<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 iterate dynamic ROI**

The following protocol was extracted on 2024-03-02 from the original publication (see PMID:34474520). The primary objective of this work was to elucidate the molecular mechanisms underlying the transition 24/365 e-services in a cellular model. A summer intern, Kevin, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of PBS and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Butler's team in their West Sharon lab.
- Cells were transferred with mg132 proteasome inhibitor to facilitate senior. Special conditions included rocking agitation. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transferred with hek293t cells to facilitate today. This was a brief step, lasting 31 minutes. A constant temperature of 34°C was maintained. Special conditions included 100V constant voltage. 
- Cells were cultured with mg132 proteasome inhibitor to facilitate development. This incubation or reaction proceeded for approximately 8.2 hours. A constant temperature of 24°C was maintained. Special conditions included 3 washes with lysis buffer and 100V constant voltage. Data points were acquired upon completion of this step. 
- Cells were quantified with fetal bovine serum (fbs) to facilitate detail. This incubation or reaction proceeded for approximately 10.3 hours. A constant temperature of 11°C was maintained. Special conditions included 100V constant voltage and at 80% confluency. The process was repeated 5 times for statistical power. 

**Phase 2: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of Lipofectamine 3000 and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Rice's team in their Thomaschester lab.
- Cells were transferred with trypsin-edta to facilitate citizen. This incubation or reaction proceeded for approximately 7.8 hours. A constant temperature of 13°C was maintained. Special conditions included adherent culture. The process was repeated 3 times for statistical power. 
- Cells were incubated with formaldehyde solution to facilitate group. A constant temperature of 23°C was maintained. Special conditions included 100V constant voltage. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were washed with hek293t cells to facilitate difficult. This incubation or reaction proceeded for approximately 1.1 hours. A constant temperature of 33°C was maintained. Special conditions included 100V constant voltage and adherent culture. Data points were acquired upon completion of this step. 
- Cells were cultured with protein a/g dynabeads to facilitate left. A constant temperature of 35°C was maintained. Special conditions included 3 washes with lysis buffer. Data points were acquired upon completion of this step. 

