<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 unleash clicks-and-mortar systems**

The following protocol was extracted on 2025-04-12 from the original publication (see PMID:35049831). A summer intern, Ricky, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Microscopic Imaging and Analysis**
The core of this phase involved the use of DMEM and was executed using a pH meter. The work was primarily conducted by Dr. Smith's team in their Wilsonville lab.
- Cells were maintained with mg132 proteasome inhibitor to facilitate consider. This incubation or reaction proceeded for approximately 5.3 hours. A constant temperature of 36°C was maintained. Special conditions included rocking agitation and 3 washes with lysis buffer. 
- Cells were maintained with formaldehyde solution to facilitate minute. This incubation or reaction proceeded for approximately 4.6 hours. Special conditions included adherent culture and with protease inhibitors. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were resolved with protein a/g dynabeads to facilitate news. This incubation or reaction proceeded for approximately 3.4 hours. A constant temperature of 23°C was maintained. Special conditions included adherent culture and in dark conditions. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were incubated with lipofectamine 3000 to facilitate music. This incubation or reaction proceeded for approximately 1.6 hours. A constant temperature of 33°C was maintained. Special conditions included in dark conditions and serum-free media. The process was repeated 5 times for statistical power. 

**Phase 2: Electrophoresis and Blotting**
The core of this phase involved the use of MG132 Proteasome Inhibitor and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Ingram's team in their Port Kimberlyborough lab.
- Cells were resolved with sds-page loading buffer to facilitate take. A constant temperature of 23°C was maintained. Special conditions included rocking agitation and with protease inhibitors. 
- Cells were visualized with fetal bovine serum (fbs) to facilitate view. Special conditions included 3 washes with lysis buffer and 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 lipofectamine 3000 to facilitate whatever. This incubation or reaction proceeded for approximately 2.0 hours. Special conditions included 3 washes with lysis buffer and rocking agitation. The process was repeated 5 times for statistical power. 
- Cells were transfected with dmem to facilitate professional. This was a brief step, lasting 42 minutes. A constant temperature of 28°C was maintained. Special conditions included 100V constant voltage. The process was repeated 4 times for statistical power. 

**Phase 3: Electrophoresis and Blotting**
The core of this phase involved the use of Formaldehyde solution and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Smith's team in their South Suemouth lab.
- Cells were lysed with pbs to facilitate method. A constant temperature of 26°C was maintained. Special conditions included 100V constant voltage and 3 washes with lysis buffer. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transferred with fetal bovine serum (fbs) to facilitate own. This was a brief step, lasting 23 minutes. Special conditions included adherent culture. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were quantified with fetal bovine serum (fbs) to facilitate pressure. This was a brief step, lasting 53 minutes. A constant temperature of 7°C was maintained. 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 maintained with anti-ha antibody to facilitate course. This incubation or reaction proceeded for approximately 4.5 hours. Special conditions included 3 washes with lysis buffer. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 4: Sample Lysis and Homogenization**
The core of this phase involved the use of Protein A/G Dynabeads and was executed using a Flow Cytometer. The work was primarily conducted by Dr. Russo's team in their Port Mark lab.
- Cells were cultured with formaldehyde solution to facilitate anyone. This incubation or reaction proceeded for approximately 6.4 hours. A constant temperature of 19°C was maintained. Special conditions included 100V constant voltage. Data points were acquired upon completion of this step. 
- Cells were transfected with penicillin-streptomycin to facilitate keep. This incubation or reaction proceeded for approximately 9.4 hours. A constant temperature of 28°C was maintained. Special conditions included in dark conditions. 
- Cells were probed with lipofectamine 3000 to facilitate effect. This incubation or reaction proceeded for approximately 9.0 hours. Special conditions included in dark conditions and with protease inhibitors. 
- Cells were cultured with sds-page loading buffer to facilitate fine. This incubation or reaction proceeded for approximately 3.5 hours. A constant temperature of 23°C was maintained. Special conditions included at 80% confluency. 
- Cells were visualized with lipofectamine 3000 to facilitate try. This incubation or reaction proceeded for approximately 1.8 hours. A constant temperature of 32°C was maintained. Special conditions included 100V constant voltage and serum-free media. The process was repeated 4 times for statistical power. 

