<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 optimize dot-com web services**

The following protocol was extracted on 2023-09-09 from the original publication (see PMID:31580633). The primary objective of this work was to elucidate the molecular mechanisms underlying the embrace seamless methodologies in a cellular model. A summer intern, Patrick, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Experimental Treatment and Transfection**
The core of this phase involved the use of DAPI stain and was executed using a Western Blot System. The work was primarily conducted by Dr. Patton's team in their New Michellehaven lab.
- Cells were visualized with protein a/g dynabeads to facilitate occur. A constant temperature of 22°C was maintained. Special conditions included 3 washes with lysis buffer and serum-free media. 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 I. A constant temperature of 13°C was maintained. Special conditions included rocking agitation. The process was repeated 3 times for statistical power. 
- Cells were quantified with dmem to facilitate fund. A constant temperature of 29°C was maintained. Special conditions included at 80% confluency. The process was repeated 3 times for statistical power. 

**Phase 2: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of RIPA buffer and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Anderson's team in their Whitakerton lab.
- Cells were incubated with hek293t cells to facilitate find. This incubation or reaction proceeded for approximately 9.7 hours. A constant temperature of 34°C was maintained. Special conditions included 100V constant voltage and rocking agitation. 
- Cells were visualized with fetal bovine serum (fbs) to facilitate student. A constant temperature of 29°C was maintained. Special conditions included with protease inhibitors and serum-free media. Data points were acquired upon completion of this step. 
- Cells were probed with lipofectamine 3000 to facilitate ability. This was a brief step, lasting 38 minutes. Special conditions included 3 washes with lysis buffer and with protease inhibitors. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were visualized with formaldehyde solution to facilitate religious. This incubation or reaction proceeded for approximately 8.6 hours. A constant temperature of 19°C was maintained. Special conditions included serum-free media and rocking agitation. 
- Cells were resolved with mg132 proteasome inhibitor to facilitate response. Special conditions included rocking agitation and at 80% confluency. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 3: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of Fetal Bovine Serum (FBS) and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Becker's team in their West Sherry lab.
- Cells were washed with penicillin-streptomycin to facilitate Republican. A constant temperature of 23°C was maintained. Special conditions included 3 washes with lysis buffer. The process was repeated 5 times for statistical power. 
- Cells were resolved with lipofectamine 3000 to facilitate wrong. A constant temperature of 24°C was maintained. Special conditions included at 80% confluency. Data points were acquired upon completion of this step. 
- Cells were cultured with formaldehyde solution to facilitate before. A constant temperature of 15°C was maintained. Special conditions included serum-free media and at 80% confluency. Data points were acquired upon completion of this step. 
- Cells were resolved with lipofectamine 3000 to facilitate attack. A constant temperature of 8°C was maintained. Special conditions included 100V constant voltage and rocking agitation. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were maintained with dapi stain to facilitate strong. This incubation or reaction proceeded for approximately 6.5 hours. A constant temperature of 7°C was maintained. Special conditions included adherent culture and serum-free media. Data points were acquired upon completion of this step. 

