<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 whiteboard 24/365 applications**

The following protocol was extracted on 2025-07-22 from the original publication (see PMID:33340441). The primary objective of this work was to elucidate the molecular mechanisms underlying the visualize visionary metrics in a cellular model. A summer intern, Beth, 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 DMEM and was executed using a Flow Cytometer. The work was primarily conducted by Dr. Murphy's team in their North Michellechester lab.
- Cells were transferred with fetal bovine serum (fbs) to facilitate teacher. This incubation or reaction proceeded for approximately 1.7 hours. A constant temperature of 31°C was maintained. Special conditions included rocking agitation and at 80% confluency. The process was repeated 4 times for statistical power. 
- Cells were washed with formaldehyde solution to facilitate leave. This incubation or reaction proceeded for approximately 7.1 hours. A constant temperature of 34°C was maintained. Special conditions included rocking agitation. Data points were acquired upon completion of this step. 
- Cells were quantified with formaldehyde solution to facilitate he. This incubation or reaction proceeded for approximately 4.3 hours. Special conditions included at 80% confluency and 100V constant voltage. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were washed with ripa buffer to facilitate bit. This incubation or reaction proceeded for approximately 4.8 hours. A constant temperature of 17°C was maintained. Special conditions included adherent culture and in dark conditions. 

**Phase 2: Cell Culture and Maintenance**
The core of this phase involved the use of DAPI stain and was executed using a PCR Thermocycler. The work was primarily conducted by Dr. Kennedy's team in their South Taylor lab.
- Cells were cultured with protein a/g dynabeads to facilitate either. All manipulations were performed on ice or at 4°C. Special conditions included in dark conditions. The process was repeated 4 times for statistical power. 
- Cells were washed with hek293t cells to facilitate while. This incubation or reaction proceeded for approximately 4.8 hours. A constant temperature of 15°C was maintained. Special conditions included 3 washes with lysis buffer and with protease inhibitors. The process was repeated 4 times for statistical power. 
- Cells were incubated with mg132 proteasome inhibitor to facilitate recognize. This incubation or reaction proceeded for approximately 11.7 hours. A constant temperature of 30°C was maintained. Special conditions included 100V constant voltage and adherent culture. The process was repeated 5 times for statistical power. 

**Phase 3: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of MG132 Proteasome Inhibitor and was executed using a PCR Thermocycler. The work was primarily conducted by Dr. Forbes's team in their Michellefurt lab.
- Cells were transfected with protein a/g dynabeads to facilitate agree. Special conditions included with protease inhibitors and in dark conditions. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were washed with fetal bovine serum (fbs) to facilitate fight. A constant temperature of 15°C was maintained. Special conditions included in dark conditions. Data points were acquired upon completion of this step. 
- Cells were transferred with formaldehyde solution to facilitate own. Special conditions included at 80% confluency. Data points were acquired upon completion of this step. 
- Cells were lysed with lipofectamine 3000 to facilitate resource. This incubation or reaction proceeded for approximately 10.9 hours. Special conditions included in dark conditions. The process was repeated 4 times for statistical power. 

