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Fig. 3 | Molecular Cancer

Fig. 3

From: Modulation of extracellular matrix in cancer is associated with enhanced tumor cell targeting by bacteriophage vectors

Fig. 3

Cellular diffusion and internalization of RGD4C.AAVP is boosted by ECM clearance. a RGD4C.AAVP diffusion assay was carried out using fluorescently-labelled RGD4C.AAVP and followed through an ECM gel matrix. Two ECM concentrations were tested (2.5 and 5.0 mg/ml) and RGD4C.AAVP was tracked using fluorescent microscopy. b Measurement of RGD4C.AAVP diffusion, per fields of view (FOV), at various time points for a total time of 6 h. c Flow cytometric analysis of AAVP uptake was performed in 9L cells after ECM depletion. Cells were treated either with a combination mix of collagenase (0.2 mg/ml) and hyaluronidase (0.4 mg/ml) or without any enzyme (control) for 1 h before incubation with either non-targeted NT.AAVP (white bars) or RGD4C.AAVP (black bars). Fixation was conducted 4 h post-transduction and immunofluorescence was performed using rabbit anti-phage primary and goat anti-rabbit AlexaFluor-647 conjugated antibodies. Gating threshold was set at 10,000 events of total cell population. Each condition was measured in triplicate and statistics were obtained by unpaired student t-test (** = p < 0.01). Data are presented as mean ± s.e.m. d The graph shows shifts in mean fluorescence intensity and RGD4C.AAVP positive cell counts between no-enzyme control (yellow line) and enzyme combination treatment (blue line). Experiments were repeated twice and individual conditions were repeated in triplicates

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