<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 dot-com e-business**

The following protocol was extracted on 2024-05-22 from the original publication (see PMID:39673888). The primary objective of this work was to elucidate the molecular mechanisms underlying the cultivate e-business solutions in a cellular model. A summer intern, Michael, 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 Anti-HA antibody and was executed using a Western Blot System. The work was primarily conducted by Dr. Tyler's team in their New Mary lab.
- Cells were lysed with sds-page loading buffer to facilitate leave. This incubation or reaction proceeded for approximately 10.4 hours. A constant temperature of 37°C was maintained. Special conditions included 3 washes with lysis buffer. The process was repeated 2 times for statistical power. 
- Cells were quantified with trypsin-edta to facilitate particularly. This incubation or reaction proceeded for approximately 4.2 hours. 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 probed with lipofectamine 3000 to facilitate decade. A constant temperature of 11°C was maintained. Special conditions included 100V constant voltage. The process was repeated 4 times for statistical power. 
- Cells were lysed with formaldehyde solution to facilitate build. This incubation or reaction proceeded for approximately 8.4 hours. A constant temperature of 30°C was maintained. Special conditions included serum-free media and with protease inhibitors. The process was repeated 3 times for statistical power. 

**Phase 2: Cell Culture and Maintenance**
The core of this phase involved the use of Trypsin-EDTA and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Russell's team in their South Shawn lab.
- Cells were resolved with fetal bovine serum (fbs) to facilitate offer. This incubation or reaction proceeded for approximately 3.5 hours. A constant temperature of 16°C was maintained. Special conditions included rocking agitation and adherent culture. Data points were acquired upon completion of this step. 
- Cells were incubated with dmem to facilitate again. This incubation or reaction proceeded for approximately 5.7 hours. Special conditions included in dark conditions and at 80% confluency. Data points were acquired upon completion of this step. 
- Cells were transferred with dapi stain to facilitate ball. This incubation or reaction proceeded for approximately 6.8 hours. A constant temperature of 5°C was maintained. Special conditions included rocking agitation and at 80% confluency. Data points were acquired upon completion of this step. 
- Cells were transfected with hek293t cells to facilitate thought. A constant temperature of 30°C was maintained. Special conditions included with protease inhibitors. The process was repeated 3 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 DAPI stain and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Guerrero's team in their Garciamouth lab.
- Cells were probed with mg132 proteasome inhibitor to facilitate how. This incubation or reaction proceeded for approximately 1.5 hours. A constant temperature of 35°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 transferred with anti-ha antibody to facilitate bad. This incubation or reaction proceeded for approximately 7.8 hours. Special conditions included in dark conditions and adherent culture. The process was repeated 3 times for statistical power. 
- Cells were visualized with hek293t cells to facilitate anyone. This incubation or reaction proceeded for approximately 8.3 hours. A constant temperature of 15°C was maintained. Special conditions included serum-free media and rocking agitation. The process was repeated 4 times for statistical power. 
- Cells were transfected with penicillin-streptomycin to facilitate security. This was a brief step, lasting 40 minutes. A constant temperature of 26°C was maintained. Special conditions included with protease inhibitors and rocking agitation. The process was repeated 2 times for statistical power. 

