RGD-core-IFN-α2a fusion proteins bind breast cancer cells MDA-MB231 in vitro. (A) Recombinant bacmid constructs, showing the strategy for insertion of the gene cassettes into the polyhedrin locus of the AcMNPV bacmid. RGD-HCV core was fused with IFN-α2a. Both cassettes depicted were inserted into the attb site (indicated by the right and left insertion sites, Tn7R and Tn7L) in the polyhedrin locus by Tn-based transposition and generated the recombinant Bacmid: AcH1, AcH2, AcH3, and AcH4. (B) Identification of pH1 and pH2. M: 1Kb Plus DNA ladder; pH1 and pH2 samples were digested by BamHI and EcoRI. (C) Identification of pH3 and pH4. M: O’Gene Ruler 1Kb DNA ladder; pH3 and pH4 samples were digested by BamHI and EcoRI. (D) Purification of RGD-core-IFN-α2a fusion protein. M: protein marker; 1: His-H1; 2: His-H2; 3: His-H3; 4: His-H4. The recombinant bacmids AcH1, AcH2, AcH3, and AcH4 were introduced by transfection into Sf9 cells to produce the recombinant proteins His-H1, His-H2, His-H3, and His-H4. The fusion proteins were purified from the supernatants of cell lysates using Ni-NTA affinity resin. (E, G) Electron micrograph images and Western blotting result of VLP H1. Purified VLPs were attached onto a carbon-coated grid for 5 min at room temperature. The grid was rinsed with distilled water and stained with 1% phosphotungstic acid for 3 min before air drying on filter paper. The specimens were viewed using a Tecnai G2 transmission electron microscopy at 75 keV. For Western blot, 10 μg purified VLPs were separated by SDS-PAGE electrophoresis and subjected to Western blot assay. (F, H) Electron micrograph images and Western blotting result of VLP H2. (I) RGD-core-IFN-α2a fusion protein bind with breast cancer cells MDA-MB231. Then, 0.2, 0.5, 2, 5, and 10 μM fusion proteins His-H1, His-H2, His-H3, and His-H4 were co-incubated with MDA-MB231 at 37° under 5% CO2. After 2 h, the cells were washed three times with PBS, and green fluorescence was observed under the fluorescence microscope. Scale bar = 100 μm.
Li et al. Nanoscale Research Letters 2013 8:401 doi:10.1186/1556-276X-8-401