The dried IGPs were resuspended in 2% ACN/20 mM NH4COOH and fractionated through high pH reverse-phase high-performance liquid chromatography with Agilent RP Zorbax 300 Å Extend C18 column (250 mm × 4.6 mm, OD 5 μm) in an Agilent 1260 LC instrument. Intact glycopeptides were first separated into 96 fractions through a gradient of mobile phases A (2% ACN/5 mM NH4COOH, pH 10) and B (90% ACN/5 mM NH4COOH, pH 10) at a flow rate of 0.2 ml/min. A total of 120 min was used for gradient elution, 0 to 2% B for 7 min, 2 to 8% B for 35 min, 8 to 16% B for 35 min, 16 to 35% B for 10 min, 35 to 95% B for 5 min, 95 to 95% B for 28 min. Then, IGPs were integrated into 12 fractions and dried by vacuum centrifuging.
1260 lc instrument
Manufactured by Agilent Technologies
Sourced in United States
The 1260 LC instrument is a liquid chromatography system designed for analytical applications. It is capable of performing high-performance liquid chromatography (HPLC) and ultra-high-performance liquid chromatography (UHPLC) separations. The instrument includes a solvent delivery system, an autosampler, a column compartment, and a detector for data acquisition.
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Lab products found in correlation
2 protocols using 1260 lc instrument
1
Glycopeptide Fractionation by HPLC
2
Polysaccharide Characterization via HPGPC
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The homogeneity and molecular weight distribution of polysaccharide fractions were determined by HPGPC on an Agilent 1260 LC instrument equipped with a refractive index detector (RID), a variable wavelength detector (VWD), coupled with an Ultrahydrogel TM 1000 column (7.8 mm × 300 mm, Waters, USA) and an Ultrahydrogel TM Linear column (7.8 mm × 300 mm, Waters, USA).
Polysaccharide solution was filtered through 0.45 μm filter prior to injection, with 0.1 M NaCl/0.02% NaN3 aqueous solution as mobile phase at a flow rate of 0.6 mL/min. The molecular weight of polysaccharides was estimated using a standard curve prepared by T-series dextrans.
According to the information obtained from HPGPC that would be discussed later in this study, we selected NCSP-50 for the following analysis.
Polysaccharide solution was filtered through 0.45 μm filter prior to injection, with 0.1 M NaCl/0.02% NaN3 aqueous solution as mobile phase at a flow rate of 0.6 mL/min. The molecular weight of polysaccharides was estimated using a standard curve prepared by T-series dextrans.
According to the information obtained from HPGPC that would be discussed later in this study, we selected NCSP-50 for the following analysis.
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