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. 2013 Apr 1;8(4):e59117. doi: 10.1371/journal.pone.0059117

Figure 1. Production and characterization of PAb.

Figure 1

(A) ELISA of PAb on ARH-77. Control rabbit IgG and PAb were incubated with ARH-77 at dilutions from 1∶2,000 to 1∶20,000. After addition of an alkaline phosphatase-conjugated secondary antibody, the absorbance was measured at 450 nm. Represented here is the mean of 4 wells to 6 wells ± standard deviation for every dilution. (B) Western blot showed the multiple protein bands recognized by PAb but not by control IgG. (C) Indirect immunofluorescence assay of PAb on myeloma and non-myeloma cell line by flow cytometry. Gray line represents 1∶2,000 PAb dilutions reacted with ARH-77 (left panel), U266 (upper part, middle panel), and Raji (upper part, right panel), human hepatocellular carcinoma cell line HepG2 (lower part, middle panel) and human pancreatic carcinoma cell line Panc-1(lower part, right panel). Black line represents control IgG diluted to 1∶2,000 used as a negative control. (D) Indirect immunofluorescence assay of antigens on ARH-77 by fluorescence microscopy with FITC-goat anti-rabbit IgG (left, green fluorescence) and with hoechst33258 (middle, blue fluorescence). Up line represents the treatment group with PAb and down line represents the treatment group with control IgG. Merged images (right) show localization of antigens on ARH-77 cells (400×).