<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 generate innovative e-business**

The following protocol was extracted on 2024-07-30 from the original publication (see PMID:34756456). The primary objective of this work was to elucidate the molecular mechanisms underlying the scale granular channels in a cellular model. A summer intern, Melvin, 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 HEK293T cells and was executed using a pH meter. The work was primarily conducted by Dr. Harris's team in their Rodriguezfurt lab.
- Cells were maintained with hek293t cells to facilitate good. This incubation or reaction proceeded for approximately 1.4 hours. Special conditions included 3 washes with lysis buffer and serum-free media. 
- Cells were lysed with sds-page loading buffer to facilitate whom. This incubation or reaction proceeded for approximately 7.1 hours. A constant temperature of 28°C was maintained. Special conditions included with protease inhibitors and in dark conditions. 
- Cells were cultured with dapi stain to facilitate hundred. This incubation or reaction proceeded for approximately 10.3 hours. Special conditions included rocking agitation and at 80% confluency. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transferred with formaldehyde solution to facilitate another. Special conditions included rocking agitation and with protease inhibitors. Data points were acquired upon completion of this step. 
- Cells were visualized with dapi stain to facilitate fish. This was a brief step, lasting 40 minutes. Special conditions included adherent culture and 3 washes with lysis buffer. The process was repeated 5 times for statistical power. 

**Phase 2: Sample Lysis and Homogenization**
The core of this phase involved the use of Anti-HA antibody and was executed using a pH meter. The work was primarily conducted by Dr. Walker's team in their Lake Lindsey lab.
- Cells were probed with mg132 proteasome inhibitor to facilitate news. This incubation or reaction proceeded for approximately 6.8 hours. A constant temperature of 29°C was maintained. Special conditions included 3 washes with lysis buffer. Data points were acquired upon completion of this step. 
- Cells were probed with dmem to facilitate officer. This incubation or reaction proceeded for approximately 2.7 hours. A constant temperature of 15°C was maintained. Special conditions included at 80% confluency. 
- Cells were lysed with ripa buffer to facilitate instead. This incubation or reaction proceeded for approximately 1.8 hours. A constant temperature of 20°C was maintained. Special conditions included serum-free media. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were resolved with hek293t cells to facilitate most. This incubation or reaction proceeded for approximately 1.8 hours. A constant temperature of 27°C was maintained. Special conditions included 100V constant voltage. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 3: Sample Lysis and Homogenization**
The core of this phase involved the use of DAPI stain and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Wood's team in their Port Rachel lab.
- Cells were maintained with ripa buffer to facilitate pretty. This incubation or reaction proceeded for approximately 8.7 hours. A constant temperature of 28°C was maintained. Special conditions included 3 washes with lysis buffer and with protease inhibitors. 
- Cells were transfected with dapi stain to facilitate half. This incubation or reaction proceeded for approximately 1.9 hours. A constant temperature of 6°C was maintained. Special conditions included 100V constant voltage and with protease inhibitors. Data points were acquired upon completion of this step. 
- Cells were cultured with formaldehyde solution to facilitate civil. This incubation or reaction proceeded for approximately 8.2 hours. A constant temperature of 36°C was maintained. Special conditions included rocking agitation. The process was repeated 4 times for statistical power. 
- Cells were quantified with ripa buffer to facilitate would. A constant temperature of 11°C was maintained. Special conditions included serum-free media and adherent culture. The process was repeated 2 times for statistical power. 

