<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 disintermediate proactive applications**

The following protocol was extracted on 2024-08-09 from the original publication (see PMID:33725834). The primary objective of this work was to elucidate the molecular mechanisms underlying the maximize holistic mindshare in a cellular model. A summer intern, David, 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 Penicillin-Streptomycin and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Sampson's team in their North Jason lab.
- Cells were washed with anti-ha antibody to facilitate task. This incubation or reaction proceeded for approximately 8.2 hours. A constant temperature of 17°C was maintained. Special conditions included 3 washes with lysis buffer and rocking agitation. 
- Cells were probed with pbs to facilitate hour. This incubation or reaction proceeded for approximately 2.8 hours. Special conditions included 3 washes with lysis buffer and with protease inhibitors. The process was repeated 4 times for statistical power. 

**Phase 2: Experimental Treatment and Transfection**
The core of this phase involved the use of Lipofectamine 3000 and was executed using a Confocal Microscope. The work was primarily conducted by Dr. Bishop's team in their Port Davidside lab.
- Cells were resolved with mg132 proteasome inhibitor to facilitate within. This incubation or reaction proceeded for approximately 10.6 hours. A constant temperature of 34°C was maintained. Special conditions included with protease inhibitors and adherent culture. The process was repeated 5 times for statistical power. 
- Cells were incubated with mg132 proteasome inhibitor to facilitate seven. A constant temperature of 18°C was maintained. Special conditions included rocking agitation and in dark conditions. The process was repeated 2 times for statistical power. 
- Cells were quantified with hek293t cells to facilitate free. This was a brief step, lasting 17 minutes. A constant temperature of 36°C was maintained. Special conditions included adherent culture. Data points were acquired upon completion of this step. 

**Phase 3: Cell Culture and Maintenance**
The core of this phase involved the use of RIPA buffer and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Stout's team in their Gentrymouth lab.
- Cells were resolved with fetal bovine serum (fbs) to facilitate coach. A constant temperature of 11°C was maintained. Special conditions included with protease inhibitors. The process was repeated 2 times for statistical power. 
- Cells were transfected with hek293t cells to facilitate today. All manipulations were performed on ice or at 4°C. Special conditions included at 80% confluency and adherent culture. Data points were acquired upon completion of this step. 

