PhyTip Columns Technologies
PhyTip Columns Technologies

N-glycans from serum IgG and total serum glycoproteins specific for endometriosis

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Protein G PhyTip Column 문의

Isolation of IgG

50 µL serum was used to isolate IgG. The serum IgG was captured using Protein G PhyTips (PTH 91-20-02 Box of 96 PhyTip columns) and a Buffer kit of PhyNexus (BUF-91-40-01) (Biotage, Uppsala, Sweden). Pre-equilibrated PhyTips were used for IgG capture (equilibration with Buffer A solution 200 µL per well, 20 cycles, at lowest speed; IgG capture with 200 µL Buffer A solution and 50 µL serum sample, 30 cycles, at lowest speed). The captured IgG in the PhyTip resin was washed with Wash buffer I (250 µL per well mixing volume, 10 cycles, at lowest speed) followed by a treatment of Wash buffer II (250 µL per well mixing volume, 10 cycles, at lowest speed). Then, IgG was eluted (250 µL per well mixing volume, 20 cycles, at lowest speed). Samples were neutralized with TRIS buffer, pH 11.0 (MERCK, NJ, USA). All solutions used for the process were from the Buffer kit of PhyNexus except the 0.1 M citric acid reagent, pH 2.5 used for elution (MERCK, NJ, USA), as this solution provided the best result after Peptide-N-Glycosidase F (PNGase F) treatment.