共查询到20条相似文献,搜索用时 15 毫秒
1.
The use of microarrays for parallel screening of nucleic acid profiles has become an industry standard. Similar efforts for screening protein-protein interactions are gaining momentum, however, they remain limited by the requirement for relatively large sample volumes. One strategy for overcoming this problem is to significantly decrease the size and consequently the sample volume of the protein interaction assay. We report here on our progress over the last two years in the construction of ultraminiaturized, functional protein capture assays. Each one micron spot in these array-based assays covers less than 1/1000(th) of the surface area of a conventional microarray spot while still maintaining enough antibodies to provide a useful dynamic range. These nanoarray assays can be read by conventional optical fluorescence microscopy as well as by novel label-free methods such as atomic force microscopy. The size reduction realized by functional protein nanoarrays also creates opportunities for novel applications including highly multiplexed single cell analysis and integration with microfluidics and other "lab-on-a-chip" technologies. 相似文献
2.
In recent years, there have been significant advances in biochemical assay miniturization and integration of microchip-based technologies with combinatorial library screening for high-throughput and large-scale applications. Small-molecule microarrays, protein arrays and cell-based arrays and conventional DNA arrays as well as microfluidic approaches in HTS are discussed in this review. 相似文献
3.
Microfluidics enables biotechnological processes to proceed on a scale (microns) at which physical processes such as osmotic
movement, electrophoretic-motility and surface interactions become enhanced. At the microscale sample volumes and assay times
are reduced, and procedural costs are lowered. The versatility of microfluidic devices allows interfacing with current methods
and technologies. Microfluidics has been applied to DNA analysis methods and shown to accelerate DNA microarray assay hybridisation
times. The linking of microfluidics to protein analysis techologies, e.g. mass spectrometry, enables picomole amounts of peptide
to be analysed within a controlled micro-environment. The flexibility of microfluidics will facilitate its exploitation in
assay development across multiple biotechnological disciplines. 相似文献
4.
Stephenie D. Prokopec John D. Watson Daryl M. Waggott Ashley B. Smith Alexander H. Wu Allan B. Okey Raimo Pohjanvirta Paul C. Boutros 《RNA (New York, N.Y.)》2013,19(1):51-62
Profiling of mRNA abundances with high-throughput platforms such as microarrays and RNA-seq has become an important tool in both basic and biomedical research. However, these platforms remain prone to systematic errors and have challenges in clinical and industrial applications. As a result, it is standard practice to validate a subset of key results using alternate technologies. Similarly, clinical and industrial applications typically involve transitions from a high-throughput discovery platform to medium-throughput validation ones. These medium-throughput validation platforms have high technical reproducibility and reduced sample input needs, and low sensitivity to sample quality (e.g., for processing FFPE specimens). Unfortunately, while medium-throughput platforms have proliferated, there are no comprehensive comparisons of them. Here we fill that gap by comparing two key medium-throughput platforms—NanoString''s nCounter Analysis System and ABI''s OpenArray System—to gold-standard quantitative real-time RT-PCR. We quantified 38 genes and positive and negative controls in 165 samples. Signal:noise ratios, correlations, dynamic range, and detection accuracy were compared across platforms. All three measurement technologies showed good concordance, but with divergent price/time/sensitivity trade-offs. This study provides the first detailed comparison of medium-throughput RNA quantification platforms and provides a template and a standard data set for the evaluation of additional technologies. 相似文献
5.
Yair Glick Dorit Avrahami Efrat Michaely Doron Gerber 《Journal of visualized experiments : JoVE》2012,(66)
Rapidly increasing fields, such as systems biology, require the development and implementation of new technologies, enabling high-throughput and high-fidelity measurements of large systems. Microfluidics promises to fulfill many of these requirements, such as performing high-throughput screening experiments on-chip, encompassing biochemical, biophysical, and cell-based assays1. Since the early days of microfluidics devices, this field has drastically evolved, leading to the development of microfluidic large-scale integration2,3. This technology allows for the integration of thousands of micromechanical valves on a single device with a postage-sized footprint (Figure 1). We have developed a high-throughput microfluidic platform for generating in vitro expression of protein arrays (Figure 2) named PING (Protein Interaction Network Generator). These arrays can serve as a template for many experiments such as protein-protein 4, protein-RNA5 or protein-DNA6 interactions.The device consist of thousands of reaction chambers, which are individually programmed using a microarrayer. Aligning of these printed microarrays to microfluidics devices programs each chamber with a single spot eliminating potential contamination or cross-reactivity Moreover, generating microarrays using standard microarray spotting techniques is also very modular, allowing for the arraying of proteins7, DNA8, small molecules, and even colloidal suspensions. The potential impact of microfluidics on biological sciences is significant. A number of microfluidics based assays have already provided novel insights into the structure and function of biological systems, and the field of microfluidics will continue to impact biology. 相似文献
6.
