Biomedical subjects

Jonas O Tegenfeldt

Publications and source records attributed to Jonas O Tegenfeldt.

9 recordsLinked to original sources

PubMed2006

Use of PLL-g-PEG in micro-fluidic devices for localizing selective and specific protein binding.

By utilizing flow-controlled PLL-g-PEG and PLL-g-PEGbiotin modification of predefined regions of a poly(dimethylsiloxane) (PDMS) micro-fluidic device, with an intentionally chosen large (approximately 1 cm2) internal surface area, we report rapid (10 min), highly localized (6 x 10(-6) cm2), and specific surface-based protein capture from a sample volume (100 microL) containing a low amount of protein (160 attomol in pure buffer and 400 attomol in serum). The design criteria for this surface modification were achieved using QCM-D (quartz crystal microbalance with energy dissipation monitoring) of serum protein adsorption onto PLL-g-PEG-modified oxidized PDMS. Equally good, or almost as good, results were obtained for oxidized SU-8, Topas, and poly(methyl metacrylate) (PMMA), demonstrating the generic potential of PLL-g-PEG for surface modification in various micro-fluidic applications.

Adsorption↗

PubMed2006

Improving the instrumental resolution of sensors based on localized surface plasmon resonance.

The colorimetric variations induced upon changes in interfacial refractive index of nanoscale noble metal structures exhibiting localized surface plasmon resonance (LSPR) provides a convenient means of label-free, affinity-based detection of biomolecular recognition reactions. However, despite being similar in nature to conventional SPR, LSPR has so far suffered from significantly lower data quality in terms of its signal-to-noise ratio (S/N) in typical biomolecular recognition analysis. In this work, generic data analysis algorithms and a simple experimental setup that provide a S/N upon protein binding that is comparable to that of state-of-the art SPR systems are presented. Specifically, it is demonstrated how temporal variations (rate approximately 0.5 Hz) in parameters proportional to the resonance peak position can be recorded simultaneously, yielding a peak position precision of <5 x 10(-4) nm and an extinction noise level of <5 x 10(-6) absorbance units (Abs). This, in turn, is shown to provide a S/N of approximately 2000 (equivalent to a detection limit of <0.1 ng/cm(2)) for typical protein binding reactions. Furthermore, the importance of utilizing changes in both peak position and magnitude is highlighted by comparing different LSPR active noble metal architectures that respond differently to bulk and interfacial refractive index changes.

Nanostructures↗

PubMed2005

Single-molecule studies of repressor-DNA interactions show long-range interactions.

We have performed single-molecule studies of GFP-LacI repressor proteins bound to bacteriophage lambda DNA containing a 256 tandem lac operator insertion confined in nanochannels. An integrated photon molecular counting method was developed to determine the number of proteins bound to DNA. By using this method, we determined the saturated mean occupancy of the 256 tandem lac operators to be 13, which constitutes only 2.5% of the available sites. This low occupancy level suggests that the repressors influence each other even when they are widely separated, at distances on the order of 200 nm, or several DNA persistence lengths.

Bacteriophage lambda↗

PubMed2004

From the Cover: The dynamics of genomic-length DNA molecules in 100-nm channels.

We show that genomic-length DNA molecules imaged in nanochannels have an extension along the channel that scales linearly with the contour length of the polymer, in agreement with the scaling arguments developed by de Gennes for self-avoiding confined polymers. This fundamental relationship allows us to measure directly the contour length of single DNA molecules confined in the channels, and the statistical analysis of the dynamics of the polymer in the nanochannel allows us to compute the SD of the mean of the extension. This statistical analysis allows us to measure the extension of lambda DNA multimers with a 130-nm SD in 1 min.

DNA↗

PubMed2004

Micro- and nanofluidics for DNA analysis.

Miniaturization to the micrometer and nanometer scale opens up the possibility to probe biology on a length scale where fundamental biological processes take place, such as the epigenetic and genetic control of single cells. To study single cells the necessary devices need to be integrated on a single chip; and, to access the relevant length scales, the devices need to be designed with feature sizes of a few nanometers up to several micrometers. We will give a few examples from the literature and from our own research in the field of miniaturized chip-based devices for DNA analysis, including dielectrophoresis for purification of DNA, artificial gel structures for rapid DNA separation, and nanofluidic channels for direct visualization of single DNA molecules.

Cell Separation↗

PubMed2002

Bacterial chromosome extraction and isolation.

We have used diffusive mixing and dielectrophoretic trapping to lyse Escherichia coli cells in a microfabricated environment and trap the E. coli chromosome. We characterize the conditions needed for efficient lysis of the cells, and conditions needed for the dielectrophoretic trapping of the chromatin without the trapping of cytoplasmic proteins.

Chromosomes, Bacterial↗

PubMed2002

Role of molecular size in ratchet fractionation.

We show the importance of finite particle size in microfluidic asymmetric continuous-flow diffusion arrays, specifically the critical nature of the particle size with respect to the barrier gaps. We show that particles much smaller than the barrier gap follow individual field lines through narrow gaps and are poorly fractionated. In contrast, particles comparable to the gap size lose memory of their incoming field line and can be fractionated with high resolution. We demonstrate this effect using a new technological approach to create very straight and narrow injection bands in such arrays, and completely resolve bands of DNA of lengths 48,500 and 16,7000 base pairs.

Bacteriophage T4↗

PubMed2002

A DNA prism for high-speed continuous fractionation of large DNA molecules.

The analysis and fractionation of large DNA molecules plays a key role in many genome projects. The standard method, pulsed-field gel electrophoresis (PFGE), is slow, with running times ranging from 10 hours to more than 200 hours. In this report, we describe a thumbnail-sized device that sorts large DNA fragments (61-209 kilobases (kb)) in 15 seconds, with a resolution of approximately 13%. An array of micron-scale posts serves as the sieving matrix, and integrated microfluidic channels spatially shape the electric fields over the matrix. Asymmetric pulsed fields are applied for continuous-flow operation, which sorts DNA molecules in different directions according to their molecular masses, much as a prism deflects light of different wavelengths at different angles. We demonstrate the robustness of the device by using it to separate large DNA inserts prepared from bacterial artificial chromosomes, a widely used DNA source for most genomics projects.

Chemical Fractionation↗

PubMed2002

Electrodeless dielectrophoresis of single- and double-stranded DNA.

Dielectrophoretic trapping of molecules is typically carried out using metal electrodes to provide high field gradients. In this paper we demonstrate dielectrophoretic trapping using insulating constrictions at far lower frequencies than are feasible with metallic trapping structures because of water electrolysis. We demonstrate that electrodeless dielectrophoresis (EDEP) can be used for concentration and patterning of both single-strand and double-strand DNA. A possible mechanism for DNA polarization in ionic solution is discussed based on the frequency, viscosity, and field dependence of the observed trapping force.

Biophysical Phenomena↗