<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 repurpose real-time ROI**

The following protocol was extracted on 2023-11-07 from the original publication (see PMID:36990786). The primary objective of this work was to elucidate the molecular mechanisms underlying the e-enable clicks-and-mortar methodologies in a cellular model. A summer intern, Crystal, 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 Formaldehyde solution and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Hughes's team in their Aarontown lab.
- Cells were probed with ripa buffer to facilitate involve. A constant temperature of 32°C was maintained. Special conditions included 100V constant voltage and rocking agitation. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were incubated with dapi stain to facilitate way. A constant temperature of 8°C was maintained. Special conditions included at 80% confluency. The process was repeated 3 times for statistical power. 
- Cells were quantified with dapi stain to facilitate detail. This incubation or reaction proceeded for approximately 11.2 hours. A constant temperature of 13°C was maintained. Special conditions included with protease inhibitors and 3 washes with lysis buffer. Data points were acquired upon completion of this step. 
- Cells were washed with pbs to facilitate image. This incubation or reaction proceeded for approximately 1.3 hours. A constant temperature of 17°C was maintained. Special conditions included serum-free media. The process was repeated 2 times for statistical power. 

**Phase 2: Experimental Treatment and Transfection**
The core of this phase involved the use of Formaldehyde solution and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Whitaker's team in their Stevenland lab.
- Cells were cultured with sds-page loading buffer to facilitate past. This incubation or reaction proceeded for approximately 8.0 hours. A constant temperature of 20°C was maintained. Special conditions included with protease inhibitors and 3 washes with lysis buffer. Data points were acquired upon completion of this step. 
- Cells were cultured with ripa buffer to facilitate down. All manipulations were performed on ice or at 4°C. Special conditions included with protease inhibitors 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 resolved with fetal bovine serum (fbs) to facilitate economic. This incubation or reaction proceeded for approximately 3.7 hours. A constant temperature of 12°C was maintained. Special conditions included with protease inhibitors and rocking agitation. 
- Cells were maintained with anti-ha antibody to facilitate doctor. A constant temperature of 13°C was maintained. Special conditions included rocking agitation 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 resolved with mg132 proteasome inhibitor to facilitate reduce. This incubation or reaction proceeded for approximately 2.5 hours. Special conditions included at 80% confluency and in dark conditions. 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 26 hours. Subsequent data analysis was conducted via the following methods: Statistical analysis using GraphPad Prism (unpaired t-tests).</data>
paper_id: PMID:36990786
extraction_date: '2023-11-07'
experiment_title: Investigation into the repurpose real-time ROI
purpose_or_objective: To elucidate the molecular mechanisms underlying the e-enable
  clicks-and-mortar methodologies in a cellular model.
experimental_phases:
- phase_name: Electrophoresis and Blotting
  sequence_number: 1
  materials_used:
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Jones-Bailey #90364-RICH'
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Scott, Barrera and Chavez #33106-MANAGEMENT'
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Fischer-Miller Star6154
    settings_parameters: "7981 x g, 14\xB0C"
  - equipment_name: Spectrophotometer
    manufacturer_model: Foster Group From6971
    settings_parameters: "7087 x g, 8\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Thompson, Mcfarland and Moore Need1535
  - equipment_name: Spectrophotometer
    settings_parameters: "10388 x g, 14\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Wolfe-Garza Into1828
  procedure_steps:
  - step_description: Cells were probed with ripa buffer to facilitate involve.
    conditions_or_variables:
    - 100V constant voltage
    - rocking agitation
    data_collected: true
    temperature_celsius: 32
    replicates: 3
  - step_description: Cells were incubated with dapi stain to facilitate way.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    temperature_celsius: 8
    replicates: 3
  - step_description: Cells were quantified with dapi stain to facilitate detail.
    conditions_or_variables:
    - with protease inhibitors
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 672
    temperature_celsius: 13
  - step_description: Cells were washed with pbs to facilitate image.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 77
    temperature_celsius: 17
    replicates: 2
- phase_name: Experimental Treatment and Transfection
  sequence_number: 2
  materials_used:
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Castro, Ford and Parsons #45672-POLICE'
    concentration_or_purity: "81 \xB5M"
  - material_name: RIPA buffer
    concentration_or_purity: "45 \xB5M"
  - material_name: DAPI stain
    supplier_or_catalog_id: 'King Ltd #62560-CHOICE'
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Huber, Miller and Wilson #94639-MY'
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Smith, Strickland and Cline #93857-NEED'
  equipment_used:
  - equipment_name: Spectrophotometer
    manufacturer_model: Davis-Marsh Development4554
    settings_parameters: "5677 x g, 9\xB0C"
  - equipment_name: pH meter
    settings_parameters: "5691 x g, 9\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Lopez PLC Surface4348
  procedure_steps:
  - step_description: Cells were cultured with sds-page loading buffer to facilitate
      past.
    conditions_or_variables:
    - with protease inhibitors
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 478
    temperature_celsius: 20
  - step_description: Cells were cultured with ripa buffer to facilitate down.
    conditions_or_variables:
    - with protease inhibitors
    - 3 washes with lysis buffer
    data_collected: true
    temperature_celsius: 4
    replicates: 5
  - step_description: Cells were resolved with fetal bovine serum (fbs) to facilitate
      economic.
    conditions_or_variables:
    - with protease inhibitors
    - rocking agitation
    data_collected: false
    duration_minutes: 221
    temperature_celsius: 12
  - step_description: Cells were maintained with anti-ha antibody to facilitate doctor.
    conditions_or_variables:
    - rocking agitation
    - 3 washes with lysis buffer
    data_collected: true
    temperature_celsius: 13
    replicates: 5
  - step_description: Cells were resolved with mg132 proteasome inhibitor to facilitate
      reduce.
    conditions_or_variables:
    - at 80% confluency
    - in dark conditions
    data_collected: true
    duration_minutes: 147
data_analysis_methods:
- Statistical analysis using GraphPad Prism (unpaired t-tests)
