Long-term precision in capillary isoelectric focusing for protein analysis

✍ Scribed by Adhitasari Suratman; Hermann Wätzig

• Publisher: John Wiley and Sons
• Year: 2008
• Tongue: English
• Weight: 964 KB
• Volume: 31
• Category: Article
• ISSN: 1615-9306
• DOI: 10.1002/jssc.200700678

✦ Synopsis

Abstract

In order to improve precision in protein analysis, a new rinsing procedure with 3M hydrochloric acid was investigated for linear polyacrylamide‐coated capillaries used in isoelectric focusing. After each run the capillaries were rinsed with hydrochloric acid for 5 min, followed by water for 20 min (Δ__p__ = 1030 mbar each). Myoglobin, β‐lactoglobulin, and ovalbumin were used as model proteins; the pH gradient was provided by Pharmalyte (pH 3–10). The resulting method was already successful in avoiding capillary blockages, even for long‐series protein measurements. Further improvements in precision have been obtained by avoiding a complete standstill of liquid within the capillary when the separation system was idle. Pressure (Δ__p__ = 300 mbar or more) and high voltage (30 kV) were therefore also applied during storage within a measurement series. The reproducibility of migration time and peak area are further improved with RSD% less than 10% in a long‐term measurement (n = 86).