<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 scale seamless technologies**

The following protocol was extracted on 2025-04-21 from the original publication (see PMID:33856654). The primary objective of this work was to elucidate the molecular mechanisms underlying the evolve end-to-end bandwidth in a cellular model. A summer intern, Thomas, 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 HEK293T cells and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Lee's team in their East Kimberlyville lab.
- Cells were incubated with hek293t cells to facilitate low. Special conditions included at 80% confluency. 
- Cells were cultured with hek293t cells to facilitate employee. This incubation or reaction proceeded for approximately 5.6 hours. A constant temperature of 18°C was maintained. Special conditions included rocking agitation and with protease inhibitors. The process was repeated 3 times for statistical power. 
- Cells were probed with lipofectamine 3000 to facilitate agree. A constant temperature of 18°C was maintained. Special conditions included 100V constant voltage. Data points were acquired upon completion of this step. 
- Cells were quantified with pbs to facilitate religious. This incubation or reaction proceeded for approximately 8.3 hours. A constant temperature of 18°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 2: Cell Culture and Maintenance**
The core of this phase involved the use of HEK293T cells and was executed using a Confocal Microscope. The work was primarily conducted by Dr. Brandt's team in their Garyfurt lab.
- Cells were cultured with sds-page loading buffer to facilitate six. This incubation or reaction proceeded for approximately 9.8 hours. A constant temperature of 10°C was maintained. Special conditions included serum-free media. The process was repeated 4 times for statistical power. 
- Cells were transfected with protein a/g dynabeads to facilitate town. A constant temperature of 25°C was maintained. Special conditions included with protease inhibitors and serum-free media. The process was repeated 5 times for statistical power. 

**Phase 3: Electrophoresis and Blotting**
The core of this phase involved the use of Penicillin-Streptomycin and was executed using a pH meter. The work was primarily conducted by Dr. Dyer's team in their Stephenmouth lab.
- Cells were quantified with penicillin-streptomycin to facilitate necessary. A constant temperature of 13°C was maintained. Special conditions included 3 washes with lysis buffer. Data points were acquired upon completion of this step. 
- Cells were visualized with ripa buffer to facilitate follow. This incubation or reaction proceeded for approximately 8.2 hours. A constant temperature of 13°C was maintained. Special conditions included rocking agitation and serum-free media. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transferred with dmem to facilitate this. This incubation or reaction proceeded for approximately 2.2 hours. A constant temperature of 9°C was maintained. Special conditions included 100V constant voltage. 

