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Biosens Bioelectron ; 54: 297-305, 2014 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-24287420


In this study, a microfluidic platform was developed to generate single layer, linear array of microbeads for multiplexed high-throughput analysis of biomolecules. The microfluidic device is comprised of eight microbead-trapping units, where microbeads were immobilized in a linear array format by the exertion of a negative pressure in the control channel connected to each sieving microstructure. Multiplexed assays were achieved by using a mixture of different spectrally-encoded microbeads functionalized with specific probes, followed by on-chip reaction and detection. The microfluidic-based microbeads array platform was employed for multiplexed analysis of DNA and proteins, as demonstrated by the simultaneous discrimination of four HPV genotypes and the parallel detection of six different proteins. Compared with the off-chip protocols, the on-chip analysis exhibited better reaction efficiency, higher sensitivity and wider linear detection range. Visual inspection and identification of functionalized microbeads were facilitated by the single layer arrangement of microbeads so that accurate data acquisition can be performed during the detection process.

Alphapapillomavirus/genética , DNA Viral/análise , Técnicas Analíticas Microfluídicas/instrumentação , Infecções por Papillomavirus/virologia , Proteínas/análise , Técnicas Biossensoriais/instrumentação , Linhagem Celular , DNA Viral/genética , Desenho de Equipamento , Genótipo , Humanos , Microesferas
Biosens Bioelectron ; 29(1): 89-96, 2011 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-21872460


A novel microfluidic device with microbeads array was developed and sensitive genotyping of human papillomavirus was demonstrated using a multiple-enzyme labeled oligonucleotide-Au nanoparticle bioconjugate as the detection tool. This method utilizes microbeads as sensing platform that was functionalized with the capture probes and modified electron rich proteins, and uses the horseradish peroxidase (HRP)-functionalized gold nanoparticles as label with a secondary DNA probe. The functionalized microbeads were independently introduced into the arrayed chambers using the loading chip slab. A single channel was used to generate weir structures to confine the microbeads and make the beads array accessible by microfluidics. Through "sandwich" hybridization, the enzyme-functionalized Au nanoparticles labels were brought close to the surface of microbeads. The oxidation of biotin-tyramine by hydrogen peroxide resulted in the deposition of multiple biotin moieties onto the surface of beads. This deposition is markedly increased in the presence of immobilized electron rich proteins. Streptavidin-labeled quantum dots were then allowed to bind to the deposited biotin moieties and displayed the signal. Enhanced detection sensitivity was achieved where the large surface area of Au nanoparticle carriers increased the amount HRP bound per sandwiched hybridization. The on-chip genotyping method could discriminate as low as 1fmol/L (10zmol/chip, SNR>3) synthesized HPV oligonucleotides DNA. The chip-based signal enhancement of the amplified assay resulted in 1000 times higher sensitivity than that of off-chip test. In addition, this on-chip format could discriminate and genotype 10copies/µL HPV genomic DNA using the PCR products. These results demonstrated that this on-chip approach can achieve highly sensitive detection and genotyping of target DNA and can be further developed for detection of disease-related biomolecules at the lowest level at their earliest incidence.

Sondas de DNA de HPV , Técnicas Analíticas Microfluídicas/métodos , Papillomaviridae/genética , Pontos Quânticos , Sequência de Bases , Colo do Útero/virologia , Sondas de DNA de HPV/genética , Desenho de Equipamento , Feminino , Genótipo , Ouro , Humanos , Nanopartículas Metálicas , Técnicas Analíticas Microfluídicas/instrumentação , Técnicas Analíticas Microfluídicas/estatística & dados numéricos , Papillomaviridae/classificação , Papillomaviridae/isolamento & purificação
Anal Bioanal Chem ; 397(7): 3143-50, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20563793


Patterning is of paramount importance in many areas of modern science and technology. As a good candidate for novel nanoscale optoelectronics and miniaturized molecule sensors, vertically aligned silicon nanowire (SiNW) with controllable location and orientation is highly desirable. In this study, we developed an effective procedure for the fabrication of vertically aligned SiNW arrays with micro-sized features by using single-step photolithography and silver nanoparticle-induced chemical etching at room temperature. We demonstrated that the vertically aligned SiNW arrays can be used as a platform for label-free DNA detection using surface-enhanced Raman spectroscopy (SERS), where the inherent "fingerprint" SERS spectra allows for the differentiation of closely related biospecies. Since the SiNW array patterns could be modified by simply varying the mask used in the photolithographic processing, it is expected that the methodology can be used to fabricate label-free DNA microarrays and may be applicable to tissue engineering, which aims to create living tissue substitutes from cells seeded onto 3D scaffolds.

DNA/análise , Nanofios/química , Silício/química , Sequência de Bases , DNA/genética , Dados de Sequência Molecular , Análise Espectral Raman
Front Biosci (Elite Ed) ; 2: 1015-27, 2010 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-20515772


Human papillomavirus (HPV) infection is a necessary factor in the development of cervical cancer. A new HPV screening method, "Human Papillomavirus Genotyping (HPG)", was developed to detect 29 HPV genotypes distribution in China. The utility of HPG was compared to Hybrid Capture 2 High-Risk HPV DNA test (HC2), and it was determined that the HPG test had been proven to be a more credible and sensitive screening HPV method than the HC2 test. HPV16, HPV 52, HPV 56, and HPV 58 were the four most common HPV genotypes in women who have suffered chronic cervicitis or abnormal vaginal bleeding in China. HPV 16 (28.57%) and 18 (17.86%) were more likely to infect multiple HPV genotypes than other HPV genotypes. Age group more than 50 years had a higher risk than other age groups.

Alphapapillomavirus/isolamento & purificação , Alphapapillomavirus/genética , Sequência de Bases , China , Primers do DNA , DNA Viral/genética , DNA Viral/isolamento & purificação , Feminino , Genótipo , Humanos , Reação em Cadeia da Polimerase