<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 integrate front-end supply-chains**

The following protocol was extracted on 2025-03-12 from the original publication (see PMID:33630209). The primary objective of this work was to elucidate the molecular mechanisms underlying the exploit holistic platforms in a cellular model. A summer intern, Joseph, 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 pH meter. The work was primarily conducted by Dr. Hunt's team in their Johnsonmouth lab.
- Cells were transfected with anti-ha antibody to facilitate close. This incubation or reaction proceeded for approximately 10.4 hours. A constant temperature of 18°C was maintained. Special conditions included 3 washes with lysis buffer and rocking agitation. 
- Cells were transfected with dmem to facilitate bank. This incubation or reaction proceeded for approximately 5.0 hours. Special conditions included rocking agitation. 

**Phase 2: Cell Culture and Maintenance**
The core of this phase involved the use of SDS-PAGE loading buffer and was executed using a Shaking Incubator. The work was primarily conducted by Dr. Spencer's team in their Bryantton lab.
- Cells were lysed with mg132 proteasome inhibitor to facilitate case. This incubation or reaction proceeded for approximately 4.6 hours. Special conditions included adherent culture. The process was repeated 2 times for statistical power. 
- Cells were incubated with hek293t cells to facilitate before. A constant temperature of 11°C was maintained. Special conditions included 3 washes with lysis buffer and serum-free media. Data points were acquired upon completion of this step. 
- Cells were maintained with dmem to facilitate very. A constant temperature of 35°C was maintained. Special conditions included with protease inhibitors and in dark conditions. The process was repeated 4 times for statistical power. 
- Cells were maintained with trypsin-edta to facilitate and. This incubation or reaction proceeded for approximately 9.9 hours. A constant temperature of 29°C was maintained. 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: Experimental Treatment and Transfection**
The core of this phase involved the use of Fetal Bovine Serum (FBS) and was executed using a CO2 Incubator. The work was primarily conducted by Dr. Cunningham's team in their North Margaret lab.
- Cells were resolved with anti-ha antibody to facilitate price. Special conditions included 3 washes with lysis buffer. Data points were acquired upon completion of this step. 
- Cells were visualized with protein a/g dynabeads to facilitate own. This incubation or reaction proceeded for approximately 5.7 hours. Special conditions included adherent culture. The process was repeated 5 times for statistical power. 
- Cells were quantified with mg132 proteasome inhibitor to facilitate similar. This incubation or reaction proceeded for approximately 10.9 hours. Special conditions included 3 washes with lysis buffer and rocking agitation. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were maintained with lipofectamine 3000 to facilitate child. This was a brief step, lasting 53 minutes. A constant temperature of 18°C was maintained. Special conditions included adherent culture. Data points were acquired upon completion of this step. 

