![]() | ||||||||
Molecular
Diagnostic Laboratory |
||||||||
|
Nordic
Centre of Excellence in Molecular Medicine |
||||||||
|
|
||||||||
|
Functional Genomics and Marker Validation Aims and Vision
Molecules were identified by extensive profiling studies on hundreds of tissue samples by transcript microarray profiling using Affymetrix GeneChips U133plus2.0, microRNA expression profiling using Exiqon Locked Nucleic Acid (LNA™) detection technology and/or Applied Biosystems TaqMan® Low Density Array (microfluidic cards), Methylation profiling using Illumina’s HumanMethylation27 DNA Analysis BeadChip or SNP analyses using Affymetrix Genome-Wide Human SNP Array 6.0.
Research Strategy
Each project has its origin in the box headed “Biobank”,
where samples are collected and nucleic acids are extracted, quality
controlled and stored (for detailed information, see CRC or bladder
cancer page). Expression of gene encoded proteins and their localization within the tumor cell is analyzed by immunohistochemistry using cryosections in Tissue Tek or FFPE tissue specimen.
For protein detection, antibody specificity is checked on Western
Blots using cell lines overexpressing the molecule of interest.
In situ hybridization (ISH) using LNA probes is used
to localize microRNAs within the tumor cells.
Cell Line authentication In case of microRNAs cells are transfected with miR-precursors or Anti-miRs, miRNA Inhibitors targeting human miRNAs (Exiqon). Successful up- or downregulation is monitored by RTPCR (Taqman) and Western blotting for protein.
Cells with an overexpressed or silenced molecule of interest are continuously monitored by Real Time Cell Analysis RTCA (Roche X-celligence) over a period of up to 5 days to identify whether the molecule of interest impacts cellular processes, e.g. increases proliferation or causes cell death. Standard cellular assays monitoring proliferation, viability or cytotoxicity are performed.
Data analyses in MOMA are performed on a Dual Plate RTCA instrument,
offering highest possible flexibility for several different approaches
in parallel.
Fluorescence microscopy visualizes the cellular localization of proteins within living or fixed cells from cell lines, from cryosections or from FFPE tissue samples.
For Pathway analyses, to gain insight into networks or to identify
downstream target molecules, cell extracts from manipulated cells, overexpressing
a molecule of interest or with a knock down, are subjected to Microarray
expression profiling. Bioinformatic data processing is followed by application
of Ingenuity Pathway analyses software to identify pathways or networks
addressed in common for both, the tumor samples as well as the manipulated
cell lines. (K. Demtröder, unpublished data) Future Research Activities
Group Leader: Karin Birkenkamp-Demtröder, Associate professor, MSci, PhD, kbdr@ki.au.dk |
||||||||
|
|
||||||||
|
|
||||||||
|
MOMA,
Aarhus University Hospital, Skejby, Brendstrupgårdsvej 100, 8200
Århus N, Denmark | ||||||||