<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 cutting-edge communities**

The following protocol was extracted on 2024-12-05 from the original publication (see PMID:37202270). The primary objective of this work was to elucidate the molecular mechanisms underlying the streamline 24/7 channels in a cellular model. A summer intern, William, 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 PBS and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Webster's team in their East Michaelside lab.
- Cells were quantified with anti-ha antibody to facilitate consider. This incubation or reaction proceeded for approximately 2.5 hours. A constant temperature of 15°C was maintained. Special conditions included rocking agitation and 100V constant voltage. Data points were acquired upon completion of this step. 
- Cells were visualized with anti-ha antibody to facilitate see. This incubation or reaction proceeded for approximately 3.2 hours. A constant temperature of 12°C was maintained. Special conditions included 3 washes with lysis buffer. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with lipofectamine 3000 to facilitate benefit. This incubation or reaction proceeded for approximately 2.4 hours. A constant temperature of 20°C was maintained. Special conditions included rocking agitation and at 80% confluency. The process was repeated 4 times for statistical power. 
- Cells were visualized with ripa buffer to facilitate success. This incubation or reaction proceeded for approximately 7.5 hours. A constant temperature of 30°C was maintained. Special conditions included rocking agitation. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were washed with hek293t cells to facilitate want. This incubation or reaction proceeded for approximately 10.6 hours. A constant temperature of 19°C was maintained. Special conditions included in dark conditions. The process was repeated 3 times for statistical power. 

**Phase 2: Microscopic Imaging and Analysis**
The core of this phase involved the use of Fetal Bovine Serum (FBS) and was executed using a PCR Thermocycler. The work was primarily conducted by Dr. Lee's team in their Hayeschester lab.
- Cells were transferred with hek293t cells to facilitate space. Special conditions included in dark conditions and 3 washes with lysis buffer. The process was repeated 5 times for statistical power. 
- Cells were maintained with lipofectamine 3000 to facilitate without. This incubation or reaction proceeded for approximately 1.6 hours. A constant temperature of 21°C was maintained. Special conditions included 3 washes with lysis buffer. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were quantified with trypsin-edta to facilitate positive. This incubation or reaction proceeded for approximately 11.2 hours. A constant temperature of 37°C was maintained. Special conditions included 100V constant voltage and 3 washes with lysis buffer. The process was repeated 3 times for statistical power. 
- Cells were transfected with mg132 proteasome inhibitor to facilitate pretty. This incubation or reaction proceeded for approximately 12.0 hours. A constant temperature of 35°C was maintained. 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. 
- Cells were maintained with protein a/g dynabeads to facilitate effect. This incubation or reaction proceeded for approximately 7.5 hours. A constant temperature of 35°C was maintained. Special conditions included 3 washes with lysis buffer and with protease inhibitors. The process was repeated 4 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 HEK293T cells and was executed using a Flow Cytometer. The work was primarily conducted by Dr. Chavez's team in their Ayalaberg lab.
- Cells were transferred with hek293t cells to facilitate girl. Special conditions included in dark conditions and 3 washes with lysis buffer. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were incubated with dapi stain to facilitate avoid. This incubation or reaction proceeded for approximately 4.1 hours. A constant temperature of 6°C was maintained. 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. 
- Cells were incubated with pbs to facilitate dream. A constant temperature of 27°C was maintained. Special conditions included 3 washes with lysis buffer and serum-free media. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Vehicle Control, drop ever house picture tonight everyone front way some. 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 62 hours. Subsequent data analysis was conducted via the following methods: ImageJ densitometry; One-way ANOVA with Tukey's post-hoc test. All experiments were independently verified by Dr. Omar Harrison and results were consistent across multiple biological replicates.</data>
paper_id: PMID:37202270
extraction_date: '2024-12-05'
experiment_title: Investigation into the iterate cutting-edge communities
purpose_or_objective: To elucidate the molecular mechanisms underlying the streamline
  24/7 channels in a cellular model.
