Anti-Human HER2/neu Recombinant Antibody scFv Fragment (MIL5) (CAT#: TAB-044CT-S(P))

Figure 1 Expression and identification of fused proteins.

Western blot analysis of purified proteins. 1: MIL5scFv; lane 2: MIL5scFv-Arg9. The predicted molecular weight of MIL5scFv-Arg9 was about 49 kDa, which was almost the same with that of MIL5scFv.

Hu, Y., Qiao, C., Lv, M., Feng, J., Yu, M., Shen, B., ... & Li, Y. (2012). Arg9 facilitates the translocation and downstream signal inhibition of an anti-HER2 single chain antibody. BMC research notes, 5(1), 336.

Figure 2 Flow cytometry analysis and confocal microscopy observation of MIL5scFv-Arg9 to penetrate NIH3T3 cells (HER2 negative).

Cells treated with MIL5scFv-FITC. The intensity of fluorescence signal was evaluated with MFI value marked on the right top of each panel. According to the fluorescence intensity, Arg9 could help MIL5scFv penetrate NIH3T3 cells. Data are representative of two independent experiments.

Hu, Y., Qiao, C., Lv, M., Feng, J., Yu, M., Shen, B., ... & Li, Y. (2012). Arg9 facilitates the translocation and downstream signal inhibition of an anti-HER2 single chain antibody. BMC research notes, 5(1), 336.

Figure 3 Flow cytometry analysis and confocal microscopy observation of MIL5scFv-Arg9 to penetrate NIH3T3 cells (HER2 negative).

Confocal microscopy observation of MIL5scFv-Arg9 to penetrate NIH3T3 cells. Cells were incubated with PBS (Control) or herceptin-FITC as negative control, MIL5scFv-Arg9-FITC (1:10) or MIL5scFv-FITC (1:10) at 37°C and collected consecutively in 0.5 h, 1 h, 2 h and 5 h. The distribution of fluorescence signal in the cells was observed under a laser scanning confocal microscopy. Fluorescent signal could be detected after 0.5 hour in MIL5scFv-Arg9 treated samples, while in MIL5scFv treated samples, weak signal could be seen after 5 hours. Each panel was merged with two photos: FITC conjugated protein (green) and nuclei (blue, cells were stained with Hoechst 33258), which were not shown respectively here.

Hu, Y., Qiao, C., Lv, M., Feng, J., Yu, M., Shen, B., ... & Li, Y. (2012). Arg9 facilitates the translocation and downstream signal inhibition of an anti-HER2 single chain antibody. BMC research notes, 5(1), 336.

Figure 4 Enhanced translocation and stronger activity of MIL5scFv-Arg9 to downregulate phospho-Akt in SKOV3 cells (HER2 positive).

Densities of phosphor-Akt were normalized based on densities of Akt shown in Figure 4B, suggesting satisfactory enhanced effect of Arg9 in inhibiting the expression level of phospho-Akt in SKOV3 cells. The data shown are calculate from two independent experiments.

Hu, Y., Qiao, C., Lv, M., Feng, J., Yu, M., Shen, B., ... & Li, Y. (2012). Arg9 facilitates the translocation and downstream signal inhibition of an anti-HER2 single chain antibody. BMC research notes, 5(1), 336.