Lc ms grade acetonitrile acn

SILuMAb universal antibody standard human (MSQC4) was purchased from Merck, Darmstadt, Germany. DEPC was obtained from Acros Organics, Geel, Belgium (Product no. 10114380). Imidazole (Product no. I2399), urea (Product no. U5378), ammonium bicarbonate (ABC; Product no. 09830), iodoacetamide (IAA) (Product no. I6125) and C18 disks- 47 mm (Product no. 66883-U) were purchased from Merck. Tris (2-carboxyethyl) phosphine (TCEP) was obtained from Alfa Aesar, Karlsruhe, Germany, and PNGaseF was purchased from Serva, Heidelberg, Germany. Sequencing grade modified trypsin (product no. V5111) was procured from Promega (Madison, WI, USA), formic acid (FA), and LC-MS grade water from VWR (Darmstadt, Germany). LC-MS grade acetonitrile (ACN) and methanol were purchased from Honeywell (Muskegon, MI, USA). Sodium chloride was obtained from Chemsolute, Renningen, Germany, and monobasic sodium phosphate from AppliChem, Darmstadt, Germany.

Subclass specific analysis of IgG Fc glycosylation included 2,579 in dividuals from 14 different countries and 25 different ethnic groups (Supplementary Table 6). We studied glycopeptides from three IgG sublessees; IgG1, IgG2 and IgG4 and each subclass was studied for various glycan features (Supplementary Tables 15, 16).
IgG glycopeptides were obtained and purified as described before [49 (link)]. Approximately 15 μg of isolated IgG was treated with 0.1 μg of sequencing grade trypsin (Promega, Fitchburg, WI) and incubated overnight at 37 °C. The reaction was stopped by dilution with 0.1 % trifluoroacetic acid (TFA; Sigma-Aldrich, St. Louis, MI). Glycopeptides were purified using a solid-phase extraction on Chromabond C-18 sorbent (Macherey-Nagel, Düren, Germany). Samples were loaded onto beads in 0.1 % TFA and washed three times using the same solvent. Glycopeptides were eluted from the phase with 20 % LC-MS grade acetonitrile (ACN; Honeywell, Morris Plains, NJ). Eluted glycopeptides were vacuum-dried and reconstituted in 20 μL of ultrapure water prior to LC-MS analysis. All glycan analyses were performed at Genos laboratory.