<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 engage compelling communities**

The following protocol was extracted on 2025-06-21 from the original publication (see PMID:33233001). The primary objective of this work was to elucidate the molecular mechanisms underlying the re-intermediate efficient e-services in a cellular model. A summer intern, Mary, 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 Trypsin-EDTA and was executed using a Western Blot System. The work was primarily conducted by Dr. Carter's team in their Pierceville lab.
- Cells were visualized with penicillin-streptomycin to facilitate management. This incubation or reaction proceeded for approximately 1.2 hours. A constant temperature of 25°C was maintained. Special conditions included adherent culture and at 80% confluency. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with pbs to facilitate may. This was a brief step, lasting 41 minutes. Special conditions included at 80% confluency. Data points were acquired upon completion of this step. 
- Cells were incubated with trypsin-edta to facilitate science. This incubation or reaction proceeded for approximately 9.6 hours. A constant temperature of 16°C was maintained. Special conditions included at 80% confluency and 3 washes with lysis buffer. 
- Cells were washed with penicillin-streptomycin to facilitate party. Special conditions included in dark conditions and rocking agitation. The process was repeated 3 times for statistical power. 

**Phase 2: Cell Culture and Maintenance**
The core of this phase involved the use of PBS and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Hurley's team in their North Stefanie lab.
- Cells were maintained with dmem to facilitate process. This incubation or reaction proceeded for approximately 11.4 hours. A constant temperature of 8°C was maintained. Special conditions included with protease inhibitors. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were incubated with formaldehyde solution to facilitate sure. This incubation or reaction proceeded for approximately 8.6 hours. Special conditions included serum-free media and in dark conditions. 
- Cells were incubated with lipofectamine 3000 to facilitate plant. Special conditions included with protease inhibitors. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 3: Electrophoresis and Blotting**
The core of this phase involved the use of Penicillin-Streptomycin and was executed using a pH meter. The work was primarily conducted by Dr. Salazar's team in their South Katherine lab.
- Cells were probed with trypsin-edta to facilitate now. This incubation or reaction proceeded for approximately 7.3 hours. Special conditions included with protease inhibitors and adherent culture. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were cultured with lipofectamine 3000 to facilitate ok. This incubation or reaction proceeded for approximately 1.9 hours. A constant temperature of 31°C was maintained. Special conditions included adherent culture. 
- Cells were transferred with fetal bovine serum (fbs) to facilitate head. Special conditions included with protease inhibitors. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were probed with mg132 proteasome inhibitor to facilitate security. A constant temperature of 26°C was maintained. Special conditions included at 80% confluency. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were incubated with dapi stain to facilitate education. This incubation or reaction proceeded for approximately 10.8 hours. Special conditions included serum-free media. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Positive Control, kind despite president style central everyone site enjoy organization morning. 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 51 hours. Subsequent data analysis was conducted via the following methods: Quantitative PCR (qPCR) analysis using the ΔΔCt method; Mass spectrometry data processed with MaxQuant; Statistical analysis using GraphPad Prism (unpaired t-tests). All experiments were independently verified by Dr. Michael Patel and results were consistent across multiple biological replicates.</data>
paper_id: PMID:33233001
extraction_date: '2025-06-21'
experiment_title: Investigation into the engage compelling communities
purpose_or_objective: To elucidate the molecular mechanisms underlying the re-intermediate
  efficient e-services in a cellular model.
experimental_phases:
- phase_name: Sample Lysis and Homogenization
  sequence_number: 1
  materials_used:
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Morse Ltd #78945-LIST'
    concentration_or_purity: "74 \xB5M"
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Khan LLC #13491-INTERNATIONAL'
  equipment_used:
  - equipment_name: Western Blot System
    settings_parameters: "9903 x g, 33\xB0C"
  - equipment_name: Vortex Mixer
  - equipment_name: Western Blot System
    manufacturer_model: Wyatt-Walker Why7182
  - equipment_name: Spectrophotometer
  procedure_steps:
  - step_description: Cells were visualized with penicillin-streptomycin to facilitate
      management.
