Protein Protocols Protein Protocols

908 Hooker and James
Fig. 1. Whole recombinant human IFN-γ analyzed by capillary electrophoresis. Recombinant
human IFN-γ glycoforms were “fingerprinted” by micellar electrokinetic capillary chromatography.
Peak groups represent IFN-γ variants with both Asn sites occupied (2N), one site occupied
(1N), or no sites occupied (0N).
4. Voltages are applied over a 0.2-min linear ramping period at a detection wavelength
of 200 nm and operating temperature of 25°C. Recombinant human IFN-γ (1 mg/mL in
50 mM borate, 50 mM SDS, pH 8.5) is injected for 5 s prior to electrophoresis at 22 kV.
Between each separation, the capillary is rinsed with 0.1 M NaOH, water, and electrophoresis
buffer for 5 min, respectively (see Note 1).
3.2. Glycosylation Analysis by MALDI-MS
There are only a few reports of the analysis of whole glycoproteins due to the limited
resolution of this technique (18). As a result, analysis of intact glycoproteins is generally
limited to those proteins that contain one glycosylation site and are