We then explored the possibility of loading biotinylated cargo proteins into already formed NGs. Biotinylated green fluorescent protein (btGFP) was used as model protein due to its fluorescent properties. The preparation of btGFPloaded nanogels (btGFP-NGs) was conducted in two steps: i) DNA-protein nanogels were formed with AlexaFluor633-functionalized streptavidin in the conditions described above, and ii) btGFPs was added and bound to remaining available streptavidin sites on the nanogels (Figure ). By AFM, the size of the resulting GFPfunctionalized nanogels was measured to be 130 ± 51 nm (Figure ). The NGs were then characterized by flow cytometry (Figures and). Colocalization of the btGFP with AlexaFluor633 fluorescence in the NGs demonstrates successful formation of GFP-functionalized NGs (btGFP-NGs-633), as 72.04% of fluorescent events were observed in the upper-right quadrant of the dot plot. The NGs were then complexed with non-fluorescent cationic liposomes in order to form the functionalized lipoproplexes btGFP-LPX-633. Flow cytometry (Figure right and S8B) demonstrated that the formation of the lipoproplexes did not interfere with GFP functionality, since the signal of functional GFP protein was still co-detected with AlexaFluor633 from the nanogels after adding the liposomes, and the percentage of colocalized fluorescence events in the upper-right quadrant of the dot plot stayed quite unaffected (74.28%). B16 cells were then incubated with the btGFP-LPX-633 and observed by confocal microscopy. Figure shows the presence of fluorescent nanogels in the cells only when they have been complexed with the liposomes. Even though the fluorescence signal of the GFP was weaker, at least 76 ± 8 % (for over 400 events) of the nanogels demonstrated colocalization of AlexaFluor633 red fluorescent signal from streptavidin with the green fluorescence from the GFP (see example of composite image and orthogonal views in the Figure ), in agreement with the flow cytometry analysis. We typically observed 7.7±4.5 of internalized particles per cell (for 140 cells) in a single confocal plane and, around 50 internalized nanogels per cell have been observed on one particular z-stack of 7 confocal planes shown in the Figure . Biotinylated GFP protein can thus been successfully delivered into the cells within DNA-streptavidin nanogels complexed with cationic liposome vector.