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2D-Electrophoresis quiz

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  • What two techniques are combined in 2D electrophoresis?
    Isoelectric focusing (IEF) and SDS-PAGE are combined in 2D electrophoresis.
  • How does isoelectric focusing (IEF) separate proteins?
    IEF separates proteins based on their isoelectric point (pI) using a stable pH gradient.
  • What property does SDS-PAGE use to separate proteins?
    SDS-PAGE separates proteins based on their molecular weight.
  • Why is 2D electrophoresis called 'two-dimensional'?
    Because it separates proteins in two perpendicular directions: first by pI, then by molecular weight.
  • What advantage does 2D electrophoresis have over IEF or SDS-PAGE alone?
    It can separate proteins with identical pI but different molecular weights, and vice versa.
  • What happens to proteins with the same pI but different molecular weights in IEF alone?
    They appear as a single band and cannot be distinguished by IEF alone.
  • How are proteins with identical molecular weights but different pIs separated in 2D electrophoresis?
    They are separated in the first dimension by IEF, which distinguishes them by their pI.
  • What is the purpose of turning the IEF gel 90 degrees before SDS-PAGE?
    Turning the gel 90 degrees allows the second separation (by molecular weight) to occur perpendicular to the first.
  • In a 2D electrophoresis gel, what do spots in the same vertical plane represent?
    They represent proteins with the same pI but different molecular weights.
  • In a 2D electrophoresis gel, what do spots in the same horizontal plane represent?
    They represent proteins with the same molecular weight but different pIs.
  • How does the pH gradient behave during isoelectric focusing?
    The pH gradient is stable and does not move during the process.
  • Where do proteins migrate during IEF?
    Proteins migrate to the region where the pH matches their isoelectric point (pI).
  • How do proteins with high molecular weight behave in SDS-PAGE?
    They travel slower and appear toward the top of the gel.
  • Why is it important to pay attention to axis labeling in 2D electrophoresis gels?
    Because the orientation of high and low pI can vary depending on how the gel is turned.
  • What is a practical application of 2D electrophoresis in biochemistry?
    It is used to analyze and characterize proteins by separating them based on both pI and molecular weight.