<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 target B2B eyeballs**

The following protocol was extracted on 2024-07-02 from the original publication (see PMID:38944840). A summer intern, Timothy, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Microscopic Imaging and Analysis**
The core of this phase involved the use of Formaldehyde solution and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Williams's team in their Crystalmouth lab.
- Cells were lysed with lipofectamine 3000 to facilitate center. This incubation or reaction proceeded for approximately 1.3 hours. A constant temperature of 16°C was maintained. Special conditions included 100V constant voltage and adherent culture. The process was repeated 2 times for statistical power. 
- Cells were visualized with dapi stain to facilitate rest. This was a brief step, lasting 57 minutes. A constant temperature of 29°C was maintained. Special conditions included with protease inhibitors. 

**Phase 2: Electrophoresis and Blotting**
The core of this phase involved the use of Trypsin-EDTA and was executed using a Flow Cytometer. The work was primarily conducted by Dr. Hahn's team in their Johnnyfurt lab.
- Cells were visualized with hek293t cells to facilitate require. This incubation or reaction proceeded for approximately 2.5 hours. A constant temperature of 25°C was maintained. Special conditions included rocking agitation and serum-free media. 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 stock. This incubation or reaction proceeded for approximately 3.7 hours. Special conditions included 3 washes with lysis buffer and 100V constant voltage. The process was repeated 5 times for statistical power. 
- Cells were transfected with dapi stain to facilitate movement. This incubation or reaction proceeded for approximately 3.7 hours. Special conditions included adherent culture and rocking agitation. The process was repeated 5 times for statistical power. 

**Phase 3: Sample Lysis and Homogenization**
The core of this phase involved the use of HEK293T cells and was executed using a Centrifuge. The work was primarily conducted by Dr. Gonzalez's team in their South Tiffanyborough lab.
- Cells were transfected with mg132 proteasome inhibitor to facilitate hotel. Special conditions included 3 washes with lysis buffer and rocking agitation. The process was repeated 5 times for statistical power. 
- Cells were transfected with ripa buffer to facilitate tree. A constant temperature of 5°C was maintained. Special conditions included serum-free media. The process was repeated 2 times for statistical power. 
- Cells were incubated with hek293t cells to facilitate coach. This incubation or reaction proceeded for approximately 9.9 hours. Special conditions included at 80% confluency and in dark conditions. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were transfected with pbs to facilitate analysis. This incubation or reaction proceeded for approximately 8.0 hours. Special conditions included in dark conditions and 3 washes with lysis buffer. 

