The iTRAQ-labeled peptide mixtures were fractionated using a Waters BEH C18 column (5 μm, 4.6 × 250 mm) with a Rigol L3000 HPLC system. The column oven temperature was set to 50 °C. Mobile phase A (2% acetonitrile adjusted to pH 10.0 using ammonium hydroxide) and mobile phase B (98% acetonitrile adjusted to pH 10.0 using ammonium hydroxide) were used to develop a gradient elution. The solvent gradient was set as follows: 3% B, 5 min; 3–8% B, 0.1 min; 8–18% B, 11.9 min; 18–32% B, 11 min; 32–45% B, 7 min; 45–80% B, 3 min; 80% B, 5 min; 80–5%, 0.1 min; and 5% B, 6.9 min. The elutes were collected every minute in a tube, and 10 fractions were pooled. All fractions were vacuum-dried and reconstituted in 0.1% (v/v) aqueous formic acid (FA) for subsequent LC–MS/MS analysis.
Beh c18 column
Manufactured by Rigol
The BEH C18 column is a high-performance liquid chromatography (HPLC) column designed for the separation and analysis of a wide range of organic compounds. It features a hybrid particle technology that combines the stability and robustness of ethylene bridged hybrid (BEH) particles with the selectivity of a C18 stationary phase.
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Lab products found in correlation
2 protocols using beh c18 column
1
Protein Fractionation and Peptide Analysis
2
iTRAQ-Based Peptide Fractionation Protocol
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The iTRAQ-labeled peptide mixtures were fractionated using a Waters BEH C18 column (5 µm, 4.6 × 250 mm) with a Rigol L3000 HPLC system. The column oven temperature was set to 50°C. Mobile phase A (2% acetonitrile adjusted to pH 10.0 using ammonium hydroxide) and mobile phase B (98% acetonitrile adjusted to pH 10.0 using ammonium hydroxide) were used to develop a gradient elution. The solvent gradient was set as follows: 3% B, 5 min; 3-8% B, 0.1 min; 8-18% B, 11.9 min; 18-32% B, 11 min; 32-45% B, 7 min; 45-80% B, 3 min; 80% B, 5 min; 80-5%, 0.1 min; and 5% B, 6.9 min. The elutes were collected every minute in a tube, and 10 fractions were pooled. All fractions were vacuum-dried and reconstituted in 0.1% (v/v) aqueous formic acid (FA) for subsequent LC-MS/MS analysis.
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