Dong Cai

Highly efficient molecular delivery into mammalian cells using carbon nanotube spearing.

Introduction of exogenous DNA into mammalian cells represents a powerful approach for manipulating signal transduction. The available techniques, however, are limited by low transduction efficiency and low cell viability after transduction. Here we report a highly efficient molecular delivery technique, named nanotube spearing, based on the penetration of nickel-embedded nanotubes into cell membranes by magnetic field driving. DNA plasmids containing the enhanced green fluorescent protein (EGFP) sequence were immobilized onto the nanotubes, and subsequently speared into targeted cells. We have achieved an unprecedented high transduction efficiency in Bal17 B-lymphoma, ex vivo B cells and primary neurons with high viability after transduction. This technique may provide a powerful tool for highly efficient gene transfer into a variety of cells, especially the hard-to-transfect cells.

A molecular-imprint nanosensor for ultrasensitive detection of proteins

Molecular imprinting is a technique for preparing polymer scaffolds that function as synthetic receptors. Imprinted polymers that can selectively bind organic compounds have proven useful in sensor development. Although creating synthetic molecular-imprinting polymers that recognize proteins remains challenging, nanodevices and nanomaterials show promise in this area. Here, we show that arrays of carbon-nanotube tips with an imprinted non-conducting polymer coating can recognize proteins with subpicogram per litre sensitivity using electrochemical impedance spectroscopy. We have developed molecular-imprinting sensors specific for human ferritin and human papillomavirus derived E7 protein. The molecular-imprinting-based nanosensor can also discriminate between Ca(2+)-induced conformational changes in calmodulin. This ultrasensitive, label-free electrochemical detection of proteins offers an alternative to biosensors based on biomolecule recognition.

Subwavelength waveguide for visible light

The authors demonstrate transmission of visible light through metallic coaxial nanostructures many wavelengths in length, with coaxial electrode spacing much less than a wavelength. Since the light frequency is well below the plasma resonance in the metal of the electrodes, the propagating mode reduces to the well-known transverse electromagnetic mode of a coaxial waveguide. They have thus achieved a faithful analog of the conventional coaxial cable for visible light.

Interaction between carbon nanotubes and mammalian cells: characterization by flow cytometry and application

We show herein that CNT-cell complexes are formed in the presence of a magnetic field. The complexes were analyzed by flow cytometry as a quantitative method for monitoring the physical interactions between CNTs and cells. We observed an increase in side scattering signals, where the amplitude was proportional to the amount of CNTs that are associated with cells. Even after the formation of CNT-cell complexes, cell viability was not significantly decreased. The association between CNTs and cells was strong enough to be used for manipulating the complexes and thereby conducting cell separation with magnetic force. In addition, the CNT-cell complexes were also utilized to facilitate electroporation. We observed a time constant from CNT-cell complexes but not from cells alone, indicating a high level of pore formation in cell membranes. Experimentally, we achieved the expression of enhanced green fluorescence protein by using a low electroporation voltage after the formation of CNT-cell complexes. These results suggest that higher transfection efficiency, lower electroporation voltage, and miniaturized setup dimension of electroporation may be accomplished through the CNT strategy outlined herein.

Carbon nanotube-mediated delivery of nucleic acids does not result in non-specific activation of B lymphocytes

The efficient delivery of genes and proteins into primary mammalian cells and tissues has represented a formidable challenge. Recent advances in the research of carbon nanotubes (CNTs) offer much promise for their use as delivery platforms into mammalian cells. Ideally, CNT-mediated applications should not result in cellular toxicity nor perturb cellular homeostasis (e.g., result in non-specific activation of primary cells). It is therefore critical to evaluate the impact of CNT exposure on the cellular metabolism, proliferation and survival of primary mammalian cells. We investigated the compatibility of a recently developed CNT-mediated delivery method, termed nanospearing, with primary ex vivo cultures of B lymphocytes. Several parameters were evaluated to assess the impact of CNTs on naive B lymphocytes, including cell survival, activation, proliferation and intracellular signal transduction. Our results indicate that nanospearing does not result in the activation of naive primary B lymphocytes nor alter survival in ex vivo cultures. Herein, B cells exposed to CNTs were capable of responding to extrinsic pro-survival signals such as interleukin-4 and signaling by the B-cell antigen receptor in a manner similar to that of B cells cultured in the absence of CNTs. Our study demonstrates the biocompatibility of the CNT-mediated nanospearing procedure with respect to primary B lymphocytes.

Ultrasensitive chemical detection using a nanocoax sensor.

We report on the design, fabrication, and performance of a nanoporous, coaxial array capacitive detector for highly sensitive chemical detection. Composed of an array of vertically aligned nanoscale coaxial electrodes constructed with porous dielectric coax annuli around carbon nanotube cores, this sensor is shown to achieve parts per billion level detection sensitivity, at room temperature, to a broad class of organic molecules. The nanoscale, 3D architecture and microscale array pitch of the sensor enable rapid access of target molecules and chip-based multiplexing capabilities, respectively.

Nanospearing – Biomolecule Delivery and Its Biocompatibility

Introduction of exogenous DNA into mammalian cells represents a powerful approach for manipulating signal transduction. However, the currently available techniques have serious limits in terms of either low transduction efficiency or low cell viability. It is found that carbon nanotubes (CNTs) can mediate molecule transportations via various mechanisms. We have reported a highly efficient molecular delivery technique, called nanotube spearing, based on the penetration of Ni-particle-embedded nanotubes into cell membranes by magnetic field driving. DNA was immobilized onto the nanotubes and subsequently speared into targeted cells. We have achieved a high transduction efficiency in Bal 17 B-lymphoma cell line, ex vivo B cells, and primary neurons with high viability. This technique may provide a powerful tool for highly efficient gene transfer in a variety of cells, especially, in the hard-to-transfect cells. However, CNTs have been associated with environmental and public health concerns which arose in the course of research on possible biomedical applications. The disturbances CNTs cause in the immune system have been met with particular interest because any ideal in vivo application of CNTs should not trigger any undesirable bodily responses. It is imperative to unravel the effects of CNTs on B cells, which represent the humoral component of acquired immunity, so that the potential risk of CNTs to public health can be thoroughly understood and advanced strategies can be employed to develop safe applications. We investigated the compatibility of the PECVD nanotubes and the nanospearing procedure in terms of cell viability, growth, and intracellular signal pathways by means of flow cytometry and biochemical analysis. No additional cell death was observed after the spearing treatment, nor had B cell activation been indicated by changes in cell size, growth, CD69 expression, and kinase phosphorylation. The post-spearing cells preserve the ability to respond to stimulation in as robust a manner as cells left untreated. Our study suggests the biocompatibility of the nanospearing procedure and PECVD nanotubes under the proposed spearing conditions with regard to the humoral component of the immune system, therefore, reducing concerns that surround in vivo applications of CNTs.