Construction and validation of the APOCHIP, a spotted oligo-microarray for the study of beta-cell apoptosis.
BACKGROUND
Type 1 diabetes mellitus (T1DM) is a autoimmune disease caused by a long-term negative balance between immune-mediated beta-cell damage and beta-cell repair/regeneration. Following immune-mediated damage the beta-cell fate depends on several genes up- or down-regulated in parallel and/or sequentially. Based on the information obtained by the analysis of several microarray experiments of beta-cells exposed to pro-apoptotic conditions (e.g. double stranded RNA (dsRNA) and cytokines), we have developed a spotted rat oligonucleotide microarray, the APOCHIP, containing 60-mer probes for 574 genes selected for the study of beta-cell apoptosis.


RESULTS
The APOCHIP was validated by a combination of approaches. First we performed an internal validation of the spotted probes based on a weighted linear regression model using dilution series experiments. Second we profiled expression measurements in ten dissimilar rat RNA samples for 515 genes that were represented on both the spotted oligonucleotide collection and on the in situ-synthesized 25-mer arrays (Affymetrix GeneChips). Internal validation showed that most of the spotted probes displayed a pattern of reaction close to that predicted by the model. By using simple rules for comparison of data between platforms we found strong correlations (rmedian= 0.84) between relative gene expression measurements made with spotted probes and in situ-synthesized 25-mer probe sets.


CONCLUSIONS
In conclusion our data suggest that there is a high reproducibility of the APOCHIP in terms of technical replication and that relative gene expression measurements obtained with the APOCHIP compare well to the Affymetrix GeneChip. The APOCHIP is available to the scientific community and is a useful tool to study the molecular mechanisms regulating beta-cell apoptosis.
Magnusson
Nils E
NE
Department of Clinical Biochemistry, Aarhus University Hospital, Denmark. nm@ki.au.dk
Cardozo
Alessandra K
AK
Kruhøffer
Mogens
M
Eizirik
Decio L
DL
Ørntoft
Torben F
TF
Jensen
Jens L
JL
D000465:Algorithms; D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism*; D003922:Diabetes Mellitus, Type 1 / Q000175:diagnosis / Q000378:metabolism*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D020869:Gene Expression Profiling / Q000295:instrumentation* / Q000379:methods; D017404:In Situ Hybridization, Fluorescence / Q000295:instrumentation* / Q000379:methods; D050417:Insulin-Secreting Cells / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation* / Q000379:methods; D051381:Rats; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D016207:Cytokines