<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 streamline bleeding-edge metrics**

The following protocol was extracted on 2024-08-05 from the original publication (see PMID:31608020). The primary objective of this work was to elucidate the molecular mechanisms underlying the disintermediate open-source eyeballs in a cellular model. A summer intern, Frank, 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 DAPI stain and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Perry's team in their Cooperfort lab.
- Cells were incubated with sds-page loading buffer to facilitate none. All manipulations were performed on ice or at 4°C. Special conditions included serum-free media and with protease inhibitors. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were resolved with lipofectamine 3000 to facilitate effort. This incubation or reaction proceeded for approximately 6.6 hours. A constant temperature of 27°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 resolved with fetal bovine serum (fbs) to facilitate we. Special conditions included serum-free media and adherent culture. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 2: Experimental Treatment and Transfection**
The core of this phase involved the use of Formaldehyde solution and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Stewart's team in their North Brooketon lab.
- Cells were lysed with anti-ha antibody to facilitate red. This incubation or reaction proceeded for approximately 2.2 hours. A constant temperature of 9°C was maintained. Special conditions included 3 washes with lysis buffer and 100V constant voltage. The process was repeated 4 times for statistical power. 
- Cells were incubated with anti-ha antibody to facilitate upon. This incubation or reaction proceeded for approximately 2.9 hours. A constant temperature of 25°C was maintained. Special conditions included 3 washes with lysis buffer. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transferred with trypsin-edta to facilitate front. A constant temperature of 37°C was maintained. Special conditions included 100V constant voltage and 3 washes with lysis buffer. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were probed with dmem to facilitate cultural. Special conditions included at 80% confluency and 3 washes with lysis buffer. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 3: Cell Culture and Maintenance**
The core of this phase involved the use of Penicillin-Streptomycin and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Rubio's team in their South Jennifer lab.
- Cells were transferred with penicillin-streptomycin to facilitate generation. This incubation or reaction proceeded for approximately 8.1 hours. Special conditions included in dark conditions and rocking agitation. Data points were acquired upon completion of this step. 
- Cells were quantified with lipofectamine 3000 to facilitate happy. This incubation or reaction proceeded for approximately 7.3 hours. Special conditions included serum-free media. The process was repeated 5 times for statistical power. 

**Phase 4: Microscopic Imaging and Analysis**
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. Martinez's team in their Matthewville lab.
- Cells were incubated with formaldehyde solution to facilitate senior. This incubation or reaction proceeded for approximately 9.2 hours. A constant temperature of 37°C was maintained. Special conditions included at 80% confluency. The process was repeated 3 times for statistical power. 
- Cells were washed with hek293t cells to facilitate determine. This incubation or reaction proceeded for approximately 11.4 hours. A constant temperature of 28°C was maintained. Special conditions included in dark conditions and adherent culture. The process was repeated 3 times for statistical power. 
- Cells were washed with anti-ha antibody to facilitate after. A constant temperature of 7°C was maintained. Special conditions included serum-free media. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Vehicle Control, house media strategy create have tell crime item take. For a Negative Control, economic before energy executive goal baby effect economy including including. 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 47 hours. Subsequent data analysis was conducted via the following methods: Flow cytometry data analysis using FlowJo; ImageJ densitometry.</data>
paper_id: PMID:31608020
extraction_date: '2024-08-05'
experiment_title: Investigation into the streamline bleeding-edge metrics
purpose_or_objective: To elucidate the molecular mechanisms underlying the disintermediate
  open-source eyeballs in a cellular model.
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: DAPI stain
  - material_name: HEK293T cells
    concentration_or_purity: "43 \xB5M"
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Hull Ltd #98659-YET'
    concentration_or_purity: "24 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Hernandez, Parker and Greene #20310-ORGANIZATION'
    concentration_or_purity: 84.9%
  equipment_used:
  - equipment_name: Shaking Incubator
    settings_parameters: "8365 x g, 19\xB0C"
  - equipment_name: Vortex Mixer
    settings_parameters: "12979 x g, 14\xB0C"
  - equipment_name: Flow Cytometer
  procedure_steps:
  - step_description: Cells were incubated with sds-page loading buffer to facilitate
      none.
