Translate this page to:
Swedish
In JoVE (1)
Other Publications (17)
- Magnetic Resonance Imaging
- Proceedings of the National Academy of Sciences of the United States of America
- Nano Letters
- Optics Letters
- Journal of the Optical Society of America. A, Optics, Image Science, and Vision
- Journal of the Optical Society of America. A, Optics, Image Science, and Vision
- Proceedings of the National Academy of Sciences of the United States of America
- Optics Express
- Applied Optics
- IEEE Journal of Selected Topics in Quantum Electronics : a Publication of the IEEE Lasers and Electro-optics Society
- Analytical Chemistry
- Journal of the Optical Society of America. A, Optics, Image Science, and Vision
- Biosensors & Bioelectronics
- Nano Letters
- Biosensors & Bioelectronics
- ACS Nano
- Nanoscale
Automatic Translation
This translation into Swedish was automatically generated.
English Version | Other Languages
Articles by Bennett B. Goldberg in JoVE
JoVE Bioengineering
Biomolekylära Detection anställa interferometriska Sensor Reflektans Imaging (IRIS)
1Department of Electrical and Computer Engineering, Boston University, 2Department of Biomedical Engineering, Boston University, 3Center for Advanced Genomics Technology, Boston University, 4Department of Medicine, Section of Infectious Diseases, Boston University School of Medicine, 5Department of Microbiology, Boston University School of Medicine, 6CNR (National Research Council), Istituto di Chimica del Riconoscimento Molecolare
Kvantitativa, hög genomströmning, i realtid, och etikett-fri biomolekylär detektion (DNA, protein, etc.) på SiO
Other articles by Bennett B. Goldberg on PubMed
MRI of the Lung Gas-space at Very Low-field Using Hyperpolarized Noble Gases
In hyperpolarized (HP) noble-gas magnetic resonance imaging, large nuclear spin polarizations, about 100,000 times that ordinarily obtainable at thermal equilibrium, are created in 3He and 129Xe. The enhanced signal that results can be employed in high-resolution MRI studies of void spaces such as in the lungs. In HP gas MRI the signal-to-noise ratio (SNR) depends only weakly on the static magnetic field (B(0)), making very low-field (VLF) MRI possible; indeed, it is possible to contemplate portable MRI using light-weight solenoids or permanent magnets. This article reports the first in vivo VLF MR images of the lungs in humans and in rats, obtained at a field of only 15 millitesla (150 Gauss).
DNA Conformation on Surfaces Measured by Fluorescence Self-interference
The conformation of DNA molecules tethered to the surface of a microarray may significantly affect the efficiency of hybridization. Although a number of methods have been applied to determine the structure of the DNA layer, they are not very sensitive to variations in the shape of DNA molecules. Here we describe the application of an interferometric technique called spectral self-interference fluorescence microscopy to the precise measurement of the average location of a fluorescent label in a DNA layer relative to the surface and thus determine specific information on the conformation of the surface-bound DNA molecules. Using spectral self-interference fluorescence microscopy, we have estimated the shape of coiled single-stranded DNA, the average tilt of double-stranded DNA of different lengths, and the amount of hybridization. The data provide important proofs of concept for the capabilities of novel optical surface analytical methods of the molecular disposition of DNA on surfaces. The determination of DNA conformations on surfaces and hybridization behavior provide information required to move DNA interfacial applications forward and thus impact emerging clinical and biotechnological fields.
Screening of Excitons in Single, Suspended Carbon Nanotubes
Resonant Raman spectroscopy of single carbon nanotubes suspended across trenches displays red-shifts of up to 30 meV of the electronic transition energies as a function of the surrounding dielectric environment. We develop a simple scaling relationship between the exciton binding energy and the external dielectric function and thus quantify the effect of screening. Our results imply that the underlying particle interaction energies change by hundreds of meV.
Diffraction of Evanescent Waves and Nanomechanical Displacement Detection
Sensitive displacement detection has emerged as a significant technological challenge in mechanical resonators with nanometer-scale dimensions. A novel nanomechanical displacement detection scheme based upon the scattering of focused evanescent fields is proposed. The sensitivity of the proposed approach is studied using diffraction theory of evanescent waves. Diffraction theory results are compared with numerical simulations.
