Reactive biomolecular divergence in genetically altered yeast cells and isolated mitochondria as measured by biocavity laser spectroscopy: rapid diagnostic method for studying cellular responses to stress and disease.

We report an analysis of four strains of baker's yeast (Saccharomyces cerevisiae) using biocavity laser spectroscopy. The four strains are grouped in two pairs (wild type and altered), in which one strain differs genetically at a single locus, affecting mitochondrial function. In one pair, the wild-type rho+ and a rho0 strain differ by complete removal of mitochondrial DNA (mtDNA). In the second pair, the wild-type rho+ and a rho- strain differ by knock-out of the nuclear gene encoding Cox4, an essential subunit of cytochrome c oxidase. The biocavity laser is used to measure the biophysical optic parameter Deltalambda, a laser wavelength shift relating to the optical density of cell or mitochondria that uniquely reflects its size and biomolecular composition. As such, Deltalambda is a powerful parameter that rapidly interrogates the biomolecular state of single cells and mitochondria. Wild-type cells and mitochondria produce Gaussian-like distributions with a single peak. In contrast, mutant cells and mitochondria produce leptokurtotic distributions that are asymmetric and highly skewed to the right. These distribution changes could be self-consistently modeled with a single, log-normal distribution undergoing a thousand-fold increase in variance of biomolecular composition. These features reflect a new state of stressed or diseased cells that we call a reactive biomolecular divergence (RBD) that reflects the vital interdependence of mitochondria and the nucleus.

Mitochondrial correlation microscopy and nanolaser spectroscopy - new tools for biophotonic detection of cancer in single cells.

Currently, pathologists rely on labor-intensive microscopic examination of tumor cells using century-old staining methods that can give false readings. Emerging BioMicroNano-technologies have the potential to provide accurate, realtime, high-throughput screening of tumor cells without the need for time-consuming sample preparation. These rapid, nano-optical techniques may play an important role in advancing early detection, diagnosis, and treatment of disease. In this report, we show that laser scanning confocal microscopy can be used to identify a previously unknown property of certain cancer cells that distinguishes them, with single-cell resolution, from closely related normal cells. This property is the correlation of light scattering and the spatial organization of mitochondria. In normal liver cells, mitochondria are highly organized within the cytoplasm and highly scattering, yielding a highly correlated signal. In cancer cells, mitochondria are more chaotically organized and poorly scattering. These differences correlate with important bioenergetic disturbances that are hallmarks of many types of cancer. In addition, we review recent work that exploits the new technology of nanolaser spectroscopy using the biocavity laser to characterize the unique spectral signatures of normal and transformed cells. These optical methods represent powerful new tools that hold promise for detecting cancer at an early stage and may help to limit delays in diagnosis and treatment.

Brief overview of BioMicroNano technologies.

This paper provides a brief overview of the fields of biological micro-electromechanical systems (bioMEMs) and associated nanobiotechnologies, collectively denoted as BioMicroNano. Although they are developing at a very rapid pace and still redefining themselves, several stabilized areas of research and development can be identified. Six major areas are delineated, and specific examples are discussed and illustrated. Various applications of the technologies are noted, and potential market sizes are compared.

Ultrafast nanolaser flow device for detecting cancer in single cells.

Currently, pathologists rely on labor-intensive microscopic examination of tumor cells using staining techniques originally devised in the 1880s that depend heavily on specimen preparation and that can give false readings. Emerging BioMicroNanotechnologies (Gourley, 2005) have the potential to provide accurate, realtime, high throughput screening of tumor cells without invasive chemical reagents. These techniques are critical to advancing early detection, diagnosis, and treatment of disease. Using a new technique to rapidly assess the properties of cells flown through a nanolaser semiconductor device, we discovered a method to rapidly assess the respiratory health of a single mammalian cell. The key discovery was the elucidation of biophotonic differences in normal and transformed (cancer) mouse liver cells by using intracellular mitochondria as biomarkers for disease. This technique holds promise for detecting cancer at a very early stage and could nearly eliminate delays in diagnosis and treatment.

Poly(dimethylsiloxane) thin films as biocompatible coatings for microfluidic devices: cell culture and flow studies with glial cells.

Oxygen plasma treatment of poly(dimethylsiloxane) (PDMS) thin films produced a hydrophilic surface that was biocompatible and resistant to biofouling in microfluidic studies. Thin film coatings of PDMS were previously developed to provide protection for semiconductor-based microoptical devices from rapid degradation by biofluids. However, the hydrophobic surface of native PDMS induced rapid clogging of microfluidic channels with glial cells. To evaluate the various issues of surface hydrophobicity and chemistry on material biocompatibility, we tested both native and oxidized PDMS (ox-PDMS) coatings as well as bare silicon and hydrophobic alkane and hydrophilic oligoethylene glycol silane monolayer coated under both cell culture and microfluidic studies. For the culture studies, the observed trend was that the hydrophilic surfaces supported cell adhesion and growth, whereas the hydrophobic ones were inhibitive. However, for the fluidic studies, a glass-silicon microfluidic device coated with the hydrophilic ox-PDMS had an unperturbed flow rate over 14 min of operation, whereas the uncoated device suffered a loss in rate of 12%, and the native PDMS coating showed a loss of nearly 40%. Possible protein modification of the surfaces from the culture medium also were examined with adsorbed films of albumin, collagen, and fibrinogen to evaluate their effect on cell adhesion.

Surface passivation of a microfluidic device to glial cell adhesion: a comparison of hydrophobic and hydrophilic SAM coatings.

Cell adhesion in a microfluidic structure can lead to catastrophic flow problems due to the comparable size of the cell with the microfabricated device. Such issues are important in the growing research area involving the merging of biological materials and MEMS devices. We have examined the surface compatibility of uncoated and coated microfabricated glass and semiconductor surfaces under static solution (cell culture) and flow experiments (microfluidic device) using glial (astrocyte and glioblastoma) cells. Bare semiconductor and glass surfaces were most attractive to cell adhesion, promoting biofouling under both static and flow conditions. Passivation of the surfaces was performed with silane coupling agents octadecyltrimethoxysilane (OTMS) or N-(triethoxysilylpropyl)-O-polyethylene oxide urethane (TESP) on SiO2 surfaces via self-assembled monolayer (SAM) deposition. The hydrophilic TESP coating was effective at inhibiting biofouling of the microfluidic structure, allowing greater than several minutes of fluid flow. The hydrophobic OTMS coating, on the other hand, promoted cell adhesion leading to restricted flow within a few minutes. Interestingly, under cell culture conditions the TESP surface exhibited biocompatible properties for glial cell adhesion and proliferation, in contrast to the OTMS surface which resisted cell growth. These studies suggest that cell adhesion is dependent upon the time domain of the cell-surface interaction.