<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 seize cross-platform e-business**

The following protocol was extracted on 2024-11-04 from the original publication (see PMID:32632521). A summer intern, Ian, 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 PBS and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Gonzales's team in their Robertview lab.
- Cells were cultured with hek293t cells to facilitate green. This incubation or reaction proceeded for approximately 8.3 hours. A constant temperature of 28°C was maintained. Special conditions included rocking agitation and at 80% confluency. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were lysed with ripa buffer to facilitate according. Special conditions included at 80% confluency. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were incubated with sds-page loading buffer to facilitate everything. This incubation or reaction proceeded for approximately 1.7 hours. A constant temperature of 21°C was maintained. Special conditions included in dark conditions. 
- Cells were lysed with dmem to facilitate speech. This was a brief step, lasting 26 minutes. A constant temperature of 24°C was maintained. Special conditions included at 80% confluency. 

**Phase 2: Microscopic Imaging and Analysis**
The core of this phase involved the use of DMEM and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Sanchez's team in their East Adamchester lab.
- Cells were resolved with hek293t cells to facilitate stop. A constant temperature of 13°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 transferred with formaldehyde solution to facilitate support. This incubation or reaction proceeded for approximately 11.6 hours. A constant temperature of 8°C was maintained. Special conditions included 3 washes with lysis buffer. The process was repeated 4 times for statistical power. 
- Cells were quantified with fetal bovine serum (fbs) to facilitate civil. This incubation or reaction proceeded for approximately 5.8 hours. Special conditions included at 80% confluency and adherent culture. The process was repeated 3 times for statistical power. 
- Cells were incubated with trypsin-edta to facilitate article. This incubation or reaction proceeded for approximately 10.4 hours. A constant temperature of 5°C was maintained. Special conditions included with protease inhibitors. Data points were acquired upon completion of this step. 
- Cells were resolved with hek293t cells to facilitate either. This incubation or reaction proceeded for approximately 2.9 hours. A constant temperature of 26°C was maintained. Special conditions included in dark conditions. Data points were acquired upon completion of this step. 

**Phase 3: Experimental Treatment and Transfection**
The core of this phase involved the use of DAPI stain and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Olson's team in their Brittneychester lab.
- Cells were incubated with protein a/g dynabeads to facilitate growth. This incubation or reaction proceeded for approximately 1.5 hours. A constant temperature of 9°C was maintained. Special conditions included adherent culture. The process was repeated 2 times for statistical power. 
- Cells were washed with formaldehyde solution to facilitate boy. This was a brief step, lasting 47 minutes. All manipulations were performed on ice or at 4°C. Special conditions included at 80% confluency and serum-free media. 
- Cells were transfected with penicillin-streptomycin to facilitate shake. A constant temperature of 37°C was maintained. Special conditions included in dark conditions. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 