Spectral Self-interference Microscopy for Low-signal Nanoscale Axial Imaging
A theoretical and numerical analysis of spectral self-interference microscopy (SSM) is presented with the goal of expanding the realm of SSM applications. In particular, this work is intended to enable SSM imaging in low-signal applications such as single-molecule studies. A comprehensive electromagnetic model for SSM is presented, allowing arbitrary forms of the excitation field, detection optics, and tensor sample response. An evanescently excited SSM system, analogous to total internal reflection microscopy, is proposed and investigated through Monte Carlo simulations. Nanometer-scale axial localization for single-emitter objects is demonstrated, even in low-signal environments. The capabilities of SSM in imaging more general objects are also considered--specifically, imaging arbitrary fluorophore distributions and two-emitter objects. A data-processing method is presented that makes SSM robust to noise and uncertainties in the detected spectral envelope.
4Pi Spectral Self-interference Microscopy
Spectral self-interference microscopy (SSM) relies on the balanced collection of light traveling two different paths from the sample to the detector, one direct and the other indirect from a reflecting substrate. The resulting spectral interference effects allow nanometer-scale axial localization of isolated emitters. To produce spectral fringes the difference between the two optical paths must be significant. Consequently, to ensure that both contributions are in focus, a low-numerical-aperture objective lens must be used, giving poor lateral resolution. Here this limitation is overcome using a 4Pi apparatus to produce the requisite two paths to the detector. The resulting instrument generalizes both SSM and 4Pi microscopy and allows a quantification of SSM resolution (rather than localization precision). Specifically, SSM is shown to be subject to the same resolution constraints as 4Pi microscopy.
Label-free and Dynamic Detection of Biomolecular Interactions for High-throughput Microarray Applications
Direct monitoring of primary molecular-binding interactions without the need for secondary reactants would markedly simplify and expand applications of high-throughput label-free detection methods. A simple interferometric technique is presented that monitors the optical phase difference resulting from accumulated biomolecular mass. As an example, 50 spots for each of four proteins consisting of BSA, human serum albumin, rabbit IgG, and protein G were dynamically monitored as they captured corresponding antibodies. Dynamic measurements were made at 26 pg/mm(2) SD per spot and with a detectable concentration of 19 ng/ml. The presented method is particularly relevant for protein microarray analysis because it is label-free, simple, sensitive, and easily scales to high-throughput.
Widefield Subsurface Microscopy of Integrated Circuits
We apply the numerical aperture increasing lens technique to widefield subsurface imaging of silicon integrated circuits. We demonstrate lateral and longitudinal resolutions well beyond the limits of conventional backside imaging. With a simple infrared widefield microscope (lambda(0) = 1.2 microm), we demonstrate a lateral spatial resolution of 0.26 microm (0.22 lambda(0)) and a longitudinal resolution of 1.24 microm (1.03 lambda(0)) for backside imaging through the silicon substrate of an integrated circuit. We present a spatial resolution comparison between widefield and confocal microscopy, which are essential in integrated circuit analysis for emission and excitation microscopy, respectively.
Hyperspectral Fourier Transform Spectrometer for Reflection Spectroscopy and Spectral Self-interference Fluorescence Microscopy
A hyperspectral Fourier transform spectrometer has been developed for studying biological material bound to optically reflecting surfaces. This instrument has two modes of operation: a white-light reflection mode and a spectral self-interference fluorescence mode. With the combined capability, information about the conformation of an ensemble of biomolecules may be determined. To the best of our knowledge, ours is the first report of this hybrid white-light reflection, spectral self-interference fluorescence measurement with any type of hyperspectral imager. The measurement technique is presented along with a full description of the system, including theoretical performance projections. Proof-of-principle measurements of artificial samples are shown, and the results are discussed.
