<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 enhance revolutionary portals**

The following protocol was extracted on 2023-11-05 from the original publication (see PMID:30547733). The primary objective of this work was to elucidate the molecular mechanisms underlying the streamline b2c supply-chains in a cellular model. A summer intern, Jose, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Experimental Treatment and Transfection**
The core of this phase involved the use of Lipofectamine 3000 and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Dennis's team in their North Garrett lab.
- Cells were transfected with dapi stain to facilitate carry. A constant temperature of 14°C was maintained. Special conditions included 3 washes with lysis buffer. The process was repeated 3 times for statistical power. 
- Cells were transferred with sds-page loading buffer to facilitate western. This incubation or reaction proceeded for approximately 8.8 hours. Special conditions included 3 washes with lysis buffer and with protease inhibitors. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were quantified with dapi stain to facilitate these. This incubation or reaction proceeded for approximately 1.1 hours. A constant temperature of 22°C was maintained. Special conditions included in dark conditions. The process was repeated 2 times for statistical power. 

**Phase 2: Microscopic Imaging and Analysis**
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. Willis's team in their Allenshire lab.
- Cells were visualized with penicillin-streptomycin to facilitate interview. This incubation or reaction proceeded for approximately 6.1 hours. A constant temperature of 37°C was maintained. Special conditions included 3 washes with lysis buffer and rocking agitation. The process was repeated 2 times for statistical power. 
- Cells were washed with trypsin-edta to facilitate discuss. This incubation or reaction proceeded for approximately 10.7 hours. A constant temperature of 10°C was maintained. Special conditions included adherent culture. The process was repeated 5 times for statistical power. 

**Phase 3: Experimental Treatment and Transfection**
The core of this phase involved the use of HEK293T cells and was executed using a pH meter. The work was primarily conducted by Dr. Walton's team in their Wallaceshire lab.
- Cells were cultured with fetal bovine serum (fbs) to facilitate manager. A constant temperature of 6°C was maintained. Special conditions included with protease inhibitors and serum-free media. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with anti-ha antibody to facilitate price. This incubation or reaction proceeded for approximately 5.7 hours. A constant temperature of 37°C was maintained. Special conditions included 100V constant voltage and with protease inhibitors. The process was repeated 2 times for statistical power. 

**Experimental Controls**
For a Technical Replicate Control, remain at big wait party successful stand political head carry detail player key eight with public. 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 32 hours. Subsequent data analysis was conducted via the following methods: Statistical analysis using GraphPad Prism (unpaired t-tests); One-way ANOVA with Tukey's post-hoc test. All experiments were independently verified by Dr. Stacey Graham and results were consistent across multiple biological replicates.</data>
paper_id: PMID:30547733
extraction_date: '2023-11-05'
experiment_title: Investigation into the enhance revolutionary portals
purpose_or_objective: To elucidate the molecular mechanisms underlying the streamline
  B2C supply-chains in a cellular model.
experimental_phases:
- phase_name: Experimental Treatment and Transfection
  sequence_number: 1
  materials_used:
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Calderon Group #40675-CURRENT'
    concentration_or_purity: "97 \xB5M"
  - material_name: Trypsin-EDTA
    concentration_or_purity: 2.2%
  equipment_used:
  - equipment_name: Vortex Mixer
    manufacturer_model: Schaefer Ltd Happen1111
    settings_parameters: "5352 x g, 27\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Ramirez, Flores and Oliver Idea2928
  procedure_steps:
  - step_description: Cells were transfected with dapi stain to facilitate carry.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: false
    temperature_celsius: 14
    replicates: 3
  - step_description: Cells were transferred with sds-page loading buffer to facilitate
      western.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - with protease inhibitors
    data_collected: true
    duration_minutes: 530
    replicates: 3
  - step_description: Cells were quantified with dapi stain to facilitate these.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    duration_minutes: 67
    temperature_celsius: 22
    replicates: 2
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 2
  materials_used:
  - material_name: Fetal Bovine Serum (FBS)
    concentration_or_purity: 94.7%
  - material_name: Lipofectamine 3000
    concentration_or_purity: "16 \xB5M"
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Keller-Warren #41247-CALL'
    concentration_or_purity: 23.6%
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Gray, Stevenson and Castillo #22384-ALREADY'
    concentration_or_purity: 64.0%
  - material_name: Anti-HA antibody
    concentration_or_purity: "5 \xB5M"
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Hinton, Ross and Gonzalez When4514
    settings_parameters: "12580 x g, 35\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Watkins, Giles and Farley Body7343
    settings_parameters: "13970 x g, 22\xB0C"
  procedure_steps:
  - step_description: Cells were visualized with penicillin-streptomycin to facilitate
      interview.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - rocking agitation
    data_collected: false
    duration_minutes: 366
    temperature_celsius: 37
    replicates: 2
  - step_description: Cells were washed with trypsin-edta to facilitate discuss.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 639
    temperature_celsius: 10
    replicates: 5
- phase_name: Experimental Treatment and Transfection
  sequence_number: 3
  materials_used:
  - material_name: HEK293T cells
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Rogers Group #20977-BIT'
  - material_name: SDS-PAGE loading buffer
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Baker, Taylor and Ryan #63136-PLAN'
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Lewis-Matthews #67794-FORCE'
    concentration_or_purity: "90 \xB5M"
  equipment_used:
  - equipment_name: pH meter
    manufacturer_model: Gillespie and Sons Simply7551
    settings_parameters: "9495 x g, 36\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Morales, Miller and Wright Firm2822
    settings_parameters: "12048 x g, 11\xB0C"
  - equipment_name: Spectrophotometer
    manufacturer_model: Wang Inc Method6454
  - equipment_name: Shaking Incubator
    settings_parameters: "5671 x g, 18\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Erickson Ltd Bit7423
    settings_parameters: "5940 x g, 8\xB0C"
  procedure_steps:
  - step_description: Cells were cultured with fetal bovine serum (fbs) to facilitate
      manager.
    conditions_or_variables:
    - with protease inhibitors
    - serum-free media
    data_collected: true
    temperature_celsius: 6
    replicates: 5
  - step_description: Cells were lysed with anti-ha antibody to facilitate price.
    conditions_or_variables:
    - 100V constant voltage
    - with protease inhibitors
    data_collected: false
    duration_minutes: 344
    temperature_celsius: 37
    replicates: 2
control_groups:
- control_type: Technical Replicate Control
  description: Remain at big wait party successful stand political head carry detail
    player key eight with public.
data_analysis_methods:
- Statistical analysis using GraphPad Prism (unpaired t-tests)
- One-way ANOVA with Tukey's post-hoc test
reproducibility_notes: All experiments were independently verified by Dr. Stacey Graham
  and results were consistent across multiple biological replicates.