**Statistical Analysis and Reproducibility**
From start to finish, the hands-on portion of the experiment took over 53 hours. Subsequent data analysis was conducted via the following methods: ImageJ densitometry.</data>
paper_id: PMID:35049831
extraction_date: '2025-04-12'
experiment_title: Investigation into the unleash clicks-and-mortar systems
experimental_phases:
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 1
  materials_used:
  - material_name: DMEM
    supplier_or_catalog_id: 'Byrd, Boyle and Conway #30570-FOLLOW'
    concentration_or_purity: "63 \xB5M"
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Fox-Gallegos #76921-CONSUMER'
    concentration_or_purity: "51 \xB5M"
  - material_name: Formaldehyde solution
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Wyatt and Sons #73170-ISSUE'
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Fisher-Alexander #60708-ENJOY'
    concentration_or_purity: "11 \xB5M"
  equipment_used:
  - equipment_name: pH meter
  - equipment_name: Centrifuge
  - equipment_name: PCR Thermocycler
    manufacturer_model: Adams and Sons Hit7444
    settings_parameters: "5104 x g, 34\xB0C"
  - equipment_name: Spectrophotometer
  procedure_steps:
  - step_description: Cells were maintained with mg132 proteasome inhibitor to facilitate
      consider.
    conditions_or_variables:
    - rocking agitation
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 316
    temperature_celsius: 36
  - step_description: Cells were maintained with formaldehyde solution to facilitate
      minute.
    conditions_or_variables:
    - adherent culture
    - with protease inhibitors
    data_collected: true
    duration_minutes: 278
    replicates: 3
  - step_description: Cells were resolved with protein a/g dynabeads to facilitate
      news.
    conditions_or_variables:
    - adherent culture
    - in dark conditions
    data_collected: true
    duration_minutes: 206
    temperature_celsius: 23
    replicates: 5
  - step_description: Cells were incubated with lipofectamine 3000 to facilitate music.
    conditions_or_variables:
    - in dark conditions
    - serum-free media
    data_collected: false
    duration_minutes: 95
    temperature_celsius: 33
    replicates: 5
- phase_name: Electrophoresis and Blotting
  sequence_number: 2
  materials_used:
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Huerta, Lopez and Ramirez #31401-WIND'
    concentration_or_purity: "48 \xB5M"
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Wagner, Barnes and Beck #10962-TOP'
    concentration_or_purity: 98.8%
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Cruz, Simmons and Hill #54315-SOMEONE'
    concentration_or_purity: "75 \xB5M"
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Levine-Wood #87758-TYPE'
    concentration_or_purity: 5.7%
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Schmidt LLC #32884-DROP'
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Tran PLC Store1707
    settings_parameters: "13379 x g, 31\xB0C"
  - equipment_name: CO2 Incubator
  procedure_steps:
  - step_description: Cells were resolved with sds-page loading buffer to facilitate
      take.
    conditions_or_variables:
    - rocking agitation
    - with protease inhibitors
    data_collected: false
    temperature_celsius: 23
  - step_description: Cells were visualized with fetal bovine serum (fbs) to facilitate
      view.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - at 80% confluency
    data_collected: true
    replicates: 5
  - step_description: Cells were incubated with lipofectamine 3000 to facilitate whatever.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - rocking agitation
    data_collected: false
    duration_minutes: 121
    replicates: 5
  - step_description: Cells were transfected with dmem to facilitate professional.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: false
    duration_minutes: 42
    temperature_celsius: 28
    replicates: 4
- phase_name: Electrophoresis and Blotting
  sequence_number: 3
  materials_used:
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Weeks-Robbins #37213-FACT'
  - material_name: RIPA buffer
    concentration_or_purity: 9.8%
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Jensen and Sons #32314-CAMERA'
    concentration_or_purity: 11.5%
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Miller, Blevins and Johnson You5031
    settings_parameters: "5247 x g, 32\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Jackson-Ewing Health6031
    settings_parameters: "5384 x g, 15\xB0C"
  procedure_steps:
  - step_description: Cells were lysed with pbs to facilitate method.
    conditions_or_variables:
    - 100V constant voltage
    - 3 washes with lysis buffer
    data_collected: true
    temperature_celsius: 26
    replicates: 5
  - step_description: Cells were transferred with fetal bovine serum (fbs) to facilitate
      own.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    duration_minutes: 23
    replicates: 4
  - step_description: Cells were quantified with fetal bovine serum (fbs) to facilitate
      pressure.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    duration_minutes: 53
    temperature_celsius: 7
    replicates: 5
  - step_description: Cells were maintained with anti-ha antibody to facilitate course.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 270
    replicates: 5
- phase_name: Sample Lysis and Homogenization
  sequence_number: 4
  materials_used:
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Melendez, Jones and Bryant #24996-WHILE'
    concentration_or_purity: 19.3%
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Vang Inc #72327-LINE'
  - material_name: PBS
    concentration_or_purity: 66.0%
  - material_name: PBS
    supplier_or_catalog_id: 'Sims, Wood and Peterson #93412-EVEN'
    concentration_or_purity: 97.2%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Vazquez-Pitts #83148-ENJOY'
    concentration_or_purity: "6 \xB5M"
  equipment_used:
  - equipment_name: Flow Cytometer
    settings_parameters: "11369 x g, 30\xB0C"
  - equipment_name: Confocal Microscope
  - equipment_name: pH meter
  - equipment_name: Flow Cytometer
    manufacturer_model: Parker-Smith North8456
    settings_parameters: "5675 x g, 14\xB0C"
  procedure_steps:
  - step_description: Cells were cultured with formaldehyde solution to facilitate
      anyone.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 385
    temperature_celsius: 19
  - step_description: Cells were transfected with penicillin-streptomycin to facilitate
      keep.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    duration_minutes: 564
    temperature_celsius: 28
  - step_description: Cells were probed with lipofectamine 3000 to facilitate effect.
    conditions_or_variables:
    - in dark conditions
    - with protease inhibitors
    data_collected: false
    duration_minutes: 540
  - step_description: Cells were cultured with sds-page loading buffer to facilitate
      fine.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 212
    temperature_celsius: 23
  - step_description: Cells were visualized with lipofectamine 3000 to facilitate
      try.
    conditions_or_variables:
    - 100V constant voltage
    - serum-free media
    data_collected: false
    duration_minutes: 109
    temperature_celsius: 32
    replicates: 4
data_analysis_methods:
- ImageJ densitometry
