Capillary lc nano esi ms

Peptide mapping was performed according to previous methods [23 (link)]. Protein bands obtained from SDS-PAGE were excised, in-gel digested with trypsin, and cleaned with Zip-Tip (Millipore, Billerica, MA, USA). Mass spectrometry analyses were performed using Capillary LC-Nano ESI-MS with a 6545 Q-TOF LC/MS (Agilent Technologies, Santa Clara, CA, USA). Samples were applied to a ZORBAX 300SB-C8 column (1 × 50 mm, 3.5 μm; Agilent) equilibrated with 0.1% (v/v) formic acid in mass grade water and eluted by a gradient between water and 100% acetonitrile at a flow rate of 10 µL/min. The tuning parameters used for mass analyses were as follows: capillary temperature 300 °C, source voltage 1.9 kV, skimmer voltage 45 V, and fragmentor voltage 175 V.

Protein bands obtained by SDS-PAGE were excised, in-gel digested with trypsin, and cleaned with Zip-Tip (Millipore, Billerica, MA, USA). Mass analyses were performed using Capillary LC-Nano ESI-MS with a 6545 Q-TOF LC/MS (Agilent Technologies, Santa Clara, CA, USA). Samples were applied to a ZORBAX 300SB-C8 column (1 × 50 mm, 3.5 μm; Agilent) equilibrated with 0.1% (v/v) formic acid in mass grade water and eluted by a gradient between water and 100% acetonitrile at a flow rate of 10 µL/min. The tuning parameters used for mass analyses were as follows: capillary temperature 300 °C, source voltage 1.9 kV, skimmer voltage 45 V, and fragmentor voltage 175 V.