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1. Factors influenced
electrophoresis mobility:
2. net charge of the
molecule
3. size and shape
4. concentration of the
molecule in solution
Amino acids have characteristic titration curves
Proton Proton
donor acceptor
Proton Proton
Fully protonated donor acceptor
form at wery low pH
Adopted from: D.L. Nelson, M.M. Cox Lehninger Principle of Biochemistry
Electrophoresis is carried out by applying a thin
layer
Electrophoresis
Densitometer tracing
density of zones is proportional
to the amount of protein
Example of application of zone electrophoresis in
clinical practice
Hypergamaglobulinemia
Hypogamaglobulinemia
Normal serum
Gel electrophoresis
Acrylamide (CH2=CH-CO-
NH2)
Polyacrylamide gel structure
held together by covalent
cross-links
Agarose gels
OH
CH2OHO O
O O
O OH
OH
O
Polyacrylamide gels
CH2=CHCONH2 + CH2(NHCOHC=CH2)2
Acrylamide N,N,N,N-methylenebisacrylamide
Ponceau S 1-2 mg
Amido Black 1-2 mg
Coomassie Blue 1.5 mg
India Ink 100 ng
Silver stain 10 ng
Colloidal gold 3 ng
Staining with Coomasie blue
1 2 3
3
An ethidium-stained gel photographed under UV light