RESUMEN
Strategies for rapid, effective nucleic acid processing hold tremendous significance to the clinical analysis of circulating tumor DNA (ctDNA), a family of important markers indicating tumorigenesis and metastasis. However, traditional techniques remain challenging to achieve efficient DNA enrichment, further bringing about complicated operation and limited detection sensitivity. Here, we developed an ion concentration polarization microplatform that enabled highly rapid, efficient enrichment and purification of ctDNA from a variety of clinical samples, including serum, urine, and feces. The platform demonstrated efficiently separating and enriching ctDNA within 30 s, with a 100-fold improvement over traditional methods. Integrating an on-chip isothermal amplification module, the platform further achieved 100-fold enhanced sensitivity in ctDNA detection, which significantly eliminated false-negative results in the serum or urine samples due to the low abundance of ctDNA. Such a simple-designed platform offers a user-friendly yet powerful diagnosis technique with a wide applicability, ranging from early tumor diagnosis to infection screening.
Asunto(s)
ADN Tumoral Circulante , Neoplasias , Ácidos Nucleicos , Humanos , Neoplasias/diagnóstico , Neoplasias/genética , ADN Tumoral Circulante/genética , Carcinogénesis , Técnicas de Amplificación de Ácido Nucleico/métodosRESUMEN
The deceptively simple concepts of mass determination and fragment analysis are the basis for the application of mass spectrometry (MS) to a boundless range of analytes, including fundamental components and polymeric forms of nucleic acids (NAs). This platform affords the intrinsic ability to observe first-hand the effects of NA-active drugs on the chemical structure, composition, and conformation of their targets, which might affect their ability to interact with cognate NAs, proteins, and other biomolecules present in a natural environment. The possibility of interfacing with high-performance separation techniques represents a multiplying factor that extends these capabilities to cover complex sample mixtures obtained from organisms that were exposed to NA-active drugs. This report provides a brief overview of these capabilities in the context of the analysis of the products of NA-drug activity and NA therapeutics. The selected examples offer proof-of-principle of the applicability of this platform to all phases of the journey undertaken by any successful NA drug from laboratory to bedside, and provide the rationale for its rapid expansion outside traditional laboratory settings in support to ever growing manufacturing operations.
Asunto(s)
Ácidos Nucleicos , Ácidos Nucleicos/química , Espectrometría de Masas/métodos , Proteínas/químicaRESUMEN
Nucleic acids are natural biopolymers of nucleotides that store, encode, transmit and express genetic information, which play central roles in diverse cellular events and diseases in living things. The analysis of nucleic acids and nucleic acids-based analysis have been widely applied in biological studies, clinical diagnosis, environmental analysis, food safety and forensic analysis. During the past decades, the field of nucleic acids analysis has been rapidly advancing with many technological breakthroughs. In this review, we focus on the methods developed for analyzing nucleic acids, nucleic acids-based analysis, device for nucleic acids analysis, and applications of nucleic acids analysis. The representative strategies for the development of new nucleic acids analysis in this field are summarized, and key advantages and possible limitations are discussed. Finally, a brief perspective on existing challenges and further research development is provided.
RESUMEN
The KRAS oncogene is involved in the pathogenesis of several types of cancer, particularly colorectal cancer (CRC). The most frequent mutations in this gene are associated with poor survival, increased tumor aggressiveness and resistance to therapy with anti-epidermal growth factor receptor (EGFR) antibodies. For this reason, KRAS mutation testing has become increasingly common in clinical practice for personalized cancer treatments of CRC patients. Detection methods for KRAS mutations are currently expensive, laborious, time-consuming and often lack of diagnostic sensitivity and specificity. In this study, we describe the development of a Lab-on-Chip assay for genotyping of KRAS mutational status. This assay, based on the In-Check platform, integrates microfluidic handling, a multiplex polymerase chain reaction (PCR) and a low-density microarray. This integrated sample-to-result system enables the detection of KRAS point mutations, including those occurring in codons 12 and 13 of exon 2, 59 and 61 of exon 3, 117 and 146 of exon 4. Thanks to its miniaturization, automation, rapid analysis, minimal risk of sample contamination, increased accuracy and reproducibility of results, this Lab-on-Chip platform may offer immediate opportunities to simplify KRAS genotyping into clinical routine.
Asunto(s)
Proteínas Proto-Oncogénicas p21(ras)/genética , Neoplasias Colorrectales , Análisis Mutacional de ADN , Genotipo , Humanos , Mutación , Reproducibilidad de los ResultadosRESUMEN
The gelatin-silver halide black and white prints represent an enormous photography heritage with a great value. Unaesthetic phenomena, the foxing stains that are caused by microbial growth on surface, have been described in stamps, drawings, books, and tissues but, until now, scarcely for photographic materials. In this study, a combination of various techniques, including culture-dependent and culture-independent approaches (RNA and DNA analysis), scanning electron microscopy-energy dispersive spectroscopy (SEM-EDS) and µ-Raman spectroscopy supported by X-ray fluorescence analysis (XRF), permitted to describe the microbial contamination dynamics of foxing stains present on the surface of two gelatin-silver halide photographs. The investigation provided also information on the effects of microbial activity on the materials' chemistry of the two prints. The action of microbial community resulted locally in either (a) formation of mixed aluminum-iron-potassium phosphate compounds that could be attributed to the hydrolytic activity of bacteria, (b) leaching of barite,