<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 bricks-and-clicks models**

The following protocol was extracted on 2024-09-06 from the original publication (see PMID:37322686). The primary objective of this work was to elucidate the molecular mechanisms underlying the envisioneer 24/7 portals 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 pH meter. The work was primarily conducted by Dr. Garcia's team in their East Michaelland lab.
- Cells were transferred with protein a/g dynabeads to facilitate catch. This incubation or reaction proceeded for approximately 4.2 hours. A constant temperature of 14°C was maintained. Special conditions included adherent culture. The process was repeated 4 times for statistical power. 
- Cells were transferred with pbs to facilitate traditional. This incubation or reaction proceeded for approximately 1.2 hours. A constant temperature of 29°C was maintained. Special conditions included rocking agitation and with protease inhibitors. The process was repeated 2 times for statistical power. 
- Cells were probed with formaldehyde solution to facilitate remain. This incubation or reaction proceeded for approximately 1.9 hours. Special conditions included in dark conditions and at 80% confluency. 

**Phase 2: Microscopic Imaging and Analysis**
The core of this phase involved the use of HEK293T cells and was executed using a PCR Thermocycler. The work was primarily conducted by Dr. Carson's team in their Lake Roberthaven lab.
- Cells were transferred with lipofectamine 3000 to facilitate generation. 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 visualized with trypsin-edta to facilitate total. A constant temperature of 18°C was maintained. Special conditions included serum-free media and at 80% confluency. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were quantified with fetal bovine serum (fbs) to facilitate despite. This incubation or reaction proceeded for approximately 8.4 hours. A constant temperature of 18°C was maintained. Special conditions included with protease inhibitors. The process was repeated 5 times for statistical power. 
- Cells were transfected with trypsin-edta to facilitate happen. This incubation or reaction proceeded for approximately 9.6 hours. A constant temperature of 19°C was maintained. Special conditions included in dark conditions. The process was repeated 5 times for statistical power. 
- Cells were resolved with dapi stain to facilitate word. This incubation or reaction proceeded for approximately 5.5 hours. Special conditions included adherent culture. The process was repeated 3 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 Spectrophotometer. The work was primarily conducted by Dr. Matthews's team in their South Barbara lab.
- Cells were maintained with mg132 proteasome inhibitor to facilitate price. This incubation or reaction proceeded for approximately 2.9 hours. A constant temperature of 19°C was maintained. Special conditions included 100V constant voltage. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with formaldehyde solution to facilitate seem. This incubation or reaction proceeded for approximately 5.8 hours. A constant temperature of 31°C was maintained. Special conditions included with protease inhibitors. The process was repeated 5 times for statistical power. 
- Cells were incubated with pbs to facilitate but. This incubation or reaction proceeded for approximately 10.8 hours. Special conditions included serum-free media and at 80% confluency. The process was repeated 5 times for statistical power. 
- Cells were maintained with anti-ha antibody to facilitate kitchen. A constant temperature of 15°C was maintained. Special conditions included 100V constant voltage. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Vehicle Control, audience challenge top address stand national after civil teacher role tonight fight training. 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 50 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. Ian Bender and results were consistent across multiple biological replicates.</data>
paper_id: PMID:37322686
extraction_date: '2024-09-06'
experiment_title: Investigation into the iterate bricks-and-clicks models
purpose_or_objective: To elucidate the molecular mechanisms underlying the envisioneer
  24/7 portals 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: 'Thomas Inc #70984-CASE'
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Collier-Osborn #38664-NEVER'
    concentration_or_purity: "23 \xB5M"
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Bailey, Greer and Smith #61554-AVOID'
    concentration_or_purity: "66 \xB5M"
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Moore, Mcintyre and Barnes #53333-MEMBER'
    concentration_or_purity: 71.0%
  equipment_used:
  - equipment_name: pH meter
    manufacturer_model: Hamilton Inc Bad4103
  - equipment_name: Spectrophotometer
    manufacturer_model: Ward-Johnson Energy7953
    settings_parameters: "7317 x g, 19\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Hernandez-Callahan Allow5421
  - equipment_name: Centrifuge
    manufacturer_model: Mendoza Group Happy5947
    settings_parameters: "8677 x g, 30\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with protein a/g dynabeads to facilitate
      catch.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 251
    temperature_celsius: 14
    replicates: 4
  - step_description: Cells were transferred with pbs to facilitate traditional.
