Abstract: Microarray data is often utilized in inferring regulatory networks. Quantile normalization (QN) is a popular method to reduce array-to-array variation. We show that in the context of time series measurements QN may not be the best choice for this task, especially not if the inference is based on continuous time ODE model. We propose an alternative normalization method that is better suited for network inference from time series data.
Abstract: Microfluidic-based micromosaic technology has allowed the pattering of recognition elements in restricted micrometer scale areas with high precision. This controlled patterning enabled the development of highly multiplexed arrays multiple analyte detection. This arraying technology was first introduced in the beginning of 2001 and holds tremendous potential to revolutionize microarray development and analyte detection. Later, several microfluidic methods were developed for microarray application. In this review we discuss these novel methods and approaches which leverage the property of microfluidic technologies to significantly improve various physical aspects of microarray technology, such as enhanced imprinting homogeneity, stability of the immobilized biomolecules, decreasing assay times, and reduction of the costs and of the bulky instrumentation.
Abstract: Colorectal cancer, a clinically diverse disease, is a leading cause of cancer-related death worldwide. Application of novel molecular diagnostic tests, which are summarized in this article, may lead to an improved survival of colorectal cancer patients. Distinction of these applications is based on the different molecular principles found in colorectal cancer (CRC). Strategies for molecular analysis of single genes (as KRAS or TP53) as well as microarray based techniques are discussed. Moreover, in addition to the fecal occult blood testing (FOBT) and colonoscopy some novel assays offer approaches for early detection of colorectal cancer like the multitarget stool DNA test or the blood-based Septin 9 DNA methylation test. Liquid biopsy analysis may also exhibit great diagnostic potential in CRC for monitoring developing resistance to treatment. These new diagnostic tools and the definition of molecular biomarkers in CRC will improve early detection and targeted therapy of colorectal cancer.
Abstract: There is virtually an unlimited number of possible Tissue Microarray (TMA) applications in basic and clinical research and ultimately in diagnostics. However, to assess the functional importance of novel markers, researchers very often turn to cell line model systems. The appropriate choice of a cell line is often a difficult task, but the use of cell microarray (CMA) technology enables a quick screening of several markers in cells of different origins, mimicking a genomic-scale analysis. In order to improve the morphological evaluations of the CMA slides we harvested the cells by conventional trypsinization, mechanical scraping and cells grown on coverslips. We show that mechanical scraping is a good evaluation method since keeps the real morphology very similar to those grown on coverslips. Immunofluorescence images are of higher quality, facilitating the reading of the biomarker cellular and subcellular localization. Here, we describe CMA technology in stem cell research.
Abstract: Molecular prognostic markers are urgently needed in order to improve therapy decisions in prostate cancer. To better understand the requirements for biomarker studies, we re-analyzed prostate cancer tissue microarray immunohistochemistry (IHC) data from 39 prognosis markers in subsets of 50 – >10,000 tumors. We found a strong association between the “prognostic power” of individual markers and the number of tissues that should be minimally included in such studies. The prognostic relevance of more than 90% of the 39 IHC markers could be detected if ≥6400 tissue samples were analyzed. Studying markers of tissue quality, including immunohistochemistry of ets-related gene (ERG) and vimentin, and fluorescence in-situ hybridization analysis of human epidermal growth factor receptor 2 (HER2), we found that 18% of tissues in our tissue microarray (TMA) showed signs of reduced tissue preservation and limited immunoreactivity. Comparing the results of Kaplan-Meier survival analyses or associations to ERG immunohistochemistry in subsets of tumors with and without exclusion of these defective tissues did not reveal statistically relevant differences. In summary, our study demonstrates that TMA-based marker validation studies using biochemical recurrence as an endpoint require at least 6400 individual tissue samples for establishing statistically relevant associations between the expression of molecular markers and patient outcome if weak to moderate prognosticators should also be reliably identified.
Abstract: With the advent of new histopathological staining techniques (histochemistry, immunohistochemistry, in situ hybridization) and the discovery of thousands of new genes, mRNA, and proteins by molecular biology, the need grew for a technique to compare many different cells or tissues on one slide in a cost effective manner and with the possibility to easily track the identity of each specimen: the tissue array (TA). Basically, a TA consists of at least two different specimens per slide. TAs differ in the kind of specimens, the number of specimens installed, the dimension of the specimens, the arrangement of the specimens, the embedding medium, the technique to prepare the specimens to be installed, and the technique to construct the TA itself. A TA can be constructed by arranging the tissue specimens in a mold and subsequently pouring the mold with the embedding medium of choice. In contrast, preformed so-called recipient blocks consisting of the embedding medium of choice have punched, drilled, or poured holes of different diameters and distances in which the cells or tissue biopsies will be deployed manually, semi-automatically, or automatically. The costs of constructing a TA differ from a few to thousands of Euros depending on the technique/equipment used. Remarkably high quality TAs can be also achieved by low cost techniques.