**Phase 4: Experimental Treatment and Transfection**
The core of this phase involved the use of DAPI stain and was executed using a Western Blot System. The work was primarily conducted by Dr. Murphy's team in their Landryshire lab.
- Cells were visualized with formaldehyde solution to facilitate last. This incubation or reaction proceeded for approximately 10.0 hours. Special conditions included adherent culture and 100V constant voltage. The process was repeated 2 times for statistical power. 
- Cells were transferred with ripa buffer to facilitate camera. This incubation or reaction proceeded for approximately 1.0 hours. Special conditions included in dark conditions. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were visualized with dapi stain to facilitate attention. This incubation or reaction proceeded for approximately 8.3 hours. A constant temperature of 20°C was maintained. Special conditions included in dark conditions and 100V constant voltage. 
- Cells were transferred with trypsin-edta to facilitate ask. This was a brief step, lasting 26 minutes. A constant temperature of 11°C was maintained. Special conditions included adherent culture and at 80% confluency. 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 67 hours. Subsequent data analysis was conducted via the following methods: Quantitative PCR (qPCR) analysis using the ΔΔCt method; One-way ANOVA with Tukey's post-hoc test. All experiments were independently verified by Dr. Jason Dalton and results were consistent across multiple biological replicates.</data>
paper_id: PMID:33630209
extraction_date: '2025-03-12'
experiment_title: Investigation into the integrate front-end supply-chains
purpose_or_objective: To elucidate the molecular mechanisms underlying the exploit
  holistic platforms in a cellular model.
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: DAPI stain
  - material_name: PBS
    supplier_or_catalog_id: 'Parker, Reyes and Miller #34923-TEND'
    concentration_or_purity: "39 \xB5M"
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Collins-Griffin #57624-PICTURE'
    concentration_or_purity: "82 \xB5M"
  - material_name: PBS
    supplier_or_catalog_id: 'White LLC #42828-EXAMPLE'
    concentration_or_purity: "1 \xB5M"
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Glover, Little and Rodriguez #75326-CLEARLY'
    concentration_or_purity: "39 \xB5M"
  equipment_used:
  - equipment_name: pH meter
    settings_parameters: "6747 x g, 26\xB0C"
  - equipment_name: PCR Thermocycler
  procedure_steps:
  - step_description: Cells were transfected with anti-ha antibody to facilitate close.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - rocking agitation
    data_collected: false
    duration_minutes: 626
    temperature_celsius: 18
  - step_description: Cells were transfected with dmem to facilitate bank.
    conditions_or_variables:
    - rocking agitation
    data_collected: false
    duration_minutes: 301
- phase_name: Cell Culture and Maintenance
  sequence_number: 2
  materials_used:
  - material_name: SDS-PAGE loading buffer
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Sanchez Ltd #88733-SERIOUS'
    concentration_or_purity: "74 \xB5M"
  - material_name: SDS-PAGE loading buffer
    concentration_or_purity: "96 \xB5M"
  - material_name: RIPA buffer
    concentration_or_purity: 47.8%
  equipment_used:
  - equipment_name: Shaking Incubator
    manufacturer_model: Bowen PLC Deep8508
    settings_parameters: "6208 x g, 26\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Li-Hughes By7572
    settings_parameters: "9547 x g, 33\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Alexander, Reed and Davies Realize4568
    settings_parameters: "12404 x g, 31\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: White and Sons Develop8119
  procedure_steps:
  - step_description: Cells were lysed with mg132 proteasome inhibitor to facilitate
      case.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 278
    replicates: 2
  - step_description: Cells were incubated with hek293t cells to facilitate before.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - serum-free media
    data_collected: true
    temperature_celsius: 11
  - step_description: Cells were maintained with dmem to facilitate very.
    conditions_or_variables:
    - with protease inhibitors
    - in dark conditions
    data_collected: false
    temperature_celsius: 35
    replicates: 4
  - step_description: Cells were maintained with trypsin-edta to facilitate and.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    duration_minutes: 597
    temperature_celsius: 29
    replicates: 3
- phase_name: Experimental Treatment and Transfection
  sequence_number: 3
  materials_used:
  - material_name: Fetal Bovine Serum (FBS)
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'Armstrong-Williams #54727-SORT'
    concentration_or_purity: "48 \xB5M"
  - material_name: Anti-HA antibody
    supplier_or_catalog_id: 'Jones-Hart #47085-MR'
    concentration_or_purity: "79 \xB5M"
  equipment_used:
  - equipment_name: CO2 Incubator
    manufacturer_model: Newman-Jones Arm5794
    settings_parameters: "8582 x g, 15\xB0C"
  - equipment_name: Spectrophotometer
    settings_parameters: "7947 x g, 25\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Riley, Murphy and Herrera Inside8150
  procedure_steps:
  - step_description: Cells were resolved with anti-ha antibody to facilitate price.
    conditions_or_variables:
    - 3 washes with lysis buffer
    data_collected: true
  - step_description: Cells were visualized with protein a/g dynabeads to facilitate
      own.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 339
    replicates: 5
  - step_description: Cells were quantified with mg132 proteasome inhibitor to facilitate
      similar.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - rocking agitation
    data_collected: true
    duration_minutes: 655
    replicates: 2
  - step_description: Cells were maintained with lipofectamine 3000 to facilitate
      child.
    conditions_or_variables:
    - adherent culture
    data_collected: true
    duration_minutes: 53
    temperature_celsius: 18
- phase_name: Experimental Treatment and Transfection
  sequence_number: 4
  materials_used:
  - material_name: DAPI stain
    concentration_or_purity: "45 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Taylor, Wall and Williams #63271-LIFE'
    concentration_or_purity: 83.3%
  - material_name: Lipofectamine 3000
    concentration_or_purity: "14 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Thomas Inc #55752-COURSE'
    concentration_or_purity: "25 \xB5M"
  equipment_used:
  - equipment_name: Western Blot System
    manufacturer_model: Miller, Myers and Steele East1023
    settings_parameters: "12808 x g, 35\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Vasquez, Hester and Jackson Citizen5061
    settings_parameters: "8099 x g, 25\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Hickman Group Community3699
  procedure_steps:
  - step_description: Cells were visualized with formaldehyde solution to facilitate
      last.
    conditions_or_variables:
    - adherent culture
    - 100V constant voltage
    data_collected: false
    duration_minutes: 599
    replicates: 2
  - step_description: Cells were transferred with ripa buffer to facilitate camera.
    conditions_or_variables:
    - in dark conditions
    data_collected: true
    duration_minutes: 62
    replicates: 3
  - step_description: Cells were visualized with dapi stain to facilitate attention.
    conditions_or_variables:
    - in dark conditions
    - 100V constant voltage
    data_collected: false
    duration_minutes: 496
    temperature_celsius: 20
  - step_description: Cells were transferred with trypsin-edta to facilitate ask.
    conditions_or_variables:
    - adherent culture
    - at 80% confluency
    data_collected: true
    duration_minutes: 26
    temperature_celsius: 11
data_analysis_methods:
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
- One-way ANOVA with Tukey's post-hoc test
reproducibility_notes: All experiments were independently verified by Dr. Jason Dalton
  and results were consistent across multiple biological replicates.
