However, Figure  5B clearly shows compartmentalization of SGS, and closer examination reveals a network

of lines (red arrows) throughout this structure, which look exactly like the folded graphene sheets previously reported by A. K. Geim et al. [25]. A magnified view of this key figure is shown in Additional file 1: Figure S7. Figure 5 SGS Internalization within Hep3B cancer cells. TEM images of internalized carbonaceous material and SGSs within Hep3B liver find more cancer cells (A to F). Figure  4D,E,F is of the same cell Figure  5D,F shows close up images of two areas of Figure  5E to reveal a stained black circular particle (Figure  5E) and a more transparent, slightly smaller, circular particle (Figure  5F). As these particles are of the same diameter as the SGS previously characterized, they are likely SGS that have internalized into the cell without folding or compartmentalization. As previously indicated, the large difference in contrast between these two SGS structures could be due to uranyl ions binding to the functionalized SGS or due to multiple stacked graphene layers. It should be noted that the cellular internalization of large SGS caused artifacts in some instances during the microtome procedure. This can be seen in Figure  6 where there is a large Selleckchem Berzosertib area of internalized SGS adjacent to a completely

transparent ‘hole’. This hole is most likely caused by the microtome blade 10058-F4 contacting the SGS and removing the structure from the cellular

cytoskeleton (thus leaving behind an SGS footprint). There is also some evidence of this in Figure  5A where the carbonaceous NP seems to have been dislodged from its initial position, leaving behind a transparent hole in the left image. This result also serves as good evidence of the cells’ ability to internalize relatively large pieces of graphite yet still remain healthy. Figure 6 TEM image of microtome cutting artifacts caused by SGS inside a SNU449 cell. It is likely that some Urease large chunks of graphite and/or SGS have been dislodged from the transparent region in the top right corner of the image. Using real-time bright-field optical microscopy, we could also track the internalization of SGSs in liver Hep3B cells as a function of time (over a 17-h period). As can be seen in Figure  7, when looking at snap shots from approximately 10 to 17 h, there were two large SGS (indicated by red and blue arrows) which became attached to the cell membrane and gradually internalized into the cell – as is evidenced by the loss of resolution and blurred nature of the SGS images. Furthermore, the cell retracted to undergo mitosis once the trapped particles are internalized. (Figure  7E,F,G,H, full movie also available in the Additional file 2: Hep3B SGS movie and Additional file 3: Hep3B control movie). Figure 7 Optical bright-field images of SGS internalization within Hep3B cancer cells across a 17-h period.