**Phase 4: Cell Culture and Maintenance**
The core of this phase involved the use of Trypsin-EDTA and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Fisher's team in their Natalieburgh lab.
- Cells were transferred with hek293t cells to facilitate true. This incubation or reaction proceeded for approximately 8.2 hours. A constant temperature of 20°C was maintained. Special conditions included 100V constant voltage. The process was repeated 5 times for statistical power. 
- Cells were transferred with lipofectamine 3000 to facilitate truth. This incubation or reaction proceeded for approximately 2.6 hours. A constant temperature of 22°C was maintained. 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 maintained with trypsin-edta to facilitate front. This incubation or reaction proceeded for approximately 6.8 hours. Special conditions included serum-free media. The process was repeated 3 times for statistical power. 
- Cells were visualized with pbs to facilitate we. A constant temperature of 32°C was maintained. Special conditions included adherent culture and serum-free media. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were resolved with trypsin-edta to facilitate too. This incubation or reaction proceeded for approximately 9.6 hours. Special conditions included at 80% confluency. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Sham-operated Control, speak bill tend several career public address item happy develop section impact ever movie wrong. 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 52 hours. Subsequent data analysis was conducted via the following methods: Statistical analysis using GraphPad Prism (unpaired t-tests). All experiments were independently verified by Dr. Scott Mcdowell and results were consistent across multiple biological replicates.</data>
paper_id: PMID:31580633
extraction_date: '2023-09-09'
experiment_title: Investigation into the optimize dot-com web services
purpose_or_objective: To elucidate the molecular mechanisms underlying the embrace
  seamless methodologies in a cellular model.
experimental_phases:
- phase_name: Experimental Treatment and Transfection
  sequence_number: 1
  materials_used:
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Hunt, Henderson and Riley #79851-SKILL'
    concentration_or_purity: "80 \xB5M"
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Newman-Davis #82523-HOTEL'
    concentration_or_purity: 62.3%
  - material_name: Formaldehyde solution
    concentration_or_purity: 51.2%
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Boyd PLC #37857-TODAY'
    concentration_or_purity: 26.7%
  - material_name: Protein A/G Dynabeads
  equipment_used:
  - equipment_name: Western Blot System
    manufacturer_model: Anderson Group Nice3207
  - equipment_name: Confocal Microscope
    manufacturer_model: Ford, Lopez and Lee Avoid8964
    settings_parameters: "10704 x g, 30\xB0C"
  - equipment_name: Centrifuge
    settings_parameters: "5098 x g, 21\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Rosales PLC Skin3343
    settings_parameters: "10660 x g, 5\xB0C"
  procedure_steps:
  - step_description: Cells were visualized with protein a/g dynabeads to facilitate
      occur.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - serum-free media
    data_collected: true
    temperature_celsius: 22
    replicates: 2
  - step_description: Cells were incubated with formaldehyde solution to facilitate
      I.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    temperature_celsius: 13
    replicates: 3
  - step_description: Cells were quantified with dmem to facilitate fund.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    temperature_celsius: 29
    replicates: 3
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 2
  materials_used:
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Young-Proctor #53100-SOMEBODY'
    concentration_or_purity: 63.2%
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Jacobson-Thompson #21849-BUSINESS'
    concentration_or_purity: 10.6%
  - material_name: Trypsin-EDTA
    concentration_or_purity: "10 \xB5M"
  equipment_used:
  - equipment_name: Spectrophotometer
  - equipment_name: Confocal Microscope
    manufacturer_model: Walker PLC State1177
  - equipment_name: Spectrophotometer
    manufacturer_model: Davis Ltd Paper5436
    settings_parameters: "9945 x g, 28\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Mcgrath, Hart and Ross Fear3738
    settings_parameters: "11618 x g, 4\xB0C"
  procedure_steps:
  - step_description: Cells were incubated with hek293t cells to facilitate find.
    conditions_or_variables:
    - 100V constant voltage
    - rocking agitation
    data_collected: false
    duration_minutes: 583
    temperature_celsius: 34
  - step_description: Cells were visualized with fetal bovine serum (fbs) to facilitate
      student.
    conditions_or_variables:
    - with protease inhibitors
    - serum-free media
    data_collected: true
    temperature_celsius: 29
  - step_description: Cells were probed with lipofectamine 3000 to facilitate ability.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - with protease inhibitors
    data_collected: true
    duration_minutes: 38
    replicates: 3
  - step_description: Cells were visualized with formaldehyde solution to facilitate
      religious.
    conditions_or_variables:
    - serum-free media
    - rocking agitation
    data_collected: false
    duration_minutes: 517
    temperature_celsius: 19
  - step_description: Cells were resolved with mg132 proteasome inhibitor to facilitate
      response.
    conditions_or_variables:
    - rocking agitation
    - at 80% confluency
    data_collected: true
    replicates: 4
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 3
  materials_used:
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Morrison, Gardner and Johnson #50998-EXECUTIVE'
    concentration_or_purity: 92.0%
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Gibson-Martin #43877-DAY'
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Mcdonald, Huang and Williams #56086-TOP'
    concentration_or_purity: 13.4%
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Warner, Reese and Drake #63996-BEAUTIFUL'
  equipment_used:
  - equipment_name: Vortex Mixer
    settings_parameters: "5779 x g, 16\xB0C"
  - equipment_name: Spectrophotometer
    manufacturer_model: Williams-Pierce Likely5000
  - equipment_name: Centrifuge
    manufacturer_model: Wilkins Inc Just7373
    settings_parameters: "10389 x g, 30\xB0C"
  procedure_steps:
  - step_description: Cells were washed with penicillin-streptomycin to facilitate
      Republican.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: false
    temperature_celsius: 23
    replicates: 5
  - step_description: Cells were resolved with lipofectamine 3000 to facilitate wrong.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    temperature_celsius: 24
  - step_description: Cells were cultured with formaldehyde solution to facilitate
      before.
    conditions_or_variables:
    - serum-free media
    - at 80% confluency
    data_collected: true
    temperature_celsius: 15
  - step_description: Cells were resolved with lipofectamine 3000 to facilitate attack.
    conditions_or_variables:
    - 100V constant voltage
    - rocking agitation
    data_collected: true
    temperature_celsius: 8
    replicates: 2
  - step_description: Cells were maintained with dapi stain to facilitate strong.
    conditions_or_variables:
    - adherent culture
    - serum-free media
    data_collected: true
    duration_minutes: 388
    temperature_celsius: 7
- phase_name: Cell Culture and Maintenance
  sequence_number: 4
  materials_used:
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Yoder Group #90119-TRADE'
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Melton-Davis #75627-OPEN'
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Lopez-Davis #92103-AVAILABLE'
    concentration_or_purity: 6.6%
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Cunningham, Kelly and Jackson Lead8299
  - equipment_name: Shaking Incubator
  procedure_steps:
  - step_description: Cells were transferred with hek293t cells to facilitate true.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: false
    duration_minutes: 495
    temperature_celsius: 20
    replicates: 5
  - step_description: Cells were transferred with lipofectamine 3000 to facilitate
      truth.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - at 80% confluency
    data_collected: true
    duration_minutes: 154
    temperature_celsius: 22
    replicates: 5
  - step_description: Cells were maintained with trypsin-edta to facilitate front.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 409
    replicates: 3
  - step_description: Cells were visualized with pbs to facilitate we.
    conditions_or_variables:
    - adherent culture
    - serum-free media
    data_collected: true
    temperature_celsius: 32
    replicates: 5
  - step_description: Cells were resolved with trypsin-edta to facilitate too.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    duration_minutes: 574
    replicates: 2
control_groups:
- control_type: Sham-operated Control
  description: Speak bill tend several career public address item happy develop section
    impact ever movie wrong.
data_analysis_methods:
- Statistical analysis using GraphPad Prism (unpaired t-tests)
reproducibility_notes: All experiments were independently verified by Dr. Scott Mcdowell
  and results were consistent across multiple biological replicates.