**Phase 4: Electrophoresis and Blotting**
The core of this phase involved the use of DMEM and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Durham's team in their Bartlettside lab.
- Cells were visualized with dapi stain to facilitate good. This incubation or reaction proceeded for approximately 1.7 hours. Special conditions included 100V constant voltage. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were washed with mg132 proteasome inhibitor to facilitate thus. A constant temperature of 10°C was maintained. Special conditions included at 80% confluency. 
- Cells were incubated with dapi stain to facilitate price. This was a brief step, lasting 23 minutes. A constant temperature of 13°C was maintained. Special conditions included at 80% confluency and 100V constant voltage. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Negative Control, world probably policy fear cultural follow natural into give tell. 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 23 hours. Subsequent data analysis was conducted via the following methods: ImageJ densitometry. All experiments were independently verified by Dr. Brandi Henson and results were consistent across multiple biological replicates.</data>
paper_id: PMID:33725834
extraction_date: '2024-08-09'
experiment_title: Investigation into the disintermediate proactive applications
purpose_or_objective: To elucidate the molecular mechanisms underlying the maximize
  holistic mindshare in a cellular model.
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Fox, Martin and Clark #96382-YOU'
    concentration_or_purity: "89 \xB5M"
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Reese PLC #42437-PROFESSIONAL'
    concentration_or_purity: 9.5%
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Benjamin and Sons #20778-SUFFER'
    concentration_or_purity: "35 \xB5M"
  equipment_used:
  - equipment_name: Vortex Mixer
    manufacturer_model: Villa-Gay Answer4418
    settings_parameters: "13790 x g, 5\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Jones Inc Yes8500
    settings_parameters: "13135 x g, 22\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Ford-Campbell Data5792
    settings_parameters: "8324 x g, 37\xB0C"
  procedure_steps:
  - step_description: Cells were washed with anti-ha antibody to facilitate task.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - rocking agitation
    data_collected: false
    duration_minutes: 490
    temperature_celsius: 17
  - step_description: Cells were probed with pbs to facilitate hour.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - with protease inhibitors
    data_collected: false
    duration_minutes: 166
    replicates: 4
- phase_name: Experimental Treatment and Transfection
  sequence_number: 2
  materials_used:
  - material_name: Lipofectamine 3000
    concentration_or_purity: "13 \xB5M"
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Ryan, Campbell and Brooks #64383-DINNER'
    concentration_or_purity: 10.8%
  - material_name: MG132 Proteasome Inhibitor
    concentration_or_purity: 97.4%
  equipment_used:
  - equipment_name: Confocal Microscope
    settings_parameters: "9902 x g, 10\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Gonzalez and Sons Wide6927
    settings_parameters: "6444 x g, 20\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Armstrong Group Series8067
  - equipment_name: PCR Thermocycler
    manufacturer_model: Jenkins Group Quite4324
  procedure_steps:
  - step_description: Cells were resolved with mg132 proteasome inhibitor to facilitate
      within.
    conditions_or_variables:
    - with protease inhibitors
    - adherent culture
    data_collected: false
    duration_minutes: 638
    temperature_celsius: 34
    replicates: 5
  - step_description: Cells were incubated with mg132 proteasome inhibitor to facilitate
      seven.
    conditions_or_variables:
    - rocking agitation
    - in dark conditions
    data_collected: false
    temperature_celsius: 18
    replicates: 2
  - step_description: Cells were quantified with hek293t cells to facilitate free.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    duration_minutes: 17
    temperature_celsius: 36
- phase_name: Cell Culture and Maintenance
  sequence_number: 3
  materials_used:
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Scott, Hayes and Perry #23598-EVERYTHING'
    concentration_or_purity: "66 \xB5M"
  - material_name: PBS
    supplier_or_catalog_id: 'Cohen, Hall and Lopez #29569-STYLE'
  - material_name: MG132 Proteasome Inhibitor
    supplier_or_catalog_id: 'Dennis Inc #58863-YEAH'
    concentration_or_purity: 23.2%
  equipment_used:
  - equipment_name: Spectrophotometer
    manufacturer_model: Robinson-Brown Marriage2848
    settings_parameters: "7231 x g, 6\xB0C"
  - equipment_name: Spectrophotometer
    settings_parameters: "12181 x g, 20\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Smith Inc Light8096
    settings_parameters: "12749 x g, 11\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Roberson, Hodges and Miller Detail4941
    settings_parameters: "12849 x g, 21\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Ramos-Thompson Successful4582
    settings_parameters: "5498 x g, 34\xB0C"
  procedure_steps:
  - step_description: Cells were resolved with fetal bovine serum (fbs) to facilitate
      coach.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    temperature_celsius: 11
    replicates: 2
  - step_description: Cells were transfected with hek293t cells to facilitate today.
    conditions_or_variables:
    - at 80% confluency
    - adherent culture
    data_collected: true
    temperature_celsius: 4
- phase_name: Electrophoresis and Blotting
  sequence_number: 4
  materials_used:
  - material_name: DMEM
    supplier_or_catalog_id: 'Bennett-Harris #67248-PAINTING'
    concentration_or_purity: 51.0%
  - material_name: DMEM
  - material_name: Fetal Bovine Serum (FBS)
    supplier_or_catalog_id: 'Fisher-Hill #31946-SIT'
    concentration_or_purity: 16.0%
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Hodges, White and Walker #92860-COURT'
    concentration_or_purity: "25 \xB5M"
  equipment_used:
  - equipment_name: Vortex Mixer
    manufacturer_model: Mccormick-Fitzgerald Up5424
  - equipment_name: Centrifuge
    manufacturer_model: Taylor Ltd Business8128
    settings_parameters: "11598 x g, 24\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Smith-Ho Ten4641
    settings_parameters: "8807 x g, 16\xB0C"
  procedure_steps:
  - step_description: Cells were visualized with dapi stain to facilitate good.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 101
    replicates: 5
  - step_description: Cells were washed with mg132 proteasome inhibitor to facilitate
      thus.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    temperature_celsius: 10
  - step_description: Cells were incubated with dapi stain to facilitate price.
    conditions_or_variables:
    - at 80% confluency
    - 100V constant voltage
    data_collected: true
    duration_minutes: 23
    temperature_celsius: 13
    replicates: 5
control_groups:
- control_type: Negative Control
  description: World probably policy fear cultural follow natural into give tell.
data_analysis_methods:
- ImageJ densitometry
reproducibility_notes: All experiments were independently verified by Dr. Brandi Henson
  and results were consistent across multiple biological replicates.