A fluorescent resonance energy transfer (FRET)-based hybridization assay for detecting multiplex ligation-dependent probe amplification (MLPA) products has been developed, extending the diagnostic power of the technique and demonstrating the possibility of combining MLPA with microarrays for the detection of multiple mutations. FRET is one of the most commonly used detection techniques for hybridization assays. To investigate the applicability of FRET based detection of MLPA products, a sandwich assay was designed to detect gene copy number by exploiting an immobilized probe labeled with an acceptor dye, Alexa Fluor 555, which hybridises to specific PCR amplicons, followed by hybridization of a second probe labeled with the donor dye, Alexa Fluor 488. Following excitation of the Alexa Fluor 488, a FRET signal was produced only if a DNA sequence specific to the BRCA1 exon 13 was present in the test sample. We have verified this assay on a DNA sample of a patient carrying a heterozygous BRCA1 exon 13 deletion using male genomic DNA as control. Here we demonstrate that the DNA sample containing the heterozygous deletion generated a considerably reduced FRET signal as compared to the control male human DNA. Our results show that the FRET design presented in this study can differentiate between reduced copy numbers any genomic DNA sequence after MLPA analysis, and the reported format is applicable to multiplex detection of MLPA products, using microarrays, or optical biosensor arrays, and future work will focus on the demonstration of this. 相似文献
7.
DNA and RNA quantifications are widely used in biological and biomedical research. In the last ten years, many technologies have been developed to enable automated and high-throughput analyses. In this review, we first give a brief overview of how DNA and RNA quantifications are carried out. Then, five technologies (microarrays, SAGE, differential display, real time PCR and real competitive PCR) are introduced, with an emphasis on how these technologies can be applied and what their limitations are. The technologies are also evaluated in terms of a few key aspects of nucleic acids quantification such as accuracy, sensitivity, specificity, cost and throughput. 相似文献
8.
Stem cells exhibit promise in numerous areas of regenerative medicine. Their fate and function are governed by a combination of intrinsic determinants and signals from the local microenvironment, or niche. An understanding of the mechanisms underlying both embryonic and adult stem cell functions has been greatly enhanced by the recent development of several high-throughput technologies: microfabricated platforms, including cellular microarrays, to investigate the combinatorial effects of microenvironmental stimuli and large-scale screens utilizing small molecules and short interfering RNAs to identify crucial genetic and signaling elements. Furthermore, the integration of these systems with other versatile platforms, such as microfluidics and lentiviral microarrays, will continue to enable the detailed elucidation of stem cell processes, and thus, greatly contribute to the development of stem cell based therapies. 相似文献
9.
Shipra Solanki Chandra M. Pandey Rajinder K. Gupta Bansi D. Malhotra 《Biotechnology journal》2020,15(5)
One of the major challenges for scientists and engineers today is to develop technologies for the improvement of human health in both developed and developing countries. However, the need for cost‐effective, high‐performance diagnostic techniques is very crucial for providing accessible, affordable, and high‐quality healthcare devices. In this context, microfluidic‐based devices (MFDs) offer powerful platforms for automation and integration of complex tasks onto a single chip. The distinct advantage of MFDs lies in precise control of the sample quantities and flow rate of samples and reagents that enable quantification and detection of analytes with high resolution and sensitivity. With these excellent properties, microfluidics (MFs) have been used for various applications in healthcare, along with other biological and medical areas. This review focuses on the emerging demands of MFs in different fields such as biomedical diagnostics, environmental analysis, food and agriculture research, etc., in the last three or so years. It also aims to reveal new opportunities in these areas and future prospects of commercial MFDs. 相似文献
10.
Xu L Yu H Akhras MS Han SJ Osterfeld S White RL Pourmand N Wang SX 《Biosensors & bioelectronics》2008,24(1):99-103
A giant magnetoresistive (GMR) biochip based on spin valve sensor array and magnetic nanoparticle labels was developed for inexpensive, sensitive and reliable DNA detection. The DNA targets detected in this experiment were PCR products amplified from Human Papillomavirus (HPV) plasmids. The concentrations of the target DNA after PCR were around 10nM in most cases, but concentrations of 10pM were also detectable, which is demonstrated by experiments with synthetic DNA samples. A mild but highly specific surface chemistry was used for probe oligonucleotide immobilization. Double modulation technique was used for signal detection in order to reduce the 1/f noise in the sensor. Twelve assays were performed with an accuracy of approximately 90%. Magnetic signals were consistent with particle coverage data measured with Scanning Electron Microscopy (SEM). More recent research on microfluidics showed the potential of reducing the assay time below one hour. This is the first demonstration of magnetic DNA detection using plasmid-derived samples. This study provides a direct proof that GMR sensors can be used for biomedical applications. 相似文献
11.
