Jump to content

User:Benjah-bmm27/degree/3/CA

From Wikipedia, the free encyclopedia

The Analysis of Macromolecules

[edit]

Separation of macromolecules

[edit]
  • Size exclusion chromatography (SEC)
  • Stationary phase is a porous material called Sephadex
  • Smaller molecules can penetrate every pore of the stationary phase, so are retained longer
  • Larger molecules cannot penetrate any of the pores, so elute faster
  • YouTube animation of SEC in action: http://www.youtube.com/watch?v=wjuWb6XpRmQ
  • It is possible to calculate the approximate molecular weight of a macromolecule using SEC
    • Determine the column void volume, Vo (volume a particle sees if it cannot penetrate any pores) by flowing dextran blue (a huge molecule, m.w. 2 MDa) through the column
    • Determine elution volumes Ve of several macromolecules of known molecular weight
    • Plot a calibration curve: log(m.w.) vs. Ve/Vo
    • Measure Ve of the analyte (a macromolecule of unknown m.w.), read off m.w. from calibration curve
  • Stokes radius

Measuring the size of macromolecules

[edit]

Movement and distance

[edit]

FRET

[edit]

Visualisation

[edit]

SEM

[edit]

TEM

[edit]