Resonant Cavity Imaging: A Means Toward High-Throughput Label-Free Protein Detection
The resonant cavity imaging biosensor (RCIB) is an optical technique for detecting molecular binding interactions label free at many locations in parallel that employs an optical resonant cavity for high sensitivity. Near-infrared light centered at 1512.5 nm couples resonantly through a Fabry-Perot cavity constructed from dielectric reflectors (Si/SiO(2)), one of which serves as the binding surface. As the wavelength is swept using a tunable laser, a near-infrared digital camera monitors cavity transmittance at each pixel. A wavelength shift in the local resonant response of the optical cavity indicates binding. Positioning the sensing surface with respect to the standing wave pattern of the electric field within the cavity controls the sensitivity with which the presence of bound molecules is detected. Transmitted intensity at thousands of pixel locations is recorded simultaneously in a 10 s, 5 nm scan. An initial proof-of-principle setup has been constructed. A test sample was fabricated with 25, 100-mum wide square features, each with a different density of 1-mum square depressions etched 12 nm into the SiO(2) surface. The average depth of each etched region was found with 0.05 nm rms precision. In a second test, avidin, bound selectively to biotin conjugated bovine serum albumin, was detected.
Direct Observation of Conformation of a Polymeric Coating with Implications in Microarray Applications
The conformation of a three-dimensional polymeric coating (copoly(DMA-NAS-MAPS)) and immobilization and hybridization of DNA strands on the polymer coated surface are investigated. A conformational change, specifically the swelling of the surface adsorbed polymer upon hydration, is quantified in conjunction with the application of this polymer coating for DNA microarray applications. Fluorescently labeled short DNA strands (23mers) covalently linked to the functional groups on the adsorbed polymer are used as probes to measure the swelling of the polymer. A fluorescence microscopy technique, Spectral Self-Interference Fluorescence Microscopy (SSFM), is utilized to directly measure the change in axial position of fluorophores due to swelling with subnanometer accuracy. Additionally, immobilization characteristics of single stranded DNA (ssDNA) and double stranded DNA (dsDNA) probes, as well as hybridization of ssDNA with target strands have been studied. The results show that ssDNA further away from the surface is hybridized more efficiently, which strengthens the earlier analysis of this polymeric coating as a simple but highly efficient and robust DNA microarray surface chemistry.
Closed-form Representations of Field Components of Fluorescent Emitters in Layered Media
Dipole radiation in and near planar stratified dielectric media is studied theoretically within the context of fluorescence microscopy, as fluorescent emitters are generally modeled by electric dipoles. Although the main emphasis of this study is placed on the closed-form representations of the field components of fluorescent emitters in layered environments in near- and far-field regions, the underlying motive is to understand the limits of spectral self-interference fluorescence microscopy in studying the dipole orientation of fluorophores. Since accurate calculations of the field components of arbitrarily polarized electric dipoles in layered environments are computationally very time-consuming, a method for finding their closed-form representations is proposed using the closed-form potential Green's functions previously developed for microwave applications. The method is verified on typical geometries used in spectral self-interference microscopy experiments, where a dipole emitter is positioned over a slab of SiO(2) on top of a Si substrate. In addition to facilitating efficient calculation of near and intermediate fields of fluorescent emitters, closed-form Green's functions for fields would also play a crucial role in developing efficient and rigorous computational analysis and design tools for optical passive devices such as optical antennas by significantly improving the computational cost of the numerical solution of the integral equation.
Quantification of DNA and Protein Adsorption by Optical Phase Shift
A primary advantage of label-free detection methods over fluorescent measurements is its quantitative detection capability, since an absolute measure of adsorbed material facilitates kinetic characterization of biomolecular interactions. Interferometric techniques relate the optical phase to biomolecular layer density on the surface, but the conversion factor has not previously been accurately determined. We present a calibration method for phase shift measurements and apply it to surface-bound bovine serum albumin, immunoglobulin G, and single-stranded DNA. Biomolecules with known concentrations dissolved in salt-free water were spotted with precise volumes on the array surface and upon evaporation of the water, left a readily calculated mass. Using our label-free technique, the calculated mass of the biolayer was compared with the measured thickness, and we observed a linear dependence over 4 orders of magnitude. We determined that the widely accepted conversion of 1 nm of thickness corresponds to approximately 1 ng/mm(2) surface density held reasonably well for these substances and through our experiments can now be further specified for different types of biomolecules. Through accurate calibration of the dependence of thickness on surface density, we have established a relation allowing precise determination of the absolute number of molecules for single-stranded DNA and two different proteins.