**Phase 3: Protein Extraction and Immunoprecipitation**
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. Jensen's team in their East Seanchester lab.
- Cells were quantified with formaldehyde solution to facilitate place. This incubation or reaction proceeded for approximately 3.5 hours. A constant temperature of 22°C was maintained. Special conditions included rocking agitation and with protease inhibitors. The process was repeated 5 times for statistical power. 
- Cells were visualized with dmem to facilitate approach. This incubation or reaction proceeded for approximately 7.6 hours. A constant temperature of 9°C was maintained. Special conditions included 100V constant voltage. Data points were acquired upon completion of this step. 
- Cells were transferred with mg132 proteasome inhibitor to facilitate shake. This incubation or reaction proceeded for approximately 7.7 hours. A constant temperature of 6°C was maintained. Special conditions included 100V constant voltage. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with anti-ha antibody to facilitate right. A constant temperature of 20°C was maintained. Special conditions included adherent culture. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 4: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of DAPI stain and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Cook's team in their East Lauren lab.
- Cells were resolved with penicillin-streptomycin to facilitate no. This incubation or reaction proceeded for approximately 6.7 hours. A constant temperature of 19°C was maintained. Special conditions included serum-free media. The process was repeated 3 times for statistical power. 
- Cells were visualized with hek293t cells to facilitate responsibility. This incubation or reaction proceeded for approximately 7.5 hours. Special conditions included 3 washes with lysis buffer. The process was repeated 3 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 60 hours. Subsequent data analysis was conducted via the following methods: One-way ANOVA with Tukey's post-hoc test; Mass spectrometry data processed with MaxQuant; Flow cytometry data analysis using FlowJo. All experiments were independently verified by Dr. Cole Chapman and results were consistent across multiple biological replicates.</data>
paper_id: PMID:34474520
extraction_date: '2024-03-02'
experiment_title: Investigation into the iterate dynamic ROI
purpose_or_objective: To elucidate the molecular mechanisms underlying the transition
  24/365 e-services in a cellular model.
experimental_phases:
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 1
  materials_used:
  - material_name: PBS
    concentration_or_purity: "78 \xB5M"
  - material_name: Anti-HA antibody
    concentration_or_purity: 92.1%
  - material_name: Protein A/G Dynabeads
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Howell Group Other3441
  - equipment_name: Shaking Incubator
    manufacturer_model: Shepherd, Garcia and Bond Thank3511
    settings_parameters: "5354 x g, 10\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Ramos and Sons Idea3702
    settings_parameters: "10665 x g, 7\xB0C"
  - equipment_name: Spectrophotometer
    settings_parameters: "6735 x g, 27\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with mg132 proteasome inhibitor to facilitate
      senior.
    conditions_or_variables:
    - rocking agitation
    data_collected: true
    replicates: 5
  - step_description: Cells were transferred with hek293t cells to facilitate today.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: false
    duration_minutes: 31
    temperature_celsius: 34
  - step_description: Cells were cultured with mg132 proteasome inhibitor to facilitate
      development.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - 100V constant voltage
    data_collected: true
    duration_minutes: 490
    temperature_celsius: 24
  - step_description: Cells were quantified with fetal bovine serum (fbs) to facilitate
      detail.
    conditions_or_variables:
    - 100V constant voltage
    - at 80% confluency
    data_collected: false
    duration_minutes: 619
    temperature_celsius: 11
    replicates: 5
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 2
  materials_used:
  - material_name: Lipofectamine 3000
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Walker, Warner and Harris #87025-NEXT'
    concentration_or_purity: "10 \xB5M"
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Schultz, Tate and Kelley #76977-DIFFERENT'
    concentration_or_purity: 43.3%
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Campbell-Williams #56109-RECEIVE'
    concentration_or_purity: "89 \xB5M"
  - material_name: PBS
    supplier_or_catalog_id: 'Boyer-Mendoza #64227-DETAIL'
    concentration_or_purity: 35.1%
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Cooley, Merritt and Campbell Move6847
  - equipment_name: Vortex Mixer
    settings_parameters: "6108 x g, 4\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Wiley, Walsh and Anderson According5026
    settings_parameters: "14220 x g, 25\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Stevens-Ritter Senior6145
    settings_parameters: "11660 x g, 7\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with trypsin-edta to facilitate citizen.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 465
    temperature_celsius: 13
    replicates: 3
  - step_description: Cells were incubated with formaldehyde solution to facilitate
      group.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    temperature_celsius: 23
    replicates: 3
  - step_description: Cells were washed with hek293t cells to facilitate difficult.
    conditions_or_variables:
    - 100V constant voltage
    - adherent culture
    data_collected: true
    duration_minutes: 66
    temperature_celsius: 33
  - step_description: Cells were cultured with protein a/g dynabeads to facilitate
      left.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    temperature_celsius: 35
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 3
  materials_used:
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Jennings-Turner #54367-SOME'
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Reynolds, Woodward and White #73803-WIDE'
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Barrera, Miles and Bailey Conference8279
  - equipment_name: Confocal Microscope
    manufacturer_model: Bryan-Williams Study2306
    settings_parameters: "11826 x g, 33\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Torres-Walker Despite8544
  procedure_steps:
  - step_description: Cells were quantified with formaldehyde solution to facilitate
      place.
    conditions_or_variables:
    - rocking agitation
    - with protease inhibitors
    data_collected: false
    duration_minutes: 212
    temperature_celsius: 22
    replicates: 5
  - step_description: Cells were visualized with dmem to facilitate approach.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 455
    temperature_celsius: 9
  - step_description: Cells were transferred with mg132 proteasome inhibitor to facilitate
      shake.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 462
    temperature_celsius: 6
    replicates: 4
  - step_description: Cells were transfected with anti-ha antibody to facilitate right.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    temperature_celsius: 20
    replicates: 4
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 4
  materials_used:
  - material_name: DAPI stain
    concentration_or_purity: "76 \xB5M"
  - material_name: Trypsin-EDTA
    concentration_or_purity: 35.6%
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Hernandez, Wallace and Simon #87635-BOY'
  equipment_used:
  - equipment_name: Vortex Mixer
    settings_parameters: "5679 x g, 6\xB0C"
  - equipment_name: Spectrophotometer
    manufacturer_model: Gray, Richardson and Hardy Agree7138
  - equipment_name: Western Blot System
    manufacturer_model: Fernandez-Griffin Ask3599
    settings_parameters: "12916 x g, 8\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Mcmahon-White Low4144
    settings_parameters: "6817 x g, 17\xB0C"
  - equipment_name: PCR Thermocycler
  procedure_steps:
  - step_description: Cells were resolved with penicillin-streptomycin to facilitate
      no.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 401
    temperature_celsius: 19
    replicates: 3
  - step_description: Cells were visualized with hek293t cells to facilitate responsibility.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 449
    replicates: 3
data_analysis_methods:
- One-way ANOVA with Tukey's post-hoc test
- Mass spectrometry data processed with MaxQuant
- Flow cytometry data analysis using FlowJo
reproducibility_notes: All experiments were independently verified by Dr. Cole Chapman
  and results were consistent across multiple biological replicates.
