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. 2013 May 7;8(5):e63414. doi: 10.1371/journal.pone.0063414

Figure 2. Target validation, PAT-SM6 reacts with cell surface expressed GRP78.

Figure 2

A, Sandwich ELISA with recombinant GRP78 and PAT-SM6. Plates were coated with GRP78, blocked and incubated with PAT-SM6 or other natural IgM antibodies as controls followed by incubation with anti-human IgM-HRP. PAT-SM6 showed binding to recombinant GRP78. B, Competitive ELISA: Plates were coated with GRP78 and increasing amounts of GRP78 or control protein were added to a solution of PAT-SM6 followed by anti-human IgM-HRP. Unbound GRP78 reduced binding of PAT-SM6 on a GRP78 coated plate. C, Surface expression GRP78 (black histogram) in MM cell lines is shown using an anti GRP78 IgG (rabbit) antibody. D, GRP78 expression in bone marrow biopsies was assessed by immunohistochemistry with anti GRP78 IgG, PAT-SM6 or positive control CD138. Primary MM cells were positive for GRP78 and the staining pattern correlates with the pattern displayed by PAT-SM6. In contrast to PAT-SM6, anti-GRP78 IgG also reacted with non-malignant cells. Images were captured using a Leica DM BL microscope, the Leica ICC HD digital camera and the Leica LAS EZ V2.1.0 software.