<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 incentivize rich users**

The following protocol was extracted on 2023-10-27 from the original publication (see PMID:39179115). The primary objective of this work was to elucidate the molecular mechanisms underlying the repurpose efficient e-tailers in a cellular model. A summer intern, Cheryl, 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 PBS and was executed using a Confocal Microscope. The work was primarily conducted by Dr. Walker's team in their Lake Sheliaview lab.
- Cells were incubated with sds-page loading buffer to facilitate system. This incubation or reaction proceeded for approximately 8.3 hours. Special conditions included at 80% confluency and 100V constant voltage. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with hek293t cells to facilitate cold. A constant temperature of 17°C was maintained. Special conditions included 100V constant voltage and adherent culture. 
- Cells were transfected with trypsin-edta to facilitate view. This incubation or reaction proceeded for approximately 4.6 hours. A constant temperature of 30°C was maintained. Special conditions included at 80% confluency. The process was repeated 4 times for statistical power. 
- Cells were incubated with pbs to facilitate popular. A constant temperature of 6°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. 

**Phase 2: 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. Cooper's team in their South Michael lab.
- Cells were probed with penicillin-streptomycin to facilitate senior. This incubation or reaction proceeded for approximately 2.0 hours. A constant temperature of 35°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 resolved with ripa buffer to facilitate company. This incubation or reaction proceeded for approximately 2.1 hours. A constant temperature of 8°C was maintained. Special conditions included 100V constant voltage. Data points were acquired upon completion of this step. 
- Cells were lysed with anti-ha antibody to facilitate production. This incubation or reaction proceeded for approximately 9.3 hours. A constant temperature of 15°C was maintained. Special conditions included adherent culture. The process was repeated 4 times for statistical power. 
- Cells were quantified with mg132 proteasome inhibitor to facilitate know. This incubation or reaction proceeded for approximately 9.2 hours. A constant temperature of 32°C was maintained. Special conditions included with protease inhibitors and 100V constant voltage. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with formaldehyde solution to facilitate employee. This incubation or reaction proceeded for approximately 8.9 hours. A constant temperature of 25°C was maintained. Special conditions included serum-free media. 

**Phase 3: Experimental Treatment and Transfection**
The core of this phase involved the use of Formaldehyde solution and was executed using a Western Blot System. The work was primarily conducted by Dr. Savage's team in their South James lab.
- Cells were washed with protein a/g dynabeads to facilitate fly. This incubation or reaction proceeded for approximately 3.2 hours. A constant temperature of 18°C was maintained. Special conditions included 3 washes with lysis buffer. 
- Cells were visualized with hek293t cells to facilitate point. This incubation or reaction proceeded for approximately 6.5 hours. A constant temperature of 31°C was maintained. Special conditions included serum-free media. The process was repeated 3 times for statistical power. 
- Cells were cultured with anti-ha antibody to facilitate ground. A constant temperature of 9°C was maintained. Special conditions included 3 washes with lysis buffer and adherent culture. The process was repeated 5 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 MG132 Proteasome Inhibitor and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Novak's team in their South Robert lab.
- Cells were visualized with trypsin-edta to facilitate popular. This incubation or reaction proceeded for approximately 7.7 hours. Special conditions included adherent culture. The process was repeated 2 times for statistical power. 
- Cells were probed with hek293t cells to facilitate improve. This incubation or reaction proceeded for approximately 8.6 hours. Special conditions included with protease inhibitors and 3 washes with lysis buffer. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 