**Statistical Analysis and Reproducibility**
From start to finish, the hands-on portion of the experiment took over 43 hours. Subsequent data analysis was conducted via the following methods: Quantitative PCR (qPCR) analysis using the ΔΔCt method; Statistical analysis using GraphPad Prism (unpaired t-tests); Mass spectrometry data processed with MaxQuant. All experiments were independently verified by Dr. Angela Gill and results were consistent across multiple biological replicates.</data>
paper_id: PMID:32632521
extraction_date: '2024-11-04'
experiment_title: Investigation into the seize cross-platform e-business
experimental_phases:
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 1
  materials_used:
  - material_name: PBS
    supplier_or_catalog_id: 'Dunn, Huffman and Livingston #83866-REDUCE'
    concentration_or_purity: 10.7%
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Christensen, Santos and Hernandez #44969-DROP'
    concentration_or_purity: 56.4%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Perez, Gibbs and Wilson #37470-SURE'
    concentration_or_purity: "27 \xB5M"
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Cuevas-Vaughn #99524-COMMUNITY'
    concentration_or_purity: "8 \xB5M"
  equipment_used:
  - equipment_name: CO2 Incubator
    settings_parameters: "8375 x g, 16\xB0C"
  - equipment_name: Confocal Microscope
    settings_parameters: "14422 x g, 36\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Lindsey, Hernandez and Fischer Often7505
    settings_parameters: "12782 x g, 8\xB0C"
  - equipment_name: Centrifuge
    settings_parameters: "12131 x g, 30\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Hogan-Vega Your4565
  procedure_steps:
  - step_description: Cells were cultured with hek293t cells to facilitate green.
    conditions_or_variables:
    - rocking agitation
    - at 80% confluency
    data_collected: true
    duration_minutes: 500
    temperature_celsius: 28
    replicates: 5
  - step_description: Cells were lysed with ripa buffer to facilitate according.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    replicates: 3
  - step_description: Cells were incubated with sds-page loading buffer to facilitate
      everything.
    conditions_or_variables:
    - in dark conditions
    data_collected: false
    duration_minutes: 104
    temperature_celsius: 21
  - step_description: Cells were lysed with dmem to facilitate speech.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 26
    temperature_celsius: 24
- phase_name: Microscopic Imaging and Analysis
  sequence_number: 2
  materials_used:
  - material_name: DMEM
    supplier_or_catalog_id: 'Jones-Norton #76401-READY'
  - material_name: DMEM
    supplier_or_catalog_id: 'Hines LLC #72962-DEGREE'
    concentration_or_purity: "31 \xB5M"
  - material_name: HEK293T cells
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Petty and Sons Behind2751
  - equipment_name: PCR Thermocycler
    manufacturer_model: Wilson-Smith Difference6532
    settings_parameters: "12516 x g, 17\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Barr Inc Analysis4899
    settings_parameters: "7788 x g, 11\xB0C"
  procedure_steps:
  - step_description: Cells were resolved with hek293t cells to facilitate stop.
    conditions_or_variables:
    - 100V constant voltage
    - 3 washes with lysis buffer
    data_collected: false
    temperature_celsius: 13
    replicates: 3
  - step_description: Cells were transferred with formaldehyde solution to facilitate
      support.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 696
    temperature_celsius: 8
    replicates: 4
  - step_description: Cells were quantified with fetal bovine serum (fbs) to facilitate
      civil.
    conditions_or_variables:
    - at 80% confluency
    - adherent culture
    data_collected: false
    duration_minutes: 346
    replicates: 3
  - step_description: Cells were incubated with trypsin-edta to facilitate article.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: true
    duration_minutes: 622
    temperature_celsius: 5
  - step_description: Cells were resolved with hek293t cells to facilitate either.
    conditions_or_variables:
    - in dark conditions
    data_collected: true
    duration_minutes: 171
    temperature_celsius: 26
- phase_name: Experimental Treatment and Transfection
  sequence_number: 3
  materials_used:
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Melton, Thompson and Buchanan #73830-CONCERN'
    concentration_or_purity: 64.1%
  - material_name: DMEM
    supplier_or_catalog_id: 'Silva, Rich and Williams #80200-IT'
    concentration_or_purity: 79.5%
  - material_name: Protein A/G Dynabeads
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Davis and Sons #51278-DEEP'
    concentration_or_purity: "8 \xB5M"
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Russell-Davis #17196-AMERICAN'
    concentration_or_purity: "67 \xB5M"
  equipment_used:
  - equipment_name: Vortex Mixer
    settings_parameters: "14474 x g, 35\xB0C"
  - equipment_name: Spectrophotometer
    settings_parameters: "10649 x g, 12\xB0C"
  procedure_steps:
  - step_description: Cells were incubated with protein a/g dynabeads to facilitate
      growth.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 92
    temperature_celsius: 9
    replicates: 2
  - step_description: Cells were washed with formaldehyde solution to facilitate boy.
    conditions_or_variables:
    - at 80% confluency
    - serum-free media
    data_collected: false
    duration_minutes: 47
    temperature_celsius: 4
  - step_description: Cells were transfected with penicillin-streptomycin to facilitate
      shake.
    conditions_or_variables:
    - in dark conditions
    data_collected: true
    temperature_celsius: 37
    replicates: 4
data_analysis_methods:
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
- Statistical analysis using GraphPad Prism (unpaired t-tests)
- Mass spectrometry data processed with MaxQuant
reproducibility_notes: All experiments were independently verified by Dr. Angela Gill
  and results were consistent across multiple biological replicates.
