A new approach to the toxicity testing of carbon-based nanomaterials—The clonogenic assay

Raman spectroscopy – a potential platform for the rapid measurement of carbon nanotube-induced cytotoxicity

“…The ratios of bands previously identified as cytotoxic markers clearly show a dose dependent response. This dose dependence 50 correlates well with that previously observed for colony size in clonogenic assays on the same samples 12 . The dose dependent response of the colony size endpoint of the clonogenic study is plotted in (Figure 10).…”

“…for consistency with previous studies [9][10][11][12] . The tip was operated at a medium level of output for a total time of 30s carried out in 10s sequential steps to minimise sample heating.…”

“…A monotonic decrease in colony size with increasing dose up to ~30mg/l is observed with a saturated 55 response at higher doses. This toxic response has been attributed to a reduced proliferative capacity as a result of medium depletion caused by adsorption of components of the cell growth medium to the SWCNTs 12 . Figure 11 The results clearly indicate that dose dependent spectral markers 15 can be identified in the Raman spectra of cellular samples exposed to SWCNTs.…”

“…Clonogenic studies have been demonstrated to be a potentially more appropriate toxicological assay and it has been demonstrated that colony-size rather than colony number is 35 a more powerful measurement of cellular toxicity post exposure to SWCNTs 12 . The implications are that the toxic response is possibly one of reduced proliferative capacity of the cells due to medium depletion rather than a reduced viability due to direct interaction with the cells 12 . Such an indirect toxicological 40 response has more recently been demonstrated 13 .…”

“…In order to allow a realistic relationship between the experimental data presented 80 here and that in vivo, the SWCNT samples were minimally processed to mimic inhalation of airborne SWCNT dust particles. SWCNT dispersion was carried out in an identical fashion to that employed in previous cytotoxicity studies [9][10][11] and cell growth and exposure conditions were identical to those employed in previous 85 clonogenic studies 12 . Dose dependent responses are examined in terms of peak ratios utilised in previous toxicity studies as well as principal component analysis.…”