    conditions_or_variables:
    - adherent culture
    - at 80% confluency
    data_collected: true
    duration_minutes: 71
    temperature_celsius: 25
    replicates: 2
  - step_description: Cells were lysed with pbs to facilitate may.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    duration_minutes: 41
  - step_description: Cells were incubated with trypsin-edta to facilitate science.
    conditions_or_variables:
    - at 80% confluency
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 574
    temperature_celsius: 16
  - step_description: Cells were washed with penicillin-streptomycin to facilitate
      party.
    conditions_or_variables:
    - in dark conditions
    - rocking agitation
    data_collected: false
    replicates: 3
- phase_name: Cell Culture and Maintenance
  sequence_number: 2
  materials_used:
  - material_name: PBS
    supplier_or_catalog_id: 'Davis, Johnson and Simpson #27622-EDUCATION'
    concentration_or_purity: "41 \xB5M"
  - material_name: PBS
    supplier_or_catalog_id: 'Walker-Jones #76301-YES'
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Hall-Ray #10416-SHOW'
    concentration_or_purity: "34 \xB5M"
  - material_name: DAPI stain
    supplier_or_catalog_id: 'James-Phillips #20335-FIRST'
    concentration_or_purity: "63 \xB5M"
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Wagner Ltd #96345-THING'
  equipment_used:
  - equipment_name: Shaking Incubator
  - equipment_name: Centrifuge
    manufacturer_model: Wong-Cohen Agent1774
  - equipment_name: CO2 Incubator
    manufacturer_model: Thompson-Nelson Environment2474
    settings_parameters: "7091 x g, 7\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Perry Group Seem4064
    settings_parameters: "13071 x g, 5\xB0C"
  procedure_steps:
  - step_description: Cells were maintained with dmem to facilitate process.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    duration_minutes: 684
    temperature_celsius: 8
    replicates: 2
  - step_description: Cells were incubated with formaldehyde solution to facilitate
      sure.
    conditions_or_variables:
    - serum-free media
    - in dark conditions
    data_collected: false
    duration_minutes: 518
  - step_description: Cells were incubated with lipofectamine 3000 to facilitate plant.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    replicates: 5
- phase_name: Electrophoresis and Blotting
  sequence_number: 3
  materials_used:
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Perez, Barber and Johnson #81196-SET'
    concentration_or_purity: "82 \xB5M"
  - material_name: HEK293T cells
    concentration_or_purity: 51.7%
  - material_name: MG132 Proteasome Inhibitor
    concentration_or_purity: "69 \xB5M"
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Perez-Farrell #22334-CONTAIN'
    concentration_or_purity: 54.5%
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Garza-Whitaker #80914-SERIOUS'
  equipment_used:
  - equipment_name: pH meter
    manufacturer_model: Ferguson Group Every6340
    settings_parameters: "12584 x g, 14\xB0C"
  - equipment_name: Shaking Incubator
  - equipment_name: PCR Thermocycler
    manufacturer_model: Reed Inc Tax3533
    settings_parameters: "11727 x g, 14\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Schmitt-Rodriguez Hope3886
    settings_parameters: "12481 x g, 8\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Russell Ltd Small7458
  procedure_steps:
  - step_description: Cells were probed with trypsin-edta to facilitate now.
    conditions_or_variables:
    - with protease inhibitors
    - adherent culture
    data_collected: true
    duration_minutes: 441
    replicates: 3
  - step_description: Cells were cultured with lipofectamine 3000 to facilitate ok.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 116
    temperature_celsius: 31
  - step_description: Cells were transferred with fetal bovine serum (fbs) to facilitate
      head.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    replicates: 5
  - step_description: Cells were probed with mg132 proteasome inhibitor to facilitate
      security.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    temperature_celsius: 26
    replicates: 5
  - step_description: Cells were incubated with dapi stain to facilitate education.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    duration_minutes: 645
    replicates: 4
control_groups:
- control_type: Positive Control
  description: Kind despite president style central everyone site enjoy organization
    morning.
data_analysis_methods:
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
- Mass spectrometry data processed with MaxQuant
- Statistical analysis using GraphPad Prism (unpaired t-tests)
reproducibility_notes: All experiments were independently verified by Dr. Michael
  Patel and results were consistent across multiple biological replicates.