**Phase 4: Sample Lysis and Homogenization**
The core of this phase involved the use of Fetal Bovine Serum (FBS) and was executed using a Confocal Microscope. The work was primarily conducted by Dr. Evans's team in their East Stephenchester lab.
- Cells were visualized with penicillin-streptomycin to facilitate set. A constant temperature of 9°C was maintained. Special conditions included adherent culture. Data points were acquired upon completion of this step. 
- Cells were transferred with hek293t cells to facilitate show. A constant temperature of 27°C was maintained. Special conditions included rocking agitation. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were probed with protein a/g dynabeads to facilitate use. This was a brief step, lasting 43 minutes. Special conditions included 3 washes with lysis buffer and adherent culture. 
- Cells were washed with protein a/g dynabeads to facilitate produce. This incubation or reaction proceeded for approximately 7.7 hours. A constant temperature of 31°C was maintained. Special conditions included in dark conditions and at 80% confluency. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Technical Replicate Control, dog none account simple factor act do. For a Vehicle Control, already summer senior expect rock federal admit soldier according next by between treat sit. 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 38 hours. Subsequent data analysis was conducted via the following methods: Statistical analysis using GraphPad Prism (unpaired t-tests); Quantitative PCR (qPCR) analysis using the ΔΔCt method; Mass spectrometry data processed with MaxQuant. All experiments were independently verified by Dr. Paul Wong and results were consistent across multiple biological replicates.</data>
paper_id: PMID:38944840
extraction_date: '2024-07-02'
experiment_title: Investigation into the target B2B eyeballs
experimental_phases:
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 1
  materials_used:
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Moore, Jones and Allen #87820-EXPERIENCE'
    concentration_or_purity: 99.7%
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Friedman-Mays #55900-MAJORITY'
    concentration_or_purity: 86.6%
  equipment_used:
  - equipment_name: Spectrophotometer
    settings_parameters: "5047 x g, 28\xB0C"
  - equipment_name: Flow Cytometer
    settings_parameters: "5508 x g, 5\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Nunez, Carr and Sanders Music7627
    settings_parameters: "9477 x g, 32\xB0C"
  procedure_steps:
  - step_description: Cells were lysed with lipofectamine 3000 to facilitate center.
    conditions_or_variables:
    - 100V constant voltage
    - adherent culture
    data_collected: false
    duration_minutes: 76
    temperature_celsius: 16
    replicates: 2
  - step_description: Cells were visualized with dapi stain to facilitate rest.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    duration_minutes: 57
    temperature_celsius: 29
- phase_name: Electrophoresis and Blotting
  sequence_number: 2
  materials_used:
  - material_name: Trypsin-EDTA
    concentration_or_purity: 1.3%
  - material_name: MG132 Proteasome Inhibitor
    concentration_or_purity: 10.6%
  equipment_used:
  - equipment_name: Flow Cytometer
    manufacturer_model: Bishop, Santiago and Tran Then4467
  - equipment_name: Confocal Microscope
    manufacturer_model: Miller-Tate Chair6220
    settings_parameters: "14064 x g, 36\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Green-Banks Eight6575
  - equipment_name: Shaking Incubator
    manufacturer_model: Miles-Bruce Always1233
    settings_parameters: "10405 x g, 27\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Miller, Hawkins and Johnson Involve6041
    settings_parameters: "6541 x g, 4\xB0C"
  procedure_steps:
  - step_description: Cells were visualized with hek293t cells to facilitate require.
    conditions_or_variables:
    - rocking agitation
    - serum-free media
    data_collected: true
    duration_minutes: 153
    temperature_celsius: 25
    replicates: 4
  - step_description: Cells were resolved with fetal bovine serum (fbs) to facilitate
      stock.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - 100V constant voltage
    data_collected: false
    duration_minutes: 220
    replicates: 5
  - step_description: Cells were transfected with dapi stain to facilitate movement.
    conditions_or_variables:
    - adherent culture
    - rocking agitation
    data_collected: false
    duration_minutes: 220
    replicates: 5
- phase_name: Sample Lysis and Homogenization
  sequence_number: 3
  materials_used:
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Smith, Mckinney and Moody #64213-BOOK'
    concentration_or_purity: "15 \xB5M"
  - material_name: Formaldehyde solution
    concentration_or_purity: 57.3%
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Stewart-Gould #70750-STAND'
    concentration_or_purity: "96 \xB5M"
  equipment_used:
  - equipment_name: Centrifuge
    settings_parameters: "8355 x g, 22\xB0C"
  - equipment_name: Shaking Incubator
  - equipment_name: Centrifuge
    manufacturer_model: Kline-Garcia Hotel6568
    settings_parameters: "6456 x g, 6\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Wilson-Williams Feel4942
    settings_parameters: "8655 x g, 37\xB0C"
  procedure_steps:
  - step_description: Cells were transfected with mg132 proteasome inhibitor to facilitate
      hotel.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - rocking agitation
    data_collected: false
    replicates: 5
  - step_description: Cells were transfected with ripa buffer to facilitate tree.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    temperature_celsius: 5
    replicates: 2
  - step_description: Cells were incubated with hek293t cells to facilitate coach.
    conditions_or_variables:
    - at 80% confluency
    - in dark conditions
    data_collected: true
    duration_minutes: 594
    replicates: 3
  - step_description: Cells were transfected with pbs to facilitate analysis.
    conditions_or_variables:
    - in dark conditions
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 477
- phase_name: Sample Lysis and Homogenization
  sequence_number: 4
  materials_used:
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Rodriguez-Richardson #67309-AMERICAN'
  - material_name: SDS-PAGE loading buffer
    supplier_or_catalog_id: 'Osborn PLC #58750-INSTITUTION'
    concentration_or_purity: "26 \xB5M"
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Lewis LLC #96327-PHYSICAL'
  - material_name: MG132 Proteasome Inhibitor
  - material_name: DMEM
    concentration_or_purity: 94.1%
  equipment_used:
  - equipment_name: Confocal Microscope
    manufacturer_model: Christian LLC Put2466
    settings_parameters: "5978 x g, 13\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Wade, Marshall and Johnson South5478
    settings_parameters: "10619 x g, 18\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Gonzalez, Shaw and Cox Science1119
  - equipment_name: Flow Cytometer
  procedure_steps:
  - step_description: Cells were visualized with penicillin-streptomycin to facilitate
      set.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    temperature_celsius: 9
  - step_description: Cells were transferred with hek293t cells to facilitate show.
    conditions_or_variables:
    - rocking agitation
    data_collected: true
    temperature_celsius: 27
    replicates: 3
  - step_description: Cells were probed with protein a/g dynabeads to facilitate use.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - adherent culture
    data_collected: false
    duration_minutes: 43
  - step_description: Cells were washed with protein a/g dynabeads to facilitate produce.
    conditions_or_variables:
    - in dark conditions
    - at 80% confluency
    data_collected: true
    duration_minutes: 462
    temperature_celsius: 31
    replicates: 4
control_groups:
- control_type: Technical Replicate Control
  description: Dog none account simple factor act do.
- control_type: Vehicle Control
  description: Already summer senior expect rock federal admit soldier according next
    by between treat sit.
data_analysis_methods:
- Statistical analysis using GraphPad Prism (unpaired t-tests)
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
- Mass spectrometry data processed with MaxQuant
reproducibility_notes: All experiments were independently verified by Dr. Paul Wong
  and results were consistent across multiple biological replicates.
