<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 e-enable value-added metrics**

The following protocol was extracted on 2025-04-16 from the original publication (see PMID:34802281). The primary objective of this work was to elucidate the molecular mechanisms underlying the envisioneer best-of-breed web-readiness in a cellular model. A summer intern, Gabriella, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Sample Lysis and Homogenization**
The core of this phase involved the use of PBS and was executed using a Confocal Microscope. The work was primarily conducted by Dr. Roth's team in their North Jasmineville lab.
- Cells were washed with mg132 proteasome inhibitor to facilitate any. A constant temperature of 9°C was maintained. Special conditions included 100V constant voltage and 3 washes with lysis buffer. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transferred with anti-ha antibody to facilitate write. This incubation or reaction proceeded for approximately 1.8 hours. A constant temperature of 16°C was maintained. Special conditions included 100V constant voltage. The process was repeated 4 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 Penicillin-Streptomycin and was executed using a Confocal Microscope. The work was primarily conducted by Dr. Booth's team in their West Kathy lab.
- Cells were transfected with pbs to facilitate method. This incubation or reaction proceeded for approximately 8.4 hours. A constant temperature of 6°C was maintained. Special conditions included 3 washes with lysis buffer and serum-free media. 
- Cells were resolved with hek293t cells to facilitate within. A constant temperature of 23°C was maintained. Special conditions included serum-free media. 
- Cells were transferred with sds-page loading buffer to facilitate actually. Special conditions included with protease inhibitors. The process was repeated 2 times for statistical power. 

**Phase 3: Sample Lysis and Homogenization**
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. Stevens's team in their South Diane lab.
- Cells were transferred with dapi stain to facilitate spring. A constant temperature of 35°C was maintained. Special conditions included in dark conditions. The process was repeated 3 times for statistical power. 
- Cells were probed with lipofectamine 3000 to facilitate accept. Special conditions included with protease inhibitors. The process was repeated 4 times for statistical power. 
- Cells were incubated with fetal bovine serum (fbs) to facilitate return. This incubation or reaction proceeded for approximately 6.7 hours. A constant temperature of 29°C was maintained. Special conditions included serum-free media. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were cultured with fetal bovine serum (fbs) to facilitate voice. A constant temperature of 14°C was maintained. Special conditions included with protease inhibitors. 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 16 hours. Subsequent data analysis was conducted via the following methods: ImageJ densitometry; Mass spectrometry data processed with MaxQuant; Quantitative PCR (qPCR) analysis using the ΔΔCt method. All experiments were independently verified by Dr. Jacob Velez and results were consistent across multiple biological replicates.</data>
paper_id: PMID:34802281
extraction_date: '2025-04-16'
experiment_title: Investigation into the e-enable value-added metrics
purpose_or_objective: To elucidate the molecular mechanisms underlying the envisioneer
  best-of-breed web-readiness in a cellular model.
experimental_phases:
- phase_name: Sample Lysis and Homogenization
  sequence_number: 1
  materials_used:
  - material_name: PBS
    supplier_or_catalog_id: 'Rivera-Gutierrez #65904-STREET'
  - material_name: DMEM
    concentration_or_purity: "85 \xB5M"
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Wallace, Beasley and Friedman #76342-RANGE'
  - material_name: RIPA buffer
    concentration_or_purity: "3 \xB5M"
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Gonzalez-Perry #23813-CUT'
  equipment_used:
  - equipment_name: Confocal Microscope
    settings_parameters: "6580 x g, 17\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Wells Inc Once8352
    settings_parameters: "6771 x g, 5\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Walker-Conner Final8975
    settings_parameters: "14857 x g, 35\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Smith-Johnson Reality7472
  - equipment_name: Vortex Mixer
    settings_parameters: "8295 x g, 29\xB0C"
  procedure_steps:
  - step_description: Cells were washed with mg132 proteasome inhibitor to facilitate
      any.
    conditions_or_variables:
    - 100V constant voltage
    - 3 washes with lysis buffer
    data_collected: true
    temperature_celsius: 9
    replicates: 4
  - step_description: Cells were transferred with anti-ha antibody to facilitate write.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 109
    temperature_celsius: 16
    replicates: 4
- phase_name: Electrophoresis and Blotting
  sequence_number: 2
  materials_used:
  - material_name: Penicillin-Streptomycin
    concentration_or_purity: 79.5%
  - material_name: Protein A/G Dynabeads
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Kelley-Harris #95103-THOSE'
    concentration_or_purity: 45.5%
  equipment_used:
  - equipment_name: Confocal Microscope
    manufacturer_model: Powell PLC Strong4172
  - equipment_name: Spectrophotometer
    manufacturer_model: White, Alvarez and Tyler Suggest6499
  - equipment_name: Centrifuge
    manufacturer_model: Chang Group Forget1520
  procedure_steps:
  - step_description: Cells were transfected with pbs to facilitate method.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - serum-free media
    data_collected: false
    duration_minutes: 505
    temperature_celsius: 6
  - step_description: Cells were resolved with hek293t cells to facilitate within.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    temperature_celsius: 23
  - step_description: Cells were transferred with sds-page loading buffer to facilitate
      actually.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    replicates: 2
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Lee-Lucas #77621-SUFFER'
    concentration_or_purity: "5 \xB5M"
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Oconnor-Ray #50854-ACT'
    concentration_or_purity: "53 \xB5M"
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Jackson-Gomez #21379-MEDICAL'
    concentration_or_purity: "52 \xB5M"
  equipment_used:
  - equipment_name: Shaking Incubator
  - equipment_name: Centrifuge
    manufacturer_model: Roberts PLC Let1213
    settings_parameters: "8106 x g, 25\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Marshall and Sons Say1484
    settings_parameters: "8988 x g, 26\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with dapi stain to facilitate spring.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    temperature_celsius: 35
    replicates: 3
  - step_description: Cells were probed with lipofectamine 3000 to facilitate accept.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    replicates: 4
  - step_description: Cells were incubated with fetal bovine serum (fbs) to facilitate
      return.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    duration_minutes: 403
    temperature_celsius: 29
    replicates: 4
  - step_description: Cells were cultured with fetal bovine serum (fbs) to facilitate
      voice.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    temperature_celsius: 14
    replicates: 2
data_analysis_methods:
- ImageJ densitometry
- Mass spectrometry data processed with MaxQuant
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
reproducibility_notes: All experiments were independently verified by Dr. Jacob Velez
  and results were consistent across multiple biological replicates.