Biaxial Strain in Graphene Adhered to Shallow Depressions
Measurements on graphene exfoliated over a substrate prepatterned with shallow depressions demonstrate that graphene does not remain free-standing but instead adheres to the substrate despite the induced biaxial strain. The strain is homogeneous over the depression bottom as determined by Raman measurements. We find higher Raman shifts and Gruneisen parameters of the phonons underlying the G and 2D bands under biaxial strain than previously reported. Interference modeling is used to determine the vertical position of the graphene and to calculate the optimum dielectric substrate stack for maximum Raman signal.
Label-free Multiplexed Virus Detection Using Spectral Reflectance Imaging
We demonstrate detection of whole viruses and viral proteins with a new label-free platform based on spectral reflectance imaging. The Interferometric Reflectance Imaging Sensor (IRIS) has been shown to be capable of sensitive protein and DNA detection in a real time and high-throughput format. Vesicular stomatitis virus (VSV) was used as the target for detection as it is well-characterized for protein composition and can be modified to express viral coat proteins from other dangerous, highly pathogenic agents for surrogate detection while remaining a biosafety level 2 agent. We demonstrate specific detection of intact VSV virions achieved with surface-immobilized antibodies acting as capture probes which is confirmed using fluorescence imaging. The limit of detection is confirmed down to 3.5 × 10(5)plaque-forming units/mL (PFUs/mL). To increase specificity in a clinical scenario, both the external glycoprotein and internal viral proteins were simultaneously detected with the same antibody arrays with detergent-disrupted purified VSV and infected cell lysate solutions. Our results show sensitive and specific virus detection with a simple surface chemistry and minimal sample preparation on a quantitative label-free interferometric platform.
Transfer of CVD-grown Monolayer Graphene Onto Arbitrary Substrates
Reproducible dry and wet transfer techniques were developed to improve the transfer of large-area monolayer graphene grown on copper foils by chemical vapor deposition (CVD). The techniques reported here allow transfer onto three different classes of substrates: substrates covered with shallow depressions, perforated substrates, and flat substrates. A novel dry transfer technique was used to make graphene-sealed microchambers without trapping liquid inside. The dry transfer technique utilizes a polydimethylsiloxane frame that attaches to the poly(methyl methacrylate) spun over the graphene film, and the monolayer graphene was transferred onto shallow depressions with 300 nm depth. The improved wet transfer onto perforated substrates with 2.7 μm diameter holes yields 98% coverage of holes covered with continuous films, allowing the ready use of Raman spectroscopy and transmission electron microscopy to study the intrinsic properties of CVD-grown monolayer graphene. Additionally, monolayer graphene transferred onto flat substrates has fewer cracks and tears, as well as lower sheet resistance than previous transfer techniques. Monolayer graphene films transferred onto glass had a sheet resistance of ∼980 Ω/sq and a transmittance of 97.6%. These transfer techniques open up possibilities for the fabrication of various graphene devices with unique configurations and enhanced performance.
Single Nanoparticle Detectors for Biological Applications
Nanoparticle research has become increasingly important in the context of bioscience and biotechnology. Practical use of nanoparticles in biology has significantly advanced our understanding about biological processes in the nanoscale as well as led to many novel diagnostic and therapeutic applications. Besides, synthetic and natural nanoparticles are of concern for their potential adverse effect on human health. Development of novel detection and characterization tools for nanoparticles will impact a broad range of disciplines in biological research from nanomedicine to nanotoxicology. In this article, we discuss the recent progress and future directions in the area of single nanoparticle detectors with an emphasis on their biological applications. A brief critical overview of electrical and mechanical detection techniques is given and a more in-depth discussion of label-free optical detection techniques is presented.