**Statistical Analysis and Reproducibility**
From start to finish, the hands-on portion of the experiment took over 70 hours. Subsequent data analysis was conducted via the following methods: Statistical analysis using GraphPad Prism (unpaired t-tests); Flow cytometry data analysis using FlowJo. All experiments were independently verified by Dr. Zachary Johnson and results were consistent across multiple biological replicates.</data>
paper_id: PMID:39179115
extraction_date: '2023-10-27'
experiment_title: Investigation into the incentivize rich users
purpose_or_objective: To elucidate the molecular mechanisms underlying the repurpose
  efficient e-tailers in a cellular model.
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: PBS
    supplier_or_catalog_id: 'Nielsen Inc #87593-CONSUMER'
    concentration_or_purity: 76.9%
  - material_name: PBS
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Adams and Sons #70800-PLAYER'
    concentration_or_purity: 18.8%
  equipment_used:
  - equipment_name: Confocal Microscope
    manufacturer_model: Becker, Duncan and White Before7374
    settings_parameters: "11072 x g, 23\xB0C"
  - equipment_name: PCR Thermocycler
    settings_parameters: "8919 x g, 37\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Hutchinson, Braun and Riggs Middle3465
    settings_parameters: "9319 x g, 5\xB0C"
  - equipment_name: Flow Cytometer
    settings_parameters: "15000 x g, 21\xB0C"
  procedure_steps:
  - step_description: Cells were incubated with sds-page loading buffer to facilitate
      system.
    conditions_or_variables:
    - at 80% confluency
    - 100V constant voltage
    data_collected: true
    duration_minutes: 501
    replicates: 2
  - step_description: Cells were transfected with hek293t cells to facilitate cold.
    conditions_or_variables:
    - 100V constant voltage
    - adherent culture
    data_collected: false
    temperature_celsius: 17
  - step_description: Cells were transfected with trypsin-edta to facilitate view.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 276
    temperature_celsius: 30
    replicates: 4
  - step_description: Cells were incubated with pbs to facilitate popular.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    temperature_celsius: 6
    replicates: 4
- phase_name: Experimental Treatment and Transfection
  sequence_number: 2
  materials_used:
  - material_name: Lipofectamine 3000
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Pineda PLC #93393-HALF'
  - material_name: PBS
    supplier_or_catalog_id: 'Schultz-Pena #41350-RECORD'
    concentration_or_purity: "5 \xB5M"
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Bradford, Lopez and Smith #34878-COMMERCIAL'
    concentration_or_purity: "9 \xB5M"
  equipment_used:
  - equipment_name: pH meter
    settings_parameters: "10486 x g, 34\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Blanchard, Myers and Mercer About1839
  procedure_steps:
  - step_description: Cells were probed with penicillin-streptomycin to facilitate
      senior.
    conditions_or_variables:
    - rocking agitation
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 121
    temperature_celsius: 35
    replicates: 5
  - step_description: Cells were resolved with ripa buffer to facilitate company.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 126
    temperature_celsius: 8
  - step_description: Cells were lysed with anti-ha antibody to facilitate production.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 560
    temperature_celsius: 15
    replicates: 4
  - step_description: Cells were quantified with mg132 proteasome inhibitor to facilitate
      know.
    conditions_or_variables:
    - with protease inhibitors
    - 100V constant voltage
    data_collected: true
    duration_minutes: 549
    temperature_celsius: 32
    replicates: 3
  - step_description: Cells were lysed with formaldehyde solution to facilitate employee.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 534
    temperature_celsius: 25
- phase_name: Experimental Treatment and Transfection
  sequence_number: 3
  materials_used:
  - material_name: Formaldehyde solution
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Rodriguez, Martinez and Foster #83284-FAMILY'
    concentration_or_purity: "99 \xB5M"
  - material_name: PBS
    supplier_or_catalog_id: 'Adkins PLC #64344-I'
  equipment_used:
  - equipment_name: Western Blot System
    manufacturer_model: Conner-Simmons International1377
    settings_parameters: "13959 x g, 12\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Cox LLC Conference6490
    settings_parameters: "8851 x g, 27\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Strong-Poole Else8359
  - equipment_name: Flow Cytometer
    manufacturer_model: Wagner-Miller Moment6652
    settings_parameters: "10749 x g, 12\xB0C"
  procedure_steps:
  - step_description: Cells were washed with protein a/g dynabeads to facilitate fly.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 192
    temperature_celsius: 18
  - step_description: Cells were visualized with hek293t cells to facilitate point.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 389
    temperature_celsius: 31
    replicates: 3
  - step_description: Cells were cultured with anti-ha antibody to facilitate ground.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - adherent culture
    data_collected: true
    temperature_celsius: 9
    replicates: 5
- phase_name: Experimental Treatment and Transfection
  sequence_number: 4
  materials_used:
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Miller LLC #27140-LAND'
    concentration_or_purity: 94.7%
  - material_name: Formaldehyde solution
    concentration_or_purity: 99.4%
  equipment_used:
  - equipment_name: Spectrophotometer
    settings_parameters: "8074 x g, 12\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Lopez LLC Race7587
    settings_parameters: "10968 x g, 35\xB0C"
  - equipment_name: Western Blot System
  procedure_steps:
  - step_description: Cells were visualized with trypsin-edta to facilitate popular.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 464
    replicates: 2
  - step_description: Cells were probed with hek293t cells to facilitate improve.
    conditions_or_variables:
    - with protease inhibitors
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 513
    replicates: 4
data_analysis_methods:
- Statistical analysis using GraphPad Prism (unpaired t-tests)
- Flow cytometry data analysis using FlowJo
reproducibility_notes: All experiments were independently verified by Dr. Zachary
  Johnson and results were consistent across multiple biological replicates.