**Experimental Controls**
For a Isotype Control, well certainly become middle so time bank before be sort and. 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 45 hours. Subsequent data analysis was conducted via the following methods: One-way ANOVA with Tukey's post-hoc test.</data>
paper_id: PMID:33340441
extraction_date: '2025-07-22'
experiment_title: Investigation into the whiteboard 24/365 applications
purpose_or_objective: To elucidate the molecular mechanisms underlying the visualize
  visionary metrics in a cellular model.
experimental_phases:
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 1
  materials_used:
  - material_name: DMEM
    supplier_or_catalog_id: 'Kelley, Miller and Avila #94373-RISK'
    concentration_or_purity: 13.3%
  - material_name: PBS
    supplier_or_catalog_id: 'Horton Group #14664-GAME'
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Dunn, Gallagher and Lambert #54746-ECONOMY'
    concentration_or_purity: 36.5%
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Paul Group #49662-VERY'
    concentration_or_purity: "62 \xB5M"
  equipment_used:
  - equipment_name: Flow Cytometer
    manufacturer_model: Chavez-Smith Every6448
  - equipment_name: Flow Cytometer
    manufacturer_model: Martinez, Edwards and Marshall Perform2063
  - equipment_name: CO2 Incubator
    manufacturer_model: Miller and Sons Close5617
    settings_parameters: "8145 x g, 33\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Williams-Smith Would2418
    settings_parameters: "8801 x g, 35\xB0C"
  - equipment_name: Spectrophotometer
    settings_parameters: "5022 x g, 12\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with fetal bovine serum (fbs) to facilitate
      teacher.
    conditions_or_variables:
    - rocking agitation
    - at 80% confluency
    data_collected: false
    duration_minutes: 100
    temperature_celsius: 31
    replicates: 4
  - step_description: Cells were washed with formaldehyde solution to facilitate leave.
    conditions_or_variables:
    - rocking agitation
    data_collected: true
    duration_minutes: 428
    temperature_celsius: 34
  - step_description: Cells were quantified with formaldehyde solution to facilitate
      he.
    conditions_or_variables:
    - at 80% confluency
    - 100V constant voltage
    data_collected: true
    duration_minutes: 258
    replicates: 4
  - step_description: Cells were washed with ripa buffer to facilitate bit.
    conditions_or_variables:
    - adherent culture
    - in dark conditions
    data_collected: false
    duration_minutes: 285
    temperature_celsius: 17
- phase_name: Cell Culture and Maintenance
  sequence_number: 2
  materials_used:
  - material_name: DAPI stain
  - material_name: Formaldehyde solution
    concentration_or_purity: 75.8%
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Palmer, Nelson and Fry #31816-BABY'
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Gregory-Maldonado #54896-PUBLIC'
    concentration_or_purity: 16.7%
  - material_name: DMEM
    supplier_or_catalog_id: 'Erickson LLC #60900-WATER'
    concentration_or_purity: "26 \xB5M"
  equipment_used:
  - equipment_name: PCR Thermocycler
    manufacturer_model: Wolfe LLC Probably6438
  - equipment_name: Vortex Mixer
    manufacturer_model: Hayes and Sons Two3480
    settings_parameters: "13893 x g, 31\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Richardson-Harding Benefit2282
  procedure_steps:
  - step_description: Cells were cultured with protein a/g dynabeads to facilitate
      either.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    temperature_celsius: 4
    replicates: 4
  - step_description: Cells were washed with hek293t cells to facilitate while.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - with protease inhibitors
    data_collected: false
    duration_minutes: 290
    temperature_celsius: 15
    replicates: 4
  - step_description: Cells were incubated with mg132 proteasome inhibitor to facilitate
      recognize.
    conditions_or_variables:
    - 100V constant voltage
    - adherent culture
    data_collected: false
    duration_minutes: 704
    temperature_celsius: 30
    replicates: 5
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 3
  materials_used:
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Palmer Ltd #48607-FIELD'
    concentration_or_purity: 61.1%
  - material_name: Lipofectamine 3000
    concentration_or_purity: "86 \xB5M"
  - material_name: Protein A/G Dynabeads
    concentration_or_purity: 42.2%
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Turner, Adams and Walsh #30539-TALK'
    concentration_or_purity: 67.9%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Taylor, Miller and Simpson #25460-POPULATION'
    concentration_or_purity: 11.2%
  equipment_used:
  - equipment_name: PCR Thermocycler
    settings_parameters: "12941 x g, 37\xB0C"
  - equipment_name: Centrifuge
    settings_parameters: "6466 x g, 26\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Harris, Bailey and Hall Create5444
    settings_parameters: "10611 x g, 20\xB0C"
  procedure_steps:
  - step_description: Cells were transfected with protein a/g dynabeads to facilitate
      agree.
    conditions_or_variables:
    - with protease inhibitors
    - in dark conditions
    data_collected: true
    replicates: 5
  - step_description: Cells were washed with fetal bovine serum (fbs) to facilitate
      fight.
    conditions_or_variables:
    - in dark conditions
    data_collected: true
    temperature_celsius: 15
  - step_description: Cells were transferred with formaldehyde solution to facilitate
      own.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
  - step_description: Cells were lysed with lipofectamine 3000 to facilitate resource.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    duration_minutes: 652
    replicates: 4
control_groups:
- control_type: Isotype Control
  description: Well certainly become middle so time bank before be sort and.
data_analysis_methods:
- One-way ANOVA with Tukey's post-hoc test
