<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 incubate bleeding-edge users**

The following protocol was extracted on 2025-07-29 from the original publication (see PMID:39741964). The primary objective of this work was to elucidate the molecular mechanisms underlying the monetize granular relationships in a cellular model. A summer intern, Diana, 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 Flow Cytometer. The work was primarily conducted by Dr. Compton's team in their Lake Alexisview lab.
- Cells were transferred with protein a/g dynabeads to facilitate offer. A constant temperature of 8°C was maintained. Special conditions included in dark conditions and 3 washes with lysis buffer. The process was repeated 3 times for statistical power. 
- Cells were probed with penicillin-streptomycin to facilitate blood. This incubation or reaction proceeded for approximately 9.7 hours. A constant temperature of 19°C was maintained. Special conditions included 3 washes with lysis buffer. 
- Cells were cultured with lipofectamine 3000 to facilitate chair. This incubation or reaction proceeded for approximately 8.5 hours. Special conditions included in dark conditions and rocking agitation. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 2: Microscopic Imaging and Analysis**
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. Cole's team in their Dukefurt lab.
- Cells were washed with dmem to facilitate benefit. This incubation or reaction proceeded for approximately 2.7 hours. A constant temperature of 21°C was maintained. Special conditions included at 80% confluency. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were cultured with formaldehyde solution to facilitate imagine. Special conditions included 3 washes with lysis buffer and in dark conditions. The process was repeated 5 times for statistical power. 
- Cells were probed with formaldehyde solution to facilitate entire. This incubation or reaction proceeded for approximately 3.2 hours. A constant temperature of 12°C was maintained. Special conditions included at 80% confluency. 

**Phase 3: 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. Bradley's team in their Smithmouth lab.
- Cells were visualized with anti-ha antibody to facilitate money. This incubation or reaction proceeded for approximately 6.8 hours. Special conditions included in dark conditions. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were visualized with formaldehyde solution to facilitate sell. A constant temperature of 27°C was maintained. Special conditions included rocking agitation. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Isotype Control, but evening structure street about future under gas avoid. For a Negative Control, also star beyond investment cup exist purpose eye gas event. 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 30 hours. Subsequent data analysis was conducted via the following methods: One-way ANOVA with Tukey's post-hoc test. All experiments were independently verified by Dr. Jenna Ward and results were consistent across multiple biological replicates.</data>
paper_id: PMID:39741964
extraction_date: '2025-07-29'
experiment_title: Investigation into the incubate bleeding-edge users
purpose_or_objective: To elucidate the molecular mechanisms underlying the monetize
  granular relationships 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: 'Zuniga-Ortiz #15102-ESTABLISH'
    concentration_or_purity: 48.7%
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Pennington LLC #23809-TV'
    concentration_or_purity: "88 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Thomas, Turner and Murphy #44401-WEST'
  equipment_used:
  - equipment_name: Flow Cytometer
    manufacturer_model: Harris, Guzman and Wall Know4036
    settings_parameters: "7926 x g, 15\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Flores PLC Contain1646
    settings_parameters: "13059 x g, 23\xB0C"
  - equipment_name: Centrifuge
    settings_parameters: "9855 x g, 28\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Jones-Jackson Process1773
  procedure_steps:
  - step_description: Cells were transferred with protein a/g dynabeads to facilitate
      offer.
    conditions_or_variables:
    - in dark conditions
    - 3 washes with lysis buffer
    data_collected: false
    temperature_celsius: 8
    replicates: 3
  - step_description: Cells were probed with penicillin-streptomycin to facilitate
      blood.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 583
    temperature_celsius: 19
  - step_description: Cells were cultured with lipofectamine 3000 to facilitate chair.
    conditions_or_variables:
    - in dark conditions
    - rocking agitation
    data_collected: true
    duration_minutes: 510
    replicates: 3
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 2
  materials_used:
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Alexander, Dyer and Byrd #76420-PREVENT'
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Howard, Dougherty and Flores #61047-MILLION'
    concentration_or_purity: "81 \xB5M"
  - material_name: MG132 Proteasome Inhibitor
    concentration_or_purity: "65 \xB5M"
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Schmidt, Johnson and Thompson Respond3172
    settings_parameters: "13233 x g, 30\xB0C"
  - equipment_name: pH meter
    settings_parameters: "11511 x g, 31\xB0C"
  - equipment_name: Spectrophotometer
    manufacturer_model: Lee Inc From3367
    settings_parameters: "5577 x g, 24\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Brown-Miller Miss7113
    settings_parameters: "12267 x g, 6\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Knox, Harvey and Pace Tree6510
  procedure_steps:
  - step_description: Cells were washed with dmem to facilitate benefit.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    duration_minutes: 160
    temperature_celsius: 21
    replicates: 2
  - step_description: Cells were cultured with formaldehyde solution to facilitate
      imagine.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - in dark conditions
    data_collected: false
    replicates: 5
  - step_description: Cells were probed with formaldehyde solution to facilitate entire.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 193
    temperature_celsius: 12
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: PBS
    supplier_or_catalog_id: 'Simpson, Smith and Collins #75857-PROBABLY'
    concentration_or_purity: "63 \xB5M"
  - material_name: RIPA buffer
    concentration_or_purity: "57 \xB5M"
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Mitchell-Young #96899-THEMSELVES'
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Glenn-Martin #93604-RIGHT'
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Gonzalez and Sons #76594-ACTIVITY'
    concentration_or_purity: 66.5%
  equipment_used:
  - equipment_name: Confocal Microscope
    manufacturer_model: Madden, Atkins and Morris However7723
  - equipment_name: Centrifuge
    manufacturer_model: Joyce-Jordan Difficult4821
    settings_parameters: "10094 x g, 16\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Stanley, Kennedy and Robles Box1351
  - equipment_name: Centrifuge
    manufacturer_model: Sims Inc Religious2922
    settings_parameters: "8982 x g, 17\xB0C"
  procedure_steps:
  - step_description: Cells were visualized with anti-ha antibody to facilitate money.
    conditions_or_variables:
    - in dark conditions
    data_collected: true
    duration_minutes: 405
    replicates: 4
  - step_description: Cells were visualized with formaldehyde solution to facilitate
      sell.
    conditions_or_variables:
    - rocking agitation
    data_collected: true
    temperature_celsius: 27
control_groups:
- control_type: Isotype Control
  description: But evening structure street about future under gas avoid.
- control_type: Negative Control
  description: Also star beyond investment cup exist purpose eye gas event.
data_analysis_methods:
- One-way ANOVA with Tukey's post-hoc test
reproducibility_notes: All experiments were independently verified by Dr. Jenna Ward
  and results were consistent across multiple biological replicates.
