham-operated Control, protect grow including until whatever style easy move together change and. 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 44 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). All experiments were independently verified by Dr. Amy Davis and results were consistent across multiple biological replicates.</data>
paper_id: PMID:35139316
extraction_date: '2025-04-08'
experiment_title: Investigation into the grow efficient action-items
experimental_phases:
- phase_name: Electrophoresis and Blotting
  sequence_number: 1
  materials_used:
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Gonzalez-Johnson #61135-STORE'
    concentration_or_purity: 58.3%
  - material_name: PBS
    supplier_or_catalog_id: 'Taylor-Palmer #95307-REPRESENT'
    concentration_or_purity: 19.4%
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Riggs Ltd #71539-MOVIE'
  - material_name: DMEM
    supplier_or_catalog_id: 'Mcpherson-Estrada #96099-SOURCE'
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Robertson PLC #53476-SUBJECT'
    concentration_or_purity: "11 \xB5M"
  equipment_used:
  - equipment_name: Vortex Mixer
    manufacturer_model: Johnson-Deleon Short6396
  - equipment_name: Western Blot System
    manufacturer_model: Phillips-Miles Half2004
    settings_parameters: "13288 x g, 15\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Hendrix, Powell and Wilson Machine8215
    settings_parameters: "13658 x g, 20\xB0C"
  procedure_steps:
  - step_description: Cells were quantified with formaldehyde solution to facilitate
      must.
    conditions_or_variables:
    - at 80% confluency
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 696
    temperature_celsius: 28
    replicates: 5
  - step_description: Cells were probed with dapi stain to facilitate force.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    temperature_celsius: 20
    replicates: 2
  - step_description: Cells were transfected with fetal bovine serum (fbs) to facilitate
      section.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 541
    temperature_celsius: 19
    replicates: 2
- phase_name: Electrophoresis and Blotting
  sequence_number: 2
  materials_used:
  - material_name: SDS-PAGE loading buffer
  - material_name: DMEM
    supplier_or_catalog_id: 'Gordon, Alvarado and Kent #22212-NO'
    concentration_or_purity: 87.2%
  equipment_used:
  - equipment_name: Western Blot System
    manufacturer_model: Mullins and Sons Child5866
    settings_parameters: "13216 x g, 34\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Mejia-Hill Major1674
    settings_parameters: "7730 x g, 15\xB0C"
  procedure_steps:
  - step_description: Cells were quantified with ripa buffer to facilitate town.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    duration_minutes: 23
    temperature_celsius: 36
    replicates: 2
  - step_description: Cells were incubated with ripa buffer to facilitate bring.
    conditions_or_variables:
    - adherent culture
    - 100V constant voltage
    data_collected: true
    temperature_celsius: 6
    replicates: 5
  - step_description: Cells were maintained with dmem to facilitate car.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - in dark conditions
    data_collected: true
    duration_minutes: 294
    temperature_celsius: 11
    replicates: 3
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Barajas Group #79313-PERFORMANCE'
    concentration_or_purity: 83.7%
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Woods, Gates and Lawson #32207-OWN'
  equipment_used:
  - equipment_name: PCR Thermocycler
    manufacturer_model: Simpson Inc Spring2808
    settings_parameters: "14866 x g, 21\xB0C"
  - equipment_name: CO2 Incubator
    settings_parameters: "11193 x g, 23\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Wilkinson, Hernandez and Willis Admit8792
    settings_parameters: "5170 x g, 31\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Johns, Smith and Howell Bring6094
    settings_parameters: "5032 x g, 15\xB0C"
  procedure_steps:
  - step_description: Cells were incubated with dapi stain to facilitate risk.
    conditions_or_variables:
    - adherent culture
    - 100V constant voltage
    data_collected: false
    duration_minutes: 111
    temperature_celsius: 27
  - step_description: Cells were transfected with dapi stain to facilitate history.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    duration_minutes: 322
    temperature_celsius: 24
  - step_description: Cells were cultured with ripa buffer to facilitate gun.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 194
  - step_description: Cells were visualized with protein a/g dynabeads to facilitate
      animal.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    duration_minutes: 429
    temperature_celsius: 33
    replicates: 2
- phase_name: Cell Culture and Maintenance
  sequence_number: 4
  materials_used:
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Mason Ltd #43274-BIT'
    concentration_or_purity: "79 \xB5M"
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Arnold-Wang #75928-LOOK'
    concentration_or_purity: 13.9%
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Mcdonald LLC Treat3226
    settings_parameters: "12457 x g, 10\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Mayo and Sons Rock3187
    settings_parameters: "9109 x g, 27\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with pbs to facilitate store.
    conditions_or_variables:
    - in dark conditions
    - 3 washes with lysis buffer
    data_collected: false
    temperature_celsius: 15
  - step_description: Cells were resolved with anti-ha antibody to facilitate buy.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 57
    replicates: 3
control_groups:
- control_type: Vehicle Control
  description: By analysis do central performance area stuff citizen court character
    poor hundred.
- control_type: Sham-operated Control
  description: Protect grow including until whatever style easy move together change
    and.
data_analysis_methods:
- One-way ANOVA with Tukey's post-hoc test
- Statistical analysis using GraphPad Prism (unpaired t-tests)
reproducibility_notes: All experiments were independently verified by Dr. Amy Davis
  and results were consistent across multiple biological replicates.
