Furthermore, the evaluation

Furthermore, the evaluation selleck chemicals Calcitriol of the huge amount of data collected by microarray analyses requires an extensive bioinformatics with multivariate statistical methods. However, the newer generation of real-time PCR instruments available with multiplex arrays enables the testing and diagnostic utilization of mRNA expression microarray data. These quantitative array real-time PCRs with 384-well plates give an opportunity for testing the selected marker panels on a large set of independent samples allowing the measuring of the expression of more than hundred genes simultaneously. For the sake of flexibility quantitative RT-PCR with multiple transcript panels are custom-designed [15]. Universal ProbeLibrary probes from Roche use a unique nucleotide chemistry called LNA (Locked Nucleic Acid), which allows very short (8�C9 bases) oligonucleotides to be efficient hybridization probes in real-time PCR assays.

Optimized primer pairs and UPL probes can make the array RT-PCR a robust, reliable, quick and cost effective gene expression analyzing method which can be suitable for daily diagnostic utilization in the future. Traditional histology may suffer from sampling bias due to biopsy orientation problems, therefore, critical areas including aberrant crypt foci, dysplastic areas or in situ carcinoma may remain hidden. Molecular based discrimination using mRNA expression can represent the whole sample to avoid this bias and support pathologists in coping with their growing workload of early cancer screening.

Furthermore, mRNA expression can reveal functional information beyond microscopy related to the biological behavior, tumor invasion, metastasic spread and therapeutic target expression in colorectal cancer. In this study, we applied whole genomic microarray analysis in order to identify gene expression profile alterations focusing on the dysplastic adenoma-carcinoma transition. Our aims were to identify characteristic transcript sets in order to develop diagnostic mRNA expression patterns for objective classification of benign and malignant colorectal diseases and to test the classificatory power of these markers on an independent sample set. Materials and Methods Patients and samples After informed consent of untreated patients, colon biopsy samples were taken during endoscopic intervention and stored in RNALater Reagent (Qiagen Inc, Germantown, US) at �C80��C.

Altogether 147 biopsy specimen (53/original set/and additionally 94 fresh frozen/independent Brefeldin_A set/samples) were analyzed in our study. Total RNA was extracted and Affymetrix microarray analysis was performed on biopsies of patients with tubulovillous/villous adenomas (n=29, 13 high-grade dysplastic and 16 with low-grade dysplasia), colorectal adenocarcinoma (n=27, 14 early and 13 advanced CRC) and of healthy normal controls (n=38).

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