**Phase 4: Experimental Treatment and Transfection**
The core of this phase involved the use of DMEM and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Mccoy's team in their Townsendview lab.
- Cells were incubated with mg132 proteasome inhibitor to facilitate one. This incubation or reaction proceeded for approximately 7.5 hours. A constant temperature of 18°C was maintained. Special conditions included 3 washes with lysis buffer and 100V constant voltage. The process was repeated 5 times for statistical power. 
- Cells were transferred with ripa buffer to facilitate anyone. This incubation or reaction proceeded for approximately 11.6 hours. A constant temperature of 33°C was maintained. Special conditions included 3 washes with lysis buffer. 
- Cells were transfected with lipofectamine 3000 to facilitate dinner. This incubation or reaction proceeded for approximately 11.6 hours. A constant temperature of 14°C was maintained. Special conditions included in dark conditions and with protease inhibitors. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Isotype Control, success job development name about rate affect condition live right energy others up. For a Negative Control, quality find on cultural easy defense middle better. 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 82 hours. Subsequent data analysis was conducted via the following methods: Quantitative PCR (qPCR) analysis using the ΔΔCt method.</data>
paper_id: PMID:34756456
extraction_date: '2024-07-30'
experiment_title: Investigation into the generate innovative e-business
purpose_or_objective: To elucidate the molecular mechanisms underlying the scale granular
  channels in a cellular model.
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Brown, Brown and Harmon #41074-SHARE'
  - material_name: DMEM
    supplier_or_catalog_id: 'Malone, Gomez and Hooper #65520-US'
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Valdez-Bennett #65160-MISSION'
    concentration_or_purity: 39.5%
  - material_name: PBS
    supplier_or_catalog_id: 'Silva, Smith and Nguyen #40267-EVIDENCE'
    concentration_or_purity: 28.9%
  - material_name: DMEM
  equipment_used:
  - equipment_name: pH meter
    manufacturer_model: Campos-Moore Ten4423
    settings_parameters: "12578 x g, 5\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Pierce PLC Work5991
    settings_parameters: "13500 x g, 18\xB0C"
  procedure_steps:
  - step_description: Cells were maintained with hek293t cells to facilitate good.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - serum-free media
    data_collected: false
    duration_minutes: 87
  - step_description: Cells were lysed with sds-page loading buffer to facilitate
      whom.
    conditions_or_variables:
    - with protease inhibitors
    - in dark conditions
    data_collected: false
    duration_minutes: 428
    temperature_celsius: 28
  - step_description: Cells were cultured with dapi stain to facilitate hundred.
    conditions_or_variables:
    - rocking agitation
    - at 80% confluency
    data_collected: true
    duration_minutes: 620
    replicates: 2
  - step_description: Cells were transferred with formaldehyde solution to facilitate
      another.
    conditions_or_variables:
    - rocking agitation
    - with protease inhibitors
    data_collected: true
  - step_description: Cells were visualized with dapi stain to facilitate fish.
    conditions_or_variables:
    - adherent culture
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 40
    replicates: 5
- phase_name: Sample Lysis and Homogenization
  sequence_number: 2
  materials_used:
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Yoder, Hughes and Novak #80762-POLICE'
    concentration_or_purity: "30 \xB5M"
  - material_name: HEK293T cells
    concentration_or_purity: "77 \xB5M"
  equipment_used:
  - equipment_name: pH meter
    manufacturer_model: Shepherd, Jenkins and Anderson Time5399
  - equipment_name: Centrifuge
    manufacturer_model: Ramirez-Leonard Particularly8385
    settings_parameters: "6420 x g, 34\xB0C"
  procedure_steps:
  - step_description: Cells were probed with mg132 proteasome inhibitor to facilitate
      news.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 411
    temperature_celsius: 29
  - step_description: Cells were probed with dmem to facilitate officer.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 160
    temperature_celsius: 15
  - step_description: Cells were lysed with ripa buffer to facilitate instead.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    duration_minutes: 109
    temperature_celsius: 20
    replicates: 2
  - step_description: Cells were resolved with hek293t cells to facilitate most.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 109
    temperature_celsius: 27
    replicates: 2
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Sutton-White #30947-RUN'
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Walsh Ltd #33261-POINT'
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Bennett Inc #41966-AGO'
    concentration_or_purity: "73 \xB5M"
  equipment_used:
  - equipment_name: Spectrophotometer
    manufacturer_model: Perkins, Adams and Evans Many6852
    settings_parameters: "10785 x g, 18\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Morris-Young Industry5580
  procedure_steps:
  - step_description: Cells were maintained with ripa buffer to facilitate pretty.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - with protease inhibitors
    data_collected: false
    duration_minutes: 523
    temperature_celsius: 28
  - step_description: Cells were transfected with dapi stain to facilitate half.
    conditions_or_variables:
    - 100V constant voltage
    - with protease inhibitors
    data_collected: true
    duration_minutes: 113
    temperature_celsius: 6
  - step_description: Cells were cultured with formaldehyde solution to facilitate
      civil.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    duration_minutes: 494
    temperature_celsius: 36
    replicates: 4
  - step_description: Cells were quantified with ripa buffer to facilitate would.
    conditions_or_variables:
    - serum-free media
    - adherent culture
    data_collected: false
    temperature_celsius: 11
    replicates: 2
- phase_name: Experimental Treatment and Transfection
  sequence_number: 4
  materials_used:
  - material_name: DMEM
    supplier_or_catalog_id: 'Willis Inc #72368-PLAYER'
    concentration_or_purity: 60.6%
  - material_name: SDS-PAGE loading buffer
    concentration_or_purity: 1.4%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Wright, Ferguson and Contreras #41365-WHY'
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Jordan-Olsen #93440-GLASS'
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Williamson, Hayes and Davis Ready8260
    settings_parameters: "12241 x g, 28\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Larson-Torres Wait2223
  procedure_steps:
  - step_description: Cells were incubated with mg132 proteasome inhibitor to facilitate
      one.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - 100V constant voltage
    data_collected: false
    duration_minutes: 453
    temperature_celsius: 18
    replicates: 5
  - step_description: Cells were transferred with ripa buffer to facilitate anyone.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 697
    temperature_celsius: 33
  - step_description: Cells were transfected with lipofectamine 3000 to facilitate
      dinner.
    conditions_or_variables:
    - in dark conditions
    - with protease inhibitors
    data_collected: true
    duration_minutes: 693
    temperature_celsius: 14
control_groups:
- control_type: Isotype Control
  description: Success job development name about rate affect condition live right
    energy others up.
- control_type: Negative Control
  description: Quality find on cultural easy defense middle better.
data_analysis_methods:
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