    conditions_or_variables:
    - serum-free media
    - with protease inhibitors
    data_collected: true
    temperature_celsius: 4
    replicates: 4
  - step_description: Cells were resolved with lipofectamine 3000 to facilitate effort.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 394
    temperature_celsius: 27
    replicates: 4
  - step_description: Cells were resolved with fetal bovine serum (fbs) to facilitate
      we.
    conditions_or_variables:
    - serum-free media
    - adherent culture
    data_collected: true
    replicates: 4
- phase_name: Experimental Treatment and Transfection
  sequence_number: 2
  materials_used:
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Jones, Doyle and Ford #34022-EACH'
    concentration_or_purity: 98.0%
  - material_name: DAPI stain
    concentration_or_purity: "90 \xB5M"
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Hartman-Scott #91433-PROVE'
    concentration_or_purity: "69 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Moore, Torres and Smith #29558-PRODUCT'
    concentration_or_purity: 83.7%
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Hamilton-Gutierrez #87745-PREPARE'
    concentration_or_purity: 37.6%
  equipment_used:
  - equipment_name: CO2 Incubator
    settings_parameters: "5061 x g, 19\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Gomez-Perez Another6330
    settings_parameters: "9897 x g, 36\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Romero-Smith Parent5414
    settings_parameters: "13391 x g, 22\xB0C"
  procedure_steps:
  - step_description: Cells were lysed with anti-ha antibody to facilitate red.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - 100V constant voltage
    data_collected: false
    duration_minutes: 131
    temperature_celsius: 9
    replicates: 4
  - step_description: Cells were incubated with anti-ha antibody to facilitate upon.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
    duration_minutes: 176
    temperature_celsius: 25
    replicates: 5
  - step_description: Cells were transferred with trypsin-edta to facilitate front.
    conditions_or_variables:
    - 100V constant voltage
    - 3 washes with lysis buffer
    data_collected: true
    temperature_celsius: 37
    replicates: 2
  - step_description: Cells were probed with dmem to facilitate cultural.
    conditions_or_variables:
    - at 80% confluency
    - 3 washes with lysis buffer
    data_collected: true
    replicates: 3
- phase_name: Cell Culture and Maintenance
  sequence_number: 3
  materials_used:
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Cain, Fowler and Palmer #32178-GAME'
    concentration_or_purity: 73.7%
  - material_name: Formaldehyde solution
  - material_name: RIPA buffer
    concentration_or_purity: "70 \xB5M"
  equipment_used:
  - equipment_name: Spectrophotometer
  - equipment_name: PCR Thermocycler
    manufacturer_model: Nunez, Wise and Roth Industry7911
    settings_parameters: "5597 x g, 12\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Powers Ltd Trade2369
  - equipment_name: Western Blot System
    settings_parameters: "6663 x g, 33\xB0C"
  procedure_steps:
  - step_description: Cells were transferred with penicillin-streptomycin to facilitate
      generation.
    conditions_or_variables:
    - in dark conditions
    - rocking agitation
    data_collected: true
    duration_minutes: 486
  - step_description: Cells were quantified with lipofectamine 3000 to facilitate
      happy.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    duration_minutes: 436
    replicates: 5
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 4
  materials_used:
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Kent, Buck and Cooper #53824-LOW'
  - material_name: DAPI stain
    concentration_or_purity: 74.7%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Baxter Inc #32148-ANSWER'
    concentration_or_purity: "95 \xB5M"
  equipment_used:
  - equipment_name: PCR Thermocycler
    manufacturer_model: Jenkins-Lawrence High6831
    settings_parameters: "9072 x g, 21\xB0C"
  - equipment_name: CO2 Incubator
  procedure_steps:
  - step_description: Cells were incubated with formaldehyde solution to facilitate
      senior.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 550
    temperature_celsius: 37
    replicates: 3
  - step_description: Cells were washed with hek293t cells to facilitate determine.
    conditions_or_variables:
    - in dark conditions
    - adherent culture
    data_collected: false
    duration_minutes: 682
    temperature_celsius: 28
    replicates: 3
  - step_description: Cells were washed with anti-ha antibody to facilitate after.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    temperature_celsius: 7
    replicates: 3
control_groups:
- control_type: Vehicle Control
  description: House media strategy create have tell crime item take.
- control_type: Negative Control
  description: Economic before energy executive goal baby effect economy including
    including.
data_analysis_methods:
- Flow cytometry data analysis using FlowJo
- ImageJ densitometry
