Peptide microarrays: A peptide library of 160,000, peptides as well other peptide microarrays for the substitutions and the KD analysis, were fabricated via the ultra-high-density peptide microarray technology of AXXELERA (Karlsruhe, Germany) [49 ]. The AXXELERA technology allows for the 30 µm-sized square peptide spots with a pitch of 60 µm that corresponds to ca. 28,000 spots/cm2.
Immunostaining: Peptide microarrays were incubated with the monoclonal anti-CD20 antibody rituximab (Sigma-Aldrich, Steinheim, Germany) with concentrations depending on the type of experiment, diluted in PBS-T (Phosphate Buffered Saline with Tween) overnight at 4 °C. A rituximab concentration of 30 µg/mL was used for the first screen of the entire resemblance-ranking peptide library. Then, the peptide chips were stained with the fluorescently labeled secondary anti-human-IgG antibodies (Jackson ImmunoResearch, Philadelphia, PA, USA). The signals from the spots were read out with the confocal scanner Innoscan 1100 AL. The scans were performed in the red channel (635 nm), with a resolution of 2 µm/pixel, a speed of 35 µm/s, and a PMT gain of 9.
Immunostaining: Peptide microarrays were incubated with the monoclonal anti-CD20 antibody rituximab (Sigma-Aldrich, Steinheim, Germany) with concentrations depending on the type of experiment, diluted in PBS-T (Phosphate Buffered Saline with Tween) overnight at 4 °C. A rituximab concentration of 30 µg/mL was used for the first screen of the entire resemblance-ranking peptide library. Then, the peptide chips were stained with the fluorescently labeled secondary anti-human-IgG antibodies (Jackson ImmunoResearch, Philadelphia, PA, USA). The signals from the spots were read out with the confocal scanner Innoscan 1100 AL. The scans were performed in the red channel (635 nm), with a resolution of 2 µm/pixel, a speed of 35 µm/s, and a PMT gain of 9.