Imaging carbon nanotubes inside a living organism

(Nanowerk Spotlight) Nanotechnology's poster child, the carbon nanotube (CNT), has been explored for use in many technical applications. Increasingly, researchers are also looking at the unique biological properties of CNTs for potential biomedical uses. For instance, the interaction between DNA and CNTs have been explored and DNA-functionalized nanotubes hold significant promise as nucleic acid sensors. Nanotubes have also been considered for use as scaffolds for cells in tissue engineering. No matter what their intended function, any material used in medicine must exhibit - among other compatibility factors - biocompatibility, non-toxicity and non-carcinogenicity. And here the jury is still out as far as CNTs are concerned. One limiting factor of toxicological studies so far has been the use of animal tissue rather than living specimen. Researchers have now succeeded in detecting single-walled CNTs (SWCNTs) inside living animals - with surprisingly benign results - paving the way for future research on the effects and fate of nanotubes inside living organisms.
"We believe that our work is the first observation of carbon nanotubes in a living animal and the first detection of individual nanotubes in biological tissues" Dr. R. Bruce Weisman explains to Nanowerk. "Although much more work must be done to investigate the effects of SWCNT exposure on higher animals (mammals), these results show an absence of negative effects in one system and suggest that it is worthwhile to continue studying SWCNTs for biomedical applications, including diagnostic imaging based on their unique near-IR fluorescence signatures."
Weisman, a professor in the Department of Chemistry at Rice University in Houston, Texas, and his group investigate the spectroscopy and photophysics of fullerenes and carbon nanotubes. Following the discovery in Weisman's lab of near-infrared nanotube fluorescence, the group has measured and unraveled the absorption and emission spectra of more than 30 semiconducting nanotube species. In his most recent work, Weisman, together with co-author Kathleen Beckingham, professor of biochemistry and cell biology, attempted the first-ever detection of nanotubes inside a living animal. Their findings have been published in a recent paper in Nano Letters ("Single-Walled Carbon Nanotubes in the Intact Organism: Near-IR Imaging and Biocompatibility Studies in Drosophila").
Single-walled carbon nanotubes in fruit flies
SWCNTs in the gut and blood system. (a,b) NIR emission (color-coded for intensity) from SWCNTs in the gut of a living larva viewed through the larval cuticle. In (a), the black branching structures are part of the trachea system that brings air in from opening in the cuticle surface (upper right). The larva was fed yeast paste containing 9 ppm SWCNTs. (b) Boluses of food containing SWCNTs in a loop of the gut of a living larva. This 0.5 s exposure was from a sequence that clearly showed peristaltic activity. Scale bars are 50 µm (a) and 100 µm (b). Note that the experimental conditions and intensity scales differ for (a) and (b). (c-e) SWCNT NIR emission showing accumulation in the dorsal vessel. Green fluorescence from GFP expressed exclusively in the dorsal vessel is shown in panel (c), and NIR fluorescence from nanotubes is shown in panel (d) (false colored in red). (e) Overlay of these two images on the corresponding bright field image, demonstrating that the SWCNTs lie within the lumen of the vessel. Scale bars are 25 µm. For (b-e), larvae were fed yeast paste containing 16 ppm SWCNTs. (Reprinted with permission from American Chemical Society)
In the study, fruit fly (Drosophila melanogaster) larvae were raised on a yeast paste that contained carbon nanotubes. The flies were fed this food from the time they hatched throughout their initial feeding phase of 4-5 days. Fruit flies are ravenous eaters during this period and gain weight continuously until they are about 200 times heavier than hatchlings. Then they become pupae. As pupae, they do not eat or grow. They mature inside pupal cases and emerge as adult flies.
Weisman lists the main results of their study:
  • "Dispersed SWCNTs in the food supply of fruit fly larvae have no adverse effects on their survival to adult stage flies or on their developmental weight gain;
  • SWCNTs can be noninvasively detected in living animals (fly larvae) by imaging the nanotubes? characteristic near-infrared fluorescence emission;
  • Under the microscope, we show that this approach even lets us image individual SWCNTs in dissected tissue specimens; and
  • Only a very tiny fraction of the ingested SWCNTs became incorporated into tissues of the fly larvae; almost all seem to have passed harmlessly through the digestive system."
  • This study addresses two issues relevant to envisioned biomedical applications of SWCNTs. "First" says Weisman, "it demonstrates that near-infrared fluorescence is a highly effective probe for disaggregated SWCNTs in biological tissues and organisms. It can detect, image, and structurally identify individual nanotubes in tissue specimens and can nondestructively image accumulations of nanotubes inside living organisms.
    "Second, our study provides new results on the effect of SWCNTs on intact organisms, relevant to possible medical uses and also to environmental contamination concerns."
    The Rice researchers found no short-term toxicity or impaired growth or viability of Drosophila larvae that had been fed dispersed SWCNTs, no matter what concentration they used in their experiments. In addition, there was no obvious impairment of fertility in matured SWCNT-fed individuals.
    Weisman points out that, although they cannot exclude subtle effects on some aspects of the Drosophila life cycle, their findings suggest that SWCNTs ingested by these insects will have negligible physiological impact.
    "Further, we estimate that only a very small fraction, ∼10-8, of the ingested nanotubes became incorporated into organs of the larvae" he says. "If these findings are valid for other insect species, then uptake by insect ingestion may prove to be an ineffective route for nanotube entry into the food chain from the environment."
    Altogether, if these findings on apparent biocompatibility of SWCNTs can be validated on a broad basis, then it seems that their sensitive in vivo detection shown in this research supports their promise for the development of novel biomedical applications.
    Michael Berger By – Michael is author of three books by the Royal Society of Chemistry:
    Nano-Society: Pushing the Boundaries of Technology,
    Nanotechnology: The Future is Tiny and
    Nanoengineering: The Skills and Tools Making Technology Invisible
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