《Expert review of proteomics》2013,10(1):41-48
DNA microarrays have changed the field of biomedical sciences over the past 10 years. For several reasons, antibody and other protein microarrays have not developed at the same rate. However, protein and antibody arrays have emerged as a powerful tool to complement DNA microarrays during the past 5 years. A genome-scale protein microarray has been demonstrated for identifying protein–protein interactions as well as for rapid identification of protein binding to a particular drug. Furthermore, protein microarrays have been shown as an efficient tool in cancer profiling, detection of bacteria and toxins, identification of allergen reactivity and autoantibodies. They have also demonstrated the ability to measure the absolute concentration of small molecules. Besides their capacity for parallel diagnostics, microarrays can be more sensitive than traditional methods such as enzyme-linked immunosorbent assay, mass spectrometry or high-performance liquid chromatography-based assays. However, for protein and antibody arrays to be successfully introduced into diagnostics, the biochemistry of immunomicroarrays must be better characterized and simplified, they must be validated in a clinical setting and be amenable to automation or integrated into easy-to-use systems, such as micrototal analysis systems or point-of-care devices. 相似文献
12.
DNA microarrays have changed the field of biomedical sciences over the past 10 years. For several reasons, antibody and other protein microarrays have not developed at the same rate. However, protein and antibody arrays have emerged as a powerful tool to complement DNA microarrays during the past 5 years. A genome-scale protein microarray has been demonstrated for identifying protein-protein interactions as well as for rapid identification of protein binding to a particular drug. Furthermore, protein microarrays have been shown as an efficient tool in cancer profiling, detection of bacteria and toxins, identification of allergen reactivity and autoantibodies. They have also demonstrated the ability to measure the absolute concentration of small molecules. Besides their capacity for parallel diagnostics, microarrays can be more sensitive than traditional methods such as enzyme-linked immunosorbent assay, mass spectrometry or high-performance liquid chromatography-based assays. However, for protein and antibody arrays to be successfully introduced into diagnostics, the biochemistry of immunomicroarrays must be better characterized and simplified, they must be validated in a clinical setting and be amenable to automation or integrated into easy-to-use systems, such as micrototal analysis systems or point-of-care devices. 相似文献
13.
Yamashita K Yamaguchi Y Miyazaki M Nakamura H Shimizu H Maeda H 《Analytical biochemistry》2004,332(2):274-279
Conformational control of macromolecules is useful for efficient chemical and biochemical reactions. This article reports a direct observation method for macromolecules, such as long-strand DNA, in microchannel flow as well as a simple method for stretching DNA strands by microfluidics. Stretching and orientation of DNA molecules by control of flow within a microchannel was observed by optical microscopy. This DNA stretching is explained by coil-stretch transition of polymer molecules. This technique is useful for creating chemical reactions with macromolecules. It offers high selectivity and efficiency that are impossible to achieve in bulk solution. We also demonstrate that our microfluidic stretching method can accomplish efficient hybridization of long-strand DNA. This method will be useful for direct hybridization assay of long-strand DNA. 相似文献
14.
Use of a mixed tissue RNA design for performance assessments on multiple microarray formats 总被引:1,自引:0,他引:1
Thompson KL Rosenzweig BA Pine PS Retief J Turpaz Y Afshari CA Hamadeh HK Damore MA Boedigheimer M Blomme E Ciurlionis R Waring JF Fuscoe JC Paules R Tucker CJ Fare T Coffey EM He Y Collins PJ Jarnagin K Fujimoto S Ganter B Kiser G Kaysser-Kranich T Sina J Sistare FD 《Nucleic acids research》2005,33(22):e187
15.
Multiphase electrodes for microbead control applications: integration of DEP and electrokinetics for bio-particle positioning 总被引:1,自引:0,他引:1
Advances in microfabrication have introduced new possibilities for automated, high-throughput biomedical investigations and analysis. Physical effects such as dielectrophoresis (DEP) and AC electrokinetics can be used to manipulate particles in solution to coordinate a sequence of bioanalytical processing steps. DEP is accomplished with non-uniform electric fields that can polarize particles (microbeads, cells, viruses, DNA, proteins, etc.) in suspension causing translational or rotational movement. AC electrokinetics is another phenomena involved with movement of particles in suspension with electric fields and is comprised of both electro-thermal and electro-osmotic effects. This paper investigates single layer electrodes that are effective for particle localization and clustering based on DEP and AC electrokinetic effects. We demonstrate a novel multi-electrode setup capable of clustering particles into an array of discrete bands using activated and electrically floating electrodes. These bands shift to adjacent regions on the electrode surface by altering the electrode activation scheme. The predictability of particle placement to specific locations provides new opportunities for integration and coordination with raster scanning lasers or a charge coupled device (CCD) for advanced biomedical diagnostic devices, and more sophisticated optical interrogation techniques. 相似文献
16.
