Allophycocyanin h7 anti cd20 clone l27

Fresh peripheral blood mononuclear cells were stained with fluorescent-labelled antibodies including Pacific blue anti-CD3 (clone UCHT1, catalogue number 558117, BD) (5 μg/ml), fluorescein isothiocyanate anti-CD19 (clone HIB19, catalogue number 555412, BD) (1:10 dilution in a 100 μl experimental sample), phycoerythrin-Cy7 anti-CD27 (clone M-T271, catalogue number 560609, BD) (1:20 dilution in a 100 μl experimental sample), allophycocyanin-H7 anti-CD20 (clone L27, catalogue number 641396, BD) (5 μg/ml) and phycoerythrin-Cy5 anti- CD38 (clone HIT2, catalogue number 555461, BD) (1:10 dilution in a 100 μl experimental sample). CD3^negCD20^negCD19^posCD27^hiCD38^hi plasmablasts were gated (Supplementary Fig. 14a) and sorted as single cells. Heavy and light chain variable domains of single plasmablasts were cloned into their respective vectors and supernatants were collected from 293T cells co-transfected with the heavy chain and light chain vectors designed to express human IgG1^{13 (link)}.

Freshly separated peripheral blood mononuclear cells (PBMCs) or thawed PBMCs were stained with fluorescent-labelled antibodies to cell surface markers purchased from BD Biosciences, USA; Pacific blue anti-CD3 (clone UCHT1, Cat. No. 558117, BD), Fluorescein isothiocyanate anti-CD19 (clone HIB19, Cat. No. 555412, BD), Phycoerythrin-Cy7 anti-CD27 (clone M-T271, Cat. No. 560609, BD), Allophycocyanin-H7 anti-CD20 (clone L27, Cat. No. 641396, BD), Phycoerythrin-Cy5 anti-CD38 (clone HIT2, Cat. No. 555461, BD) and Phycoerythrin anti-human IgG (clone G18-145, Cat. No. 555787, BD). The CD3^negCD19^posCD20^negCD27^hiCD38^hiIgG^pos plasmablasts were gated and isolated in chamber as single cells as previously described [32 (link)].