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Affinity Capillary Electrophoresis 329
2. The linear dynamic range of the detector is decreased when using buffers with high
UV-absorbance. The result is a decrease in peak height, resolution and increase in
noise in the electropherogram.
3. ESI is a mild ionization method compared with fast atom bombardment. ESI
facilitates characterization of non-covalent molecular complexes in the gas phase.
4. To maintain the electrical circuit at the MS electrospray source, addition of surplus
electrolyte is crucial for the liquid junction and coaxial sheath–flow interface. The
analyte thus is diluted, and this gives a decrease in detection sensitivity as well as
an interference with the resolution of the CE separation.
5. Heparins are strongly anionic, highly sulphated glycosaminoglycans. Their charges
make them ideal for use as ligands in ACE. Heparin preparations contain mixtures
of polymers of different chain length and of different sulphation and carboxylation
(98). Heparin from bovine lung represents one of the most highly sulphated types
(Dorothe Spillmann, personal communication).
6. CE buffers should routinely be prepared using deionized water (of Milli Q quality)
and should be filtered through 0.22-pore size filters (e.g. cellulose acetate filter
system (Corning 430767)) before use. The buffers usually can then be kept at 4ºC
for months.
8. Other useful binding buffers are
(a) Isotonic borate, pH 7.4
(A) 10ml0.05MNa 2 B 4 O 7 (19.11 g/l of Na 2 B 4 O 7 ·10 H 2 O)
(B) 90ml0.2MHBO 4 (12.40 g/l)
(C) 270 mg NaCl
(b) To scan for analyte recovery at a range of electrophoresis buffer pH values
(31), borate buffers of the following compositions may be helpful:
pH 6.8: 3 ml (A), 97 ml (B), 270 mg (C)
pH 7.8: 20ml (A), 80 ml (B), 260 mg (C)
pH 8.1: 30 ml (A), 70 ml (B), 240 mg (C)
pH 8.4: 45 ml (A), 55 ml (B), 210 mg (C)
pH 8.6: 55 ml (A), 45 ml (B), 190 mg (C)
pH 8.8: 70 ml (A), 30 ml (B), 140 mg (C)
pH 9.1: 90 ml (A), 10 ml (B), 70 mg (C)
(c) HEPES, pH 7.4: 10 mM N-2-hydroxyethylpiperazine-N´-ethanesulphonic acid
(HEPES) (2.38 g/l), adjusted with NaOH to pH 7.4, 150 mM NaCl (8.77 g/l).
(d) Tricine, pH 8.15: This buffer will absorb strongly at 200 nm, 20 mM N-
Tris(hydroxymethyl)methylglycine (Tricine) (3.58 g/l) adjusted with NaOH
to pH 8.15, 150 mM NaCl (8.77 g/l).
(e) Tris-buffered saline, pH 7.4: This buffer will absorb strongly at 200 nm. 5 mM
Tris(hydroxymethyl)aminomethane (Tris) (0.61 g/l) adjusted with HCl to pH
7.4, 150 mM NaCl (8.77 g/l)