Devarajan K 《PLoS computational biology》2008,4(7):e1000029
In the last decade, advances in high-throughput technologies such as DNA microarrays have made it possible to simultaneously measure the expression levels of tens of thousands of genes and proteins. This has resulted in large amounts of biological data requiring analysis and interpretation. Nonnegative matrix factorization (NMF) was introduced as an unsupervised, parts-based learning paradigm involving the decomposition of a nonnegative matrix V into two nonnegative matrices, W and H, via a multiplicative updates algorithm. In the context of a pxn gene expression matrix V consisting of observations on p genes from n samples, each column of W defines a metagene, and each column of H represents the metagene expression pattern of the corresponding sample. NMF has been primarily applied in an unsupervised setting in image and natural language processing. More recently, it has been successfully utilized in a variety of applications in computational biology. Examples include molecular pattern discovery, class comparison and prediction, cross-platform and cross-species analysis, functional characterization of genes and biomedical informatics. In this paper, we review this method as a data analytical and interpretive tool in computational biology with an emphasis on these applications. 相似文献
17.
Design and fabrication of a silica on silicon integrated optical biochip as a fluorescence microarray platform 总被引:2,自引:0,他引:2
Ruano JM Glidle A Cleary A Walmsley A Aitchison JS Cooper JM 《Biosensors & bioelectronics》2003,18(2-3):175-184
Previous research into the use of Flame Hydrolysis Deposition (FHD) of glasses in integrated optics has focused on the successful commercial exploitation of low cost optical devices within the field of telecommunications and optoelectronics. Recently we have sought to apply these fabrication technologies to the development of optical biochips, utilising their ability to be integrated with microfluidics as a 'Lab-on-a-chip' platform. In this paper, we carry this development forward by seeking to create a microarray of integrated optical sensing elements, addressed using a glass-polymer hybrid technology in which poly(dimethylsiloxane), PDMS, is used as an elastomeric packaging over-layer. In particular, we describe the wide range of modelling and microfabrication processes required for the successful manufacture, integration and packaging of such arrays. The integration of both optical and fluidic circuits in this device avoids precise alignment requirements and results in a compact, robust and reliable device. Finally, in this paper, we describe the implementation of a pumping system for delivering small amounts of fluid across the array together with an optical signal treatment. 相似文献
18.
Gabibov A Yakubovskaya E Lukin M Favorov P Reshetnyak A Monastyrsky M 《The FEBS journal》2005,272(24):6336-6343
A catalytic turnover of supercoiled DNA (scDNA) transformation mediated by topoisomerases leads to changes in the linking number (Lk) of the polymeric substrate by 1 or 2 per cycle. As a substrate of the topoisomerization reaction it is chemically identical to its product; even a single catalytic event results in the quantum leap in the scDNA topology. Non-intrusive continuous assay to measure the kinetics of the scDNA topoisomerization was performed. The development of such a technique was hindered because of multiple DNA species of intermediate topology present in the reaction mixture. The interrelation of DNA topology, its hydrodynamics, and optical anisotropy enable us to use the flow linear dichroism technique (FLD) for continuous monitoring of the scDNA topoisomerization reaction. This approach permits us to study the kinetics of DNA transformation catalyzed by eukaryotic topoisomerases I and II, as well as mechanistic characteristics of these enzymes and their interactions with anticancer drugs. Moreover, FLD assay can be applied to any enzymatic reaction that involves scDNA as a substrate. It also provides a new way of screening drugs dynamically and is likely to be potent in various biomedical applications. 相似文献
19.
20.
Dielectrophoretic platforms for bio-microfluidic systems 总被引:2,自引:0,他引:2
Khoshmanesh K Nahavandi S Baratchi S Mitchell A Kalantar-zadeh K 《Biosensors & bioelectronics》2011,26(5):1800-1814
Dielectrophoresis, the induced motion of polarisable particles in a nonuniform electric field, has been proven as a versatile mechanism to transport, accumulate, separate and characterise micro/nano scale bioparticles in microfluidic systems. The integration of DEP systems into the microfluidics enables the inexpensive, fast, highly sensitive, highly selective and label-free detection and analysis of target bioparticles. This review provides an in-depth overview of state-of-the-art dielectrophoretic (DEP) platforms integrated into microfluidics aimed towards different biomedical applications. It classifies the current DEP systems in terms of different microelectrode configurations and operating strategies devised to generate and employ DEP forces in such processes, and compares the features of each approach. Finally, it suggests the future trends and potential applications of DEP systems in single cell analysis, stem cell research, establishing novel devices, and realising fully DEP-activated lab-on-a-chip systems. 相似文献