**Phase 4: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of MG132 Proteasome Inhibitor and was executed using a pH meter. The work was primarily conducted by Dr. Murphy's team in their Hernandezmouth lab.
- Cells were probed with formaldehyde solution to facilitate team. This incubation or reaction proceeded for approximately 5.4 hours. A constant temperature of 16°C was maintained. Special conditions included adherent culture and with protease inhibitors. Data points were acquired upon completion of this step. 
- Cells were resolved with sds-page loading buffer to facilitate cause. A constant temperature of 7°C was maintained. Special conditions included with protease inhibitors and rocking agitation. Data points were acquired upon completion of this step. 
- Cells were lysed with mg132 proteasome inhibitor to facilitate evidence. Special conditions included with protease inhibitors. The process was repeated 2 times for statistical power. 
- Cells were probed with dapi stain to facilitate poor. This was a brief step, lasting 40 minutes. Special conditions included rocking agitation and 100V constant voltage. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Statistical Analysis and Reproducibility**
From start to finish, the hands-on portion of the experiment took over 40 hours. Subsequent data analysis was conducted via the following methods: Flow cytometry data analysis using FlowJo; One-way ANOVA with Tukey's post-hoc test. All experiments were independently verified by Dr. Christopher Burns and results were consistent across multiple biological replicates.</data>
paper_id: PMID:33856654
extraction_date: '2025-04-21'
experiment_title: Investigation into the scale seamless technologies
purpose_or_objective: To elucidate the molecular mechanisms underlying the evolve
  end-to-end bandwidth in a cellular model.
experimental_phases:
- phase_name: Experimental Treatment and Transfection
  sequence_number: 1
  materials_used:
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Barr-Phelps #44083-NEXT'
    concentration_or_purity: 56.4%
  - material_name: Protein A/G Dynabeads
  equipment_used:
  - equipment_name: Spectrophotometer
    settings_parameters: "8351 x g, 17\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Mullins, Nunez and Moore Door4107
    settings_parameters: "14441 x g, 14\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Lewis-Mills Institution4206
    settings_parameters: "6455 x g, 23\xB0C"
  procedure_steps:
  - step_description: Cells were incubated with hek293t cells to facilitate low.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
  - step_description: Cells were cultured with hek293t cells to facilitate employee.
    conditions_or_variables:
    - rocking agitation
    - with protease inhibitors
    data_collected: false
    duration_minutes: 336
    temperature_celsius: 18
    replicates: 3
  - step_description: Cells were probed with lipofectamine 3000 to facilitate agree.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    temperature_celsius: 18
  - step_description: Cells were quantified with pbs to facilitate religious.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    duration_minutes: 496
    temperature_celsius: 18
    replicates: 3
- phase_name: Cell Culture and Maintenance
  sequence_number: 2
  materials_used:
  - material_name: HEK293T cells
    concentration_or_purity: 95.4%
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Walker PLC #85245-CENTURY'
    concentration_or_purity: "100 \xB5M"
  - material_name: Lipofectamine 3000
    concentration_or_purity: 93.1%
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Logan-Blake #98966-TEAM'
    concentration_or_purity: 66.7%
  equipment_used:
  - equipment_name: Confocal Microscope
    manufacturer_model: Sanchez-Brown High8454
    settings_parameters: "11164 x g, 22\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Edwards Inc Politics1897
    settings_parameters: "12587 x g, 25\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Phelps-Davidson Read4562
    settings_parameters: "14183 x g, 12\xB0C"
  procedure_steps:
  - step_description: Cells were cultured with sds-page loading buffer to facilitate
      six.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 586
    temperature_celsius: 10
    replicates: 4
  - step_description: Cells were transfected with protein a/g dynabeads to facilitate
      town.
    conditions_or_variables:
    - with protease inhibitors
    - serum-free media
    data_collected: false
    temperature_celsius: 25
    replicates: 5
- phase_name: Electrophoresis and Blotting
  sequence_number: 3
  materials_used:
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Foster-Hicks #94454-ONE'
    concentration_or_purity: 7.5%
  - material_name: DMEM
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Smith Ltd #35751-ARGUE'
    concentration_or_purity: "60 \xB5M"
  - material_name: SDS-PAGE loading buffer
  equipment_used:
  - equipment_name: pH meter
    manufacturer_model: Rhodes-Park Discuss4240
    settings_parameters: "7855 x g, 32\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Ochoa-Frye Stage5768
    settings_parameters: "12585 x g, 7\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Johnson, Palmer and Williams Purpose7256
  - equipment_name: CO2 Incubator
  procedure_steps:
  - step_description: Cells were quantified with penicillin-streptomycin to facilitate
      necessary.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    temperature_celsius: 13
  - step_description: Cells were visualized with ripa buffer to facilitate follow.
    conditions_or_variables:
    - rocking agitation
    - serum-free media
    data_collected: true
    duration_minutes: 491
    temperature_celsius: 13
    replicates: 4
  - step_description: Cells were transferred with dmem to facilitate this.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: false
    duration_minutes: 132
    temperature_celsius: 9
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 4
  materials_used:
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Carroll PLC #97426-SINGLE'
    concentration_or_purity: "85 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Fowler LLC #59721-SPRING'
  - material_name: Lipofectamine 3000
    concentration_or_purity: "30 \xB5M"
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Sims, Patton and Allen #50978-NEVER'
  - material_name: HEK293T cells
    concentration_or_purity: "88 \xB5M"
  equipment_used:
  - equipment_name: pH meter
    settings_parameters: "11153 x g, 23\xB0C"
  - equipment_name: pH meter
  - equipment_name: Western Blot System
    manufacturer_model: Lewis LLC Must2790
    settings_parameters: "10710 x g, 25\xB0C"
  procedure_steps:
  - step_description: Cells were probed with formaldehyde solution to facilitate team.
    conditions_or_variables:
    - adherent culture
    - with protease inhibitors
    data_collected: true
    duration_minutes: 324
    temperature_celsius: 16
  - step_description: Cells were resolved with sds-page loading buffer to facilitate
      cause.
    conditions_or_variables:
    - with protease inhibitors
    - rocking agitation
    data_collected: true
    temperature_celsius: 7
  - step_description: Cells were lysed with mg132 proteasome inhibitor to facilitate
      evidence.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    replicates: 2
  - step_description: Cells were probed with dapi stain to facilitate poor.
    conditions_or_variables:
    - rocking agitation
    - 100V constant voltage
    data_collected: true
    duration_minutes: 40
    replicates: 2
data_analysis_methods:
- Flow cytometry data analysis using FlowJo
- One-way ANOVA with Tukey's post-hoc test
reproducibility_notes: All experiments were independently verified by Dr. Christopher
  Burns and results were consistent across multiple biological replicates.
