Enrico Alessi

Novel Biochip Platform for Nucleic Acid Analysis

This manuscript describes the use of a novel biochip platform for the rapid analysis/identification of nucleic acids, including DNA and microRNAs, with very high specificity. This approach combines a unique dynamic chemistry approach for nucleic acid testing and analysis developed by DestiNA Genomics with the STMicroelectronics In-Check platform, which comprises two microfluidic optimized and independent PCR reaction chambers, and a sequential microarray area for nucleic acid capture and identification by fluorescence. With its compact bench-top “footprint” requiring only a single technician to operate, the biochip system promises to transform and expand routine clinical diagnostic testing and screening for genetic diseases, cancers, drug toxicology and heart disease, as well as employment in the emerging companion diagnostics market.

Is this the real time for genomics

In the last decades, molecular biology has moved from gene-by-gene analysis to more complex studies using a genome-wide scale. Thanks to high-throughput genomic technologies, such as microarrays and next-generation sequencing, a huge amount of information has been generated, expanding our knowledge on the genetic basis of various diseases. Although some of this information could be transferred to clinical diagnostics, the technologies available are not suitable for this purpose. In this review, we will discuss the drawbacks associated with the use of traditional DNA microarrays in diagnostics, pointing out emerging platforms that could overcome these obstacles and offer a more reproducible, qualitative and quantitative multigenic analysis. New miniaturized and automated devices, called Lab-on-Chip, begin to integrate PCR and microarray on the same platform, offering integrated sample-to-result systems. The introduction of this kind of innovative devices may facilitate the transition of genome-based tests into clinical routine.

Early genomics of learning and memory: a review

The characterization of the molecular mechanisms whereby our brain codes, stores and retrieves memories remains a fundamental puzzle in neuroscience. Despite the knowledge that memory storage involves gene induction, the identification and characterization of the effector genes has remained elusive. The completion of the Human Genome Project and a variety of new technologies are revolutionizing the way these mechanisms can be explored. This review will examine how a genomic approach can be used to dissect and analyze the complex dynamic interactions involved in gene regulation during learning and memory. This innovative approach is providing information on a new class of genes associated with learning and memory in health and disease and is elucidating new molecular targets and pathways whose pharmacological modulation may allow new therapeutic approaches for improving cognition.

Developments of the in-check platform for diagnostic applications

In-Check is STMicroelectronics proprietary platform for molecular diagnostics. In-Check lays its foundations on the monolithic integration of microelectronics and micromachining technology MEMS, with microfluidic and optical features, bio-chemical surface functionalization and molecular biology. It comprises a core lab-on-chip device, control and reading instrumentation, a complete suite of software modules, and application protocols. Leveraging on such capabilities, In-Check enables fast, highly sensitive and specific, multi-analytical capability of nucleic acid analysis. The platform provides a unique combination of nucleic acid amplification, by polymerase-chain-reaction and target identification and typing by DNA microarray. These integrated biological functionalities together with top quality standard and process control are key features for a platform to be accepted by the highly demanding modern medical diagnostic. This paper describes recent developments of In-Check and some core biological characterizations.

Genotyping of KRAS Mutational Status by the In-Check Lab-on-Chip Platform

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.

4.2.2 InCheck System: a Highly Integrated Silicon Labonchip for Sample Preparation, PCR Amplification and Microarray Detection Towards the Molecular Diagnostics Pointofcare

The In-Check System is based on a miniaturized silicon lab-on-chip (LoC) where the Polymerase Chain reactor lives together with a customizable microarray module for running a seamless nucleic acid test [1]. This device is designed for accurate temperature performances control, such as accuracy and heating rate provided by both a sophisticated chip calibration process and a precise control by the Temperature Control System (TCS). In addition, the device, is optimized for a microarray fluorescence reading operation by an external instrument, the optical reader (OR). Finally, it is based on microfluidic features that enable to load the chip and fill the reaction chambers without the risk of bubble formation or leaks. In this manuscript are reported the experimental results for the detection of human betaglobine gene (HBB) directly from human cells in less than 2 hours in a silicon reactor. The sample preparation process was entirely performed in one single step into the silicon reactor. It was fully characterized by RT-qPCR. We performed also a comparison study showing higher performances in the LoC silicon reactor than the standard tube. Moreover, the DNA extracted was amplified by PCR, and the resulting product hybridized on the microarray. All the results suggest that the hybridization reactions performed on the silicon LoC can be used to exploit the discriminatory power of microarrays for a specific gene detection.