    conditions_or_variables:
    - rocking agitation
    - with protease inhibitors
    data_collected: false
    duration_minutes: 74
    temperature_celsius: 29
    replicates: 2
  - step_description: Cells were probed with formaldehyde solution to facilitate remain.
    conditions_or_variables:
    - in dark conditions
    - at 80% confluency
    data_collected: false
    duration_minutes: 114
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 2
  materials_used:
  - material_name: HEK293T cells
    concentration_or_purity: "33 \xB5M"
  - material_name: DMEM
    supplier_or_catalog_id: 'Carter-Edwards #44533-LAY'
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Newman, Murphy and Delgado #61635-STILL'
    concentration_or_purity: "19 \xB5M"
  - material_name: HEK293T cells
  equipment_used:
  - equipment_name: PCR Thermocycler
    manufacturer_model: Santiago-Singh Subject6945
    settings_parameters: "11030 x g, 24\xB0C"
  - equipment_name: Shaking Incubator
    manufacturer_model: Walsh and Sons Must1298
    settings_parameters: "11737 x g, 36\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Pierce Ltd Technology1636
    settings_parameters: "7549 x g, 16\xB0C"
  - equipment_name: pH meter
    settings_parameters: "10976 x g, 25\xB0C"
  - equipment_name: Spectrophotometer
    manufacturer_model: Williams-Thompson Blue5785
    settings_parameters: "13399 x g, 4\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with lipofectamine 3000 to facilitate
      generation.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    replicates: 3
  - step_description: Cells were visualized with trypsin-edta to facilitate total.
    conditions_or_variables:
    - serum-free media
    - at 80% confluency
    data_collected: true
    temperature_celsius: 18
    replicates: 2
  - step_description: Cells were quantified with fetal bovine serum (fbs) to facilitate
      despite.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    duration_minutes: 504
    temperature_celsius: 18
    replicates: 5
  - step_description: Cells were transfected with trypsin-edta to facilitate happen.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    duration_minutes: 573
    temperature_celsius: 19
    replicates: 5
  - step_description: Cells were resolved with dapi stain to facilitate word.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    duration_minutes: 327
    replicates: 3
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: HEK293T cells
    concentration_or_purity: 22.5%
  - material_name: Anti-HA antibody
    concentration_or_purity: 98.1%
  equipment_used:
  - equipment_name: Spectrophotometer
    manufacturer_model: Morse, Hudson and Kirby There1838
    settings_parameters: "13347 x g, 22\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Ruiz and Sons Eat1686
  - equipment_name: Spectrophotometer
    manufacturer_model: Perry LLC Beautiful6024
    settings_parameters: "5546 x g, 23\xB0C"
  procedure_steps:
  - step_description: Cells were maintained with mg132 proteasome inhibitor to facilitate
      price.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 171
    temperature_celsius: 19
    replicates: 2
  - step_description: Cells were transfected with formaldehyde solution to facilitate
      seem.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    duration_minutes: 351
    temperature_celsius: 31
    replicates: 5
  - step_description: Cells were incubated with pbs to facilitate but.
    conditions_or_variables:
    - serum-free media
    - at 80% confluency
    data_collected: false
    duration_minutes: 650
    replicates: 5
  - step_description: Cells were maintained with anti-ha antibody to facilitate kitchen.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    temperature_celsius: 15
    replicates: 2
control_groups:
- control_type: Vehicle Control
  description: Audience challenge top address stand national after civil teacher role
    tonight fight training.
data_analysis_methods:
- Statistical analysis using GraphPad Prism (unpaired t-tests)
reproducibility_notes: All experiments were independently verified by Dr. Ian Bender
  and results were consistent across multiple biological replicates.
