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. 2021 Oct 23;22(21):11452. doi: 10.3390/ijms222111452

Figure 5.

Figure 5

Specific uptake of anti-uPARAP mAb in MM cells. (a) Analysis of mAb uptake by flow cytometry. Cells were analyzed after uptake of Alexa Fluor 647 conjugated mAb 9b7 (AF647-9b7) against uPARAP. The AF647-9b7 uptake is shown as histograms (in pink) for all of the cell lines examined (MM cell lines ONE58, JL-1 and H-Meso-1 and osteosarcoma lines 143B +/+ and 143B -/-). Histograms for cells without antibody (orange) and with cells pre-incubated with competing anti-uPARAP mAb 5f4 before the addition of AF647-9b7 (light blue) are included as controls. The bar chart shows the mean fluorescence intensity of the analyzed samples (MFI ± SD). (b) Internalization of anti-uPARAP mAb in H-Meso-1, JL-1 and ONE58 cells, shown by confocal imaging. Cells were analyzed after uptake of AF647-9b7 against uPARAP or Alexa Fluor 647-conjugated mAb aTNP (AF647-aTNP) as a negative control. In the right panel, the merged images are displayed after incubation with AF647-9b7 (red) and AF647-aTNP (red); cells were stained with WGA Alexa Fluor 488-conjugate for surface staining (green) and Hoechst stain 33258 for visualization of the cell nuclei (blue). The internalized amount is displayed in greyscale representation (left panels). Scale bar: 20 μm.