Long-term precision in capillary isoelectric focusing for protein analysis.
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 (Deltap = 1030 mbar each). Myoglobin, beta-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 (Deltap = 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).
Suratman
Adhitasari
A
Institute of Pharmacy, TU Braunschweig, Braunschweig, Germany. Fax: +49-531-391-2799.
Wätzig
Hermann
H
D000818:Animals; D002417:Cattle; D002623:Chemistry Techniques, Analytical / Q000379:methods; D002645:Chickens; D019075:Electrophoresis, Capillary / Q000379:methods*; D006736:Horses; D006851:Hydrochloric Acid / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing / Q000379:methods*; D007782:Lactoglobulins / Q000737:chemistry; D009211:Myoglobin / Q000737:chemistry; D010047:Ovalbumin / Q000737:chemistry; D011312:Pressure; D011506:Proteins / Q000032:analysis* / Q000737:chemistry*; D014867:Water / Q000737:chemistry
D007782:Lactoglobulins; D009211:Myoglobin; D011506:Proteins; D014867:Water; D010047:Ovalbumin; D006851:Hydrochloric Acid