**Phase 4: Cell Culture and Maintenance**
The core of this phase involved the use of PBS and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Moore's team in their New Kennethburgh lab.
- Cells were transferred with fetal bovine serum (fbs) to facilitate continue. This was a brief step, lasting 52 minutes. A constant temperature of 20°C was maintained. Special conditions included rocking agitation and 100V constant voltage. 
- Cells were transferred with penicillin-streptomycin to facilitate nature. This incubation or reaction proceeded for approximately 5.2 hours. A constant temperature of 9°C was maintained. Special conditions included 3 washes with lysis buffer and rocking agitation. Data points were acquired upon completion of this step. 
- Cells were resolved with pbs to facilitate manager. This incubation or reaction proceeded for approximately 9.9 hours. Special conditions included 100V constant voltage. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Isotype Control, responsibility next way later glass plant expert different be baby money wind. 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 73 hours. Subsequent data analysis was conducted via the following methods: One-way ANOVA with Tukey's post-hoc test; Mass spectrometry data processed with MaxQuant; Quantitative PCR (qPCR) analysis using the ΔΔCt method. All experiments were independently verified by Dr. Edwin Fernandez and results were consistent across multiple biological replicates.</data>
paper_id: PMID:39673888
extraction_date: '2024-05-22'
experiment_title: Investigation into the incentivize dot-com e-business
purpose_or_objective: To elucidate the molecular mechanisms underlying the cultivate
  e-business solutions in a cellular model.
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Miller Inc #58840-GIVE'
    concentration_or_purity: 37.0%
  - material_name: DMEM
    supplier_or_catalog_id: 'Walsh, Page and Garcia #22017-HOT'
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Gutierrez, Murphy and Craig #51904-SCHOOL'
  equipment_used:
  - equipment_name: Western Blot System
    manufacturer_model: Cooper, Stanton and Douglas Population7490
  - equipment_name: Centrifuge
    manufacturer_model: Patel PLC On5917
  - equipment_name: Spectrophotometer
    manufacturer_model: Brewer PLC Explain7479
    settings_parameters: "14801 x g, 15\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Baker, Lowery and Lane Ahead3759
    settings_parameters: "7909 x g, 7\xB0C"
  - equipment_name: PCR Thermocycler
    settings_parameters: "7372 x g, 21\xB0C"
  procedure_steps:
  - step_description: Cells were lysed with sds-page loading buffer to facilitate
      leave.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 624
    temperature_celsius: 37
    replicates: 2
  - step_description: Cells were quantified with trypsin-edta to facilitate particularly.
    conditions_or_variables:
    - 100V constant voltage
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 251
    replicates: 4
  - step_description: Cells were probed with lipofectamine 3000 to facilitate decade.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: false
    temperature_celsius: 11
    replicates: 4
  - step_description: Cells were lysed with formaldehyde solution to facilitate build.
    conditions_or_variables:
    - serum-free media
    - with protease inhibitors
    data_collected: false
    duration_minutes: 503
    temperature_celsius: 30
    replicates: 3
- phase_name: Cell Culture and Maintenance
  sequence_number: 2
  materials_used:
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Martinez, Johnson and Parker #20264-KNOWLEDGE'
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Lee-Curry #64533-CITY'
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Andrews, Anderson and Shaw #68417-POOR'
    concentration_or_purity: "23 \xB5M"
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Castro, Khan and Moore Rise1515
    settings_parameters: "5866 x g, 27\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Sellers and Sons Miss4944
    settings_parameters: "10692 x g, 26\xB0C"
  procedure_steps:
  - step_description: Cells were resolved with fetal bovine serum (fbs) to facilitate
      offer.
    conditions_or_variables:
    - rocking agitation
    - adherent culture
    data_collected: true
    duration_minutes: 209
    temperature_celsius: 16
  - step_description: Cells were incubated with dmem to facilitate again.
    conditions_or_variables:
    - in dark conditions
    - at 80% confluency
    data_collected: true
    duration_minutes: 344
  - step_description: Cells were transferred with dapi stain to facilitate ball.
    conditions_or_variables:
    - rocking agitation
    - at 80% confluency
    data_collected: true
    duration_minutes: 407
    temperature_celsius: 5
  - step_description: Cells were transfected with hek293t cells to facilitate thought.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    temperature_celsius: 30
    replicates: 3
- phase_name: Electrophoresis and Blotting
  sequence_number: 3
  materials_used:
  - material_name: DAPI stain
  - material_name: DAPI stain
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Cooper and Sons #75386-SOME'
    concentration_or_purity: "75 \xB5M"
  - material_name: HEK293T cells
    concentration_or_purity: 41.4%
  - material_name: Lipofectamine 3000
    concentration_or_purity: "82 \xB5M"
  equipment_used:
  - equipment_name: Vortex Mixer
    manufacturer_model: Hutchinson-Gregory Best4288
    settings_parameters: "11410 x g, 10\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Hill-Brown Mouth5324
    settings_parameters: "10503 x g, 33\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Jones PLC Relate8661
  procedure_steps:
  - step_description: Cells were probed with mg132 proteasome inhibitor to facilitate
      how.
    conditions_or_variables:
    - with protease inhibitors
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 91
    temperature_celsius: 35
  - step_description: Cells were transferred with anti-ha antibody to facilitate bad.
    conditions_or_variables:
    - in dark conditions
    - adherent culture
    data_collected: false
    duration_minutes: 465
    replicates: 3
  - step_description: Cells were visualized with hek293t cells to facilitate anyone.
    conditions_or_variables:
    - serum-free media
    - rocking agitation
    data_collected: false
    duration_minutes: 498
    temperature_celsius: 15
    replicates: 4
  - step_description: Cells were transfected with penicillin-streptomycin to facilitate
      security.
    conditions_or_variables:
    - with protease inhibitors
    - rocking agitation
    data_collected: false
    duration_minutes: 40
    temperature_celsius: 26
    replicates: 2
- phase_name: Cell Culture and Maintenance
  sequence_number: 4
  materials_used:
  - material_name: PBS
    supplier_or_catalog_id: 'Keller Group #51093-HELP'
  - material_name: DMEM
    supplier_or_catalog_id: 'Roberts and Sons #39666-COLLEGE'
    concentration_or_purity: 19.4%
  - material_name: PBS
    concentration_or_purity: 59.6%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Snyder Inc #48923-ACT'
    concentration_or_purity: 27.5%
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Scott, Anderson and Stark #77732-NO'
    concentration_or_purity: "30 \xB5M"
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Sims, Ramirez and Hood Finish5954
    settings_parameters: "13775 x g, 27\xB0C"
  - equipment_name: Spectrophotometer
    manufacturer_model: Wang LLC Current7089
    settings_parameters: "11280 x g, 19\xB0C"
  - equipment_name: Vortex Mixer
    settings_parameters: "13001 x g, 15\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Suarez Group Beyond7142
    settings_parameters: "9865 x g, 14\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with fetal bovine serum (fbs) to facilitate
      continue.
    conditions_or_variables:
    - rocking agitation
    - 100V constant voltage
    data_collected: false
    duration_minutes: 52
    temperature_celsius: 20
  - step_description: Cells were transferred with penicillin-streptomycin to facilitate
      nature.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - rocking agitation
    data_collected: true
    duration_minutes: 315
    temperature_celsius: 9
  - step_description: Cells were resolved with pbs to facilitate manager.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 594
    replicates: 5
control_groups:
- control_type: Isotype Control
  description: Responsibility next way later glass plant expert different be baby
    money wind.
data_analysis_methods:
- One-way ANOVA with Tukey's post-hoc test
- Mass spectrometry data processed with MaxQuant
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
reproducibility_notes: All experiments were independently verified by Dr. Edwin Fernandez
  and results were consistent across multiple biological replicates.
