Micro-punches versus micro-slices for serial sampling of human dentine: Striking a balance between improved temporal resolution and measuring additional isotope systems.

RATIONALE: The last decade has seen a dramatic increase in the application of serial sampling of human dentine in archaeology. Rapid development in the field has provided many improvements in the methodology, in terms of both time resolution as well as the ability to integrate more isotope systems in the analysis. This study provides a comparison of two common sampling approaches, allowing researchers to select the most suitable approach for addressing specific research questions. METHODS: Two common approaches for sequential sampling of human dentine (micro-punches and micro-slices) are compared in terms of viability and efficacy. Using archaeological deciduous second molars and permanent first molars, this study demonstrates how the two approaches capture aspects of the weaning process in different ways. In addition, different aspects related to the extraction protocols, such as the thickness of the central slide and the solubilisation step, are also evaluated. RESULTS: While both approaches show similar intra-tooth isotopic patterns, the micro-punches approach is preferable for research that requires a very fine temporal resolution, while the micro-slices approach is best for research where δ34 S values are needed, or when the samples are poorly preserved. In addition, the solubilisation step has a large effect on collagen yield, and, to a lesser extent, on isotopic compositions. Therefore, it is important to ensure that only samples that have undergone the same pre-treatment protocol are directly compared. CONCLUSIONS: We present the pros and cons of the two micro-sampling approaches and offer possible mitigation strategies to address some of the most important issues related to each approach.

Collagen-carbon nanotube composite materials as scaffolds in tissue engineering.

Carbon nanotubes (CNT) are attractive for use in fiber-reinforced composite materials due to their very high aspect ratio, combined with outstanding mechanical and electrical properties. Composite materials comprising a collagen matrix with embedded CNT were prepared by mixing solubilized Type I collagen with solutions of carboxylated single-walled carbon nanotubes (SWNT) at concentrations of 0, 0.2, 0.4, 0.8, and 2.0 weight percent. Living smooth muscle cells were incorporated at the time of collagen gelation to produce cell-seeded collagen-CNT composite matrices. Constructs containing 2.0 wt % CNT exhibited delayed gel compaction, relative to lower concentrations that compacted at the same rate as pure collagen controls. Cell viability in all constructs was consistently above 85% at both Day 3 and Day 7, whereas cell number in CNT-containing constructs was lower than in control constructs at Day 3, though statistically unchanged by Day 7. Scanning electron microscopy showed physical interactions between CNT and collagen matrix. Raman spectroscopy confirmed the presence of CNT at the expected diameter (0.85-1.30 nm), but did not indicate strong molecular interactions between the collagen and CNT components. Such collagen-CNT composite matrices may have utility as scaffolds in tissue engineering, or as components of biosensors or other medical devices.

Carbon nanotubes increase the electrical conductivity of fibroblast-seeded collagen hydrogels.

Carbon nanotubes are attractive as additives in fiber-reinforced composites due to their high aspect ratio, strength and electrical conductivity. In the present study, solubilized collagen Type I was polymerized in the presence of dispersed single-walled carbon nanotubes (SWNT) and human dermal fibroblast cells (HDF) to produce collagen-SWNT composite biomaterials with HDF embedded directly in the matrix. The resulting constructs, with SWNT loadings of 0 (control), 0.8, 2.0 and 4.0 wt.% SWNT, were cultured and electrical properties were evaluated in the frequency range 5-500 kHz at days 3 and 7. All collagen-SWNT hydrogel matrices underwent HDF-mediated gel compaction over time in culture, but the presence of SWNT significantly decreased the rate and extent of gel compaction. Viability of HDF in all constructs was consistently high and cell morphology was not affected by the presence of SWNT. However, cell number at day 7 in culture decreased with increasing SWNT loading. Electrical conductivity of the constructs varied from 3 to 7 mS cm(-1), depending on SWNT loading level. Conductivity increased uniformly with increasing wt.% of SWNT (R=0.78) and showed a modest frequency dependence, suggesting that the electrical percolation threshold had not been reached in these materials. These data demonstrate that the electrical conductivity of cell-seeded collagen gels can be increased through the incorporation of carbon nanotubes. Protein-SWNT composite materials may have application as scaffolds for tissue engineering, as substrates to study electrical stimulation of cells, and as transducers or leads for biosensors.

Directional conductivity in SWNT-collagen-fibrin composite biomaterials through strain-induced matrix alignment.

Composite biomaterials incorporating fibroblast cells, collagen Type I, fibrin, and 2 wt % carboxylated SWNT were created, and their properties were compared with similar control constructs without SWNT. Alignment of the matrix was stimulated by application of 8% cyclic strain for three 12-h periods over three days. All constructs underwent cell-mediated gel compaction to 15-20% of their initial volume, which was not affected by SWNT loading. Mechanical strain increased the rate of compaction, and strained constructs were significantly more compacted than unstrained controls by day 3. Cell viability and morphology were similar in both control and SWNT-loaded constructs, but unstrained samples exhibited a more stellate appearance with more numerous cellular projections. Application of mechanical strain caused clear alignment of both the cells and matrix in the direction of the applied strain. Bioimpedance measurements showed that SWNT loading increased the electrical conductivity of composite constructs, and that mechanically-induced alignment of the matrix/SWNT caused a further increase in conductivity. These results demonstrate that SWNT can be used to augment the electrical properties of 3D protein hydrogels, and that anisotropy in the matrix further enhances these properties. Such electrically conductive biopolymers may have a variety of applications in tissue engineering and biosensor development.