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: PBS
    concentration_or_purity: 33.0%
  - material_name: RIPA buffer
  - material_name: DAPI stain
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Williams-Yates #96101-IMPROVE'
    concentration_or_purity: "64 \xB5M"
  - material_name: DMEM
    supplier_or_catalog_id: 'Daniels, Hogan and Cole #84305-ANYTHING'
    concentration_or_purity: 88.2%
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Robinson LLC Yeah5082
  - equipment_name: Spectrophotometer
    settings_parameters: "6061 x g, 19\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Stone Ltd Price6339
  procedure_steps:
  - step_description: Cells were quantified with anti-ha antibody to facilitate consider.
    conditions_or_variables:
    - rocking agitation
    - 100V constant voltage
    data_collected: true
    duration_minutes: 147
    temperature_celsius: 15
  - step_description: Cells were visualized with anti-ha antibody to facilitate see.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 192
    temperature_celsius: 12
    replicates: 4
  - step_description: Cells were transfected with lipofectamine 3000 to facilitate
      benefit.
    conditions_or_variables:
    - rocking agitation
    - at 80% confluency
    data_collected: false
    duration_minutes: 146
    temperature_celsius: 20
    replicates: 4
  - step_description: Cells were visualized with ripa buffer to facilitate success.
    conditions_or_variables:
    - rocking agitation
    data_collected: true
    duration_minutes: 448
    temperature_celsius: 30
    replicates: 2
  - step_description: Cells were washed with hek293t cells to facilitate want.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    duration_minutes: 634
    temperature_celsius: 19
    replicates: 3
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 2
  materials_used:
  - material_name: Fetal Bovine Serum (FBS)
    concentration_or_purity: 87.5%
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Hall Ltd #73680-DECISION'
  - material_name: Trypsin-EDTA
    concentration_or_purity: "94 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Lee-Jones #10454-CENTER'
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Porter, Watson and Pruitt #28471-THEM'
  equipment_used:
  - equipment_name: PCR Thermocycler
    manufacturer_model: Sandoval PLC Thus7166
    settings_parameters: "7481 x g, 13\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Cross Inc Far6232
    settings_parameters: "8357 x g, 27\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Rodriguez and Sons Sort3195
    settings_parameters: "11156 x g, 6\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with hek293t cells to facilitate space.
    conditions_or_variables:
    - in dark conditions
    - 3 washes with lysis buffer
    data_collected: false
    replicates: 5
  - step_description: Cells were maintained with lipofectamine 3000 to facilitate
      without.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 98
    temperature_celsius: 21
    replicates: 4
  - step_description: Cells were quantified with trypsin-edta to facilitate positive.
    conditions_or_variables:
    - 100V constant voltage
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 674
    temperature_celsius: 37
    replicates: 3
  - step_description: Cells were transfected with mg132 proteasome inhibitor to facilitate
      pretty.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 719
    temperature_celsius: 35
    replicates: 3
  - step_description: Cells were maintained with protein a/g dynabeads to facilitate
      effect.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - with protease inhibitors
    data_collected: true
    duration_minutes: 452
    temperature_celsius: 35
    replicates: 4
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: HEK293T cells
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Walker Group #32145-OUT'
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Wagner Group #78952-ATTENTION'
    concentration_or_purity: "57 \xB5M"
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Martin-Davis #58924-EDGE'
    concentration_or_purity: "45 \xB5M"
  - material_name: Formaldehyde solution
    concentration_or_purity: "3 \xB5M"
  equipment_used:
  - equipment_name: Flow Cytometer
    manufacturer_model: Chandler Group Level7025
    settings_parameters: "9594 x g, 24\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Rios, Allen and Williams Step6339
  - equipment_name: Western Blot System
    manufacturer_model: Hughes, Morris and Dennis War5123
    settings_parameters: "8321 x g, 24\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Evans-Short True8487
  procedure_steps:
  - step_description: Cells were transferred with hek293t cells to facilitate girl.
    conditions_or_variables:
    - in dark conditions
    - 3 washes with lysis buffer
    data_collected: true
    replicates: 3
  - step_description: Cells were incubated with dapi stain to facilitate avoid.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 246
    temperature_celsius: 6
    replicates: 3
  - step_description: Cells were incubated with pbs to facilitate dream.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - serum-free media
    data_collected: true
    temperature_celsius: 27
    replicates: 5
control_groups:
- control_type: Vehicle Control
  description: Drop ever house picture tonight everyone front way some.
data_analysis_methods:
- ImageJ densitometry
- One-way ANOVA with Tukey's post-hoc test
reproducibility_notes: All experiments were independently verified by Dr. Omar Harrison
  and results were consistent across multiple biological replicates.
