The Blanket Effect: How Turning the World Upside Down Reveals the Nature of Graphene Oxide Cytocompatibility

Abstract

Extensive cytocompatibility testing of 2D nanocarbon materials including graphene oxide (GO) has been performed, but results remain contradictory. Literature has yet to account for settling—although sedimentation is visible to the eye and physics suggests that even individual graphenic flakes will settle. To investigate settling, a series of functional graphenic materials (FGMs) with differing oxidation levels, functionalities, and physical dimensions are synthesized. Though zeta potential indicates colloidal stability, significant gravitational settling of the FGMs is theoretically and experimentally demonstrated. By creating a setup to culture cells in traditional and inverted orientations in the same well, a “blanket effect” is demonstrated in which FGMs settle out of solution and cover cells at the bottom of the well, ultimately reducing viability. Inverted cells protected from the blanket effect are unaffected. Therefore, these results demonstrate that settling is a crucial factor that must be considered for FGM cytocompatibility experiments.

Upside-Down Preference in the Forskolin-Induced In Vitro Differentiation of 50B11 Sensory Neurons: A Morphological Investigation by Label-Free Non-Linear Microscopy

“…The most impressive feature we noted in our experiments is the peculiar appearance of the cells growing under the supports, even if it is relatively common to observe cells attached under the glass slides. Indeed, the growth of cells under the supports was previously observed using the inverted cell culture [35], transwell assay [36], and ceiling culture methods [37]. Moreover, in accordance with our observations, the inverted culture method was also previously employed to grow mouse primary hippocampal neurons showing an improved neurogenesis process and an increment in cell survival in this condition.…”

Sedimentation of colloidal nanoparticles in fluids: efficient and robust numerical evaluation of analytic solutions of the Mason-Weaver equation

“…As pointed out earlier, [2][5] [6] the series converges rapidly for large values of {τ, α}, but slowly for small values. A large number of terms for the series (e.g., > 100000) might be evaluated and added to the final result in the hope of reaching convergence, as was done by Holt et al [5]. However, it was found in the present work using standard double precision floating-point arithmetic that the factor exp (−αm 2 π 2 τ )m…”

“…Very recently, Holt et al used the analytic solution in their work on the rôle of nanoparticle sedimentation in cytotoxicity studies. [5] They reported that they needed to numerically sum a great number of terms (over 10 5 ) in the expression by M&W to obtain satisfactory results, but did not give further details. Here, the numerical evaluation of the expressions derived by Mason & Weaver is further investigated over a wide range of parameters.…”

Adenylate kinase immobilized on graphene oxide impairs progression of human lung carcinoma epithelial cells through adenosinergic pathway

“…Our previous studies, and results from other research groups, clearly demonstrated that graphene oxide (GO) fulfilled most of the conditions required for enzyme immobilization support, that is, it bound the protein with high yield, and did not decrease catalytic activity, even after storage or lyophilization 1–3 . It is of a great importance that this material is cytocompatible and exerts no toxic effects on cell viability 4,5 . In the case of enzymes as adenylate kinase (AK), expected to act in extracellular environment, GO seems to be the most convenient support preventing the cellular internalization of AK‐GO system 6 .…”

“…[1][2][3] It is of a great importance that this material is cytocompatible and exerts no toxic effects on cell viability. 4,5 In the case of enzymes as adenylate kinase (AK), expected to act in extracellular environment, GO seems to be the most convenient support preventing the cellular internalization of AK-GO system. 6 The preliminary in vitro evaluation confirmed the positive effect of AK-GO nanobiocatalyst on the cultured cells, especially when exposed to high, cytotoxic ATP concentrations.…”