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Capillary Electrophoresis - Coggle Diagram

- - - - Consisting of large molecules or ultramicroscopic particles of one substance dispersed through a second substance
  - - - Sample applied broadly, not in concentrated zone
      - Significant heating of electrolyte - convection currents cause mixing
  - - - Agarose
      - Polyacrylamide
      - Gels formed in tubes + slabs
      - Wells provide sample application zones
      - Stacking in SDS-PAGE concentrates samples
      - Analytes separated into distinct zones or bands
    - - Capillary developed
        
        Narrow tubes have low conductance (generate less heat)
        
        Anti-conductive
        
        Separation occurs in narrow bore fused-silica capillary
        
        Anti-convective media not needed
        
        Internal diameter 25-75\(\mu\)m
- - - - General cathodic migration of all species
  - - - Electrophoretic mobility
      - Electroosmotic mobility
    - - Cations are attracted to cathode at the negative end of capillary
      - Anions attracted to anode at positive end
      - Depends on charge and size
        
        Neutrals are unaffected by electric field + do not move
      - Electroosmotic flow (EOF) results from general flow of solvent to cathode end
        
        Cations tend to 'drag' solvent along as they move
        
        Bulk solvent flow results , carrying all species in this order - cations, neutrals, anions
        
        2 layers
        
        Fixed/stern
        
        Diffuse
        
        When high voltage electric field applied, diffuse layer cations attracted to cathode
        
        Hydrated so when they move they drag water, inducing EOF
        
        Potential difference forms between the layers - across the plane of shear (zeta or \(\zeta\)
        
        EOF is proportional to magnitude of \(\zeta\)
        
        Increases with pH
        
        Suppressed by increasing ionic strength
        
        Influence on capillary zone electrophoresis (CZE)
        
        +ve charge cations move towards cathode by electrostatic attraction
        
        Neutral + -ve charged components drawn through capillary by EOF
        
        Anions separated based on relative migration variance
        
        Balance of cathodal EOF and anodal electrostatic attraction
      - EOF influenced by chemistry of walls
        
        Silanoyl groups readily ionised above pH 3
        
        Extent increases with pH
        
        EOF significant above pH 4
        
        Anionic Si-O\(^-\) groups attract buffer cations
        
        Form double electrical layer at capillary surface
- - - - Dissipates with length
  - - - Similar molecules start to move at diff. rates
      - Peaks broaden
    - - Less pronounced peak broadening
      - Better resolution
  - - - Neutral species
      - Weak acids and bases
- - - - Lumbar spine
      - Skull
      - Long bones
      - Pelvis
  - - - Pepper pot skull on x-ray
  - - - Serum and/or urine paraprotein
      - Osteolytic lesions on x-ray
      - 25% plasma cells in bone marrow (or clonal population)
  - - - CRP
        
        Complement
        
        Transferrin
        
        Hemopexin
        
        \(\beta\)-lipoprotein
        
        \(\alpha_2\)-macroglobulin
        
        1 more item...
  - - - IgG >20g/L
      - IgA/M >10g/L
      - IgD/E paraprotein at any conc.
      - Serum protein electrophoresis detects abnormal levels of immunoglobulin
      - Quantification of paraprotein by densitometry
        
        Relate density of band to measured total protein
      - By turbidi