New MI PathArray™ assays focus on research area-specific pathways in development and disease, bringing the highest microarray sensitivity to your research. Gene expression analysis is often performed using whole-genome microarrays resulting in very large data sets that generate information irrelevant to the focused researcher. Alternative methods for focused gene expression currently utilize qPCR, but these tools have limited gene coverage due to high assay costs and limited instrument capabilities. The MI PathArray microarray platform provides an alternative : a focused gene pathway tool with current scientifically-relevant gene content providing manageable data sets for focused analysis in a cost-effective multi-assay platform.
Each multi-assay slide includes four discrete, identical arrays containing pathway-specific collection of 70mer oligonucleotides selected through a vigorous curation process from the MI collection of human gene probes. Probes and controls are printed in triplicate in each assay area offering increased sensitivity and statistical power. Control probes for every assay include 17 probes for widely expressed genes (positive controls), 6 negative probes, and a series of 8 probes for use with spike-in controls used to determine the limits of detectability.
Designed for use with pre-labeled cDNA derived from total RNA, detailed sample preparation protocols for MI PathArray assays to achieve semi-quantitative results are provided. RNA spike quantities recommended for inclusion included in the reverse transcription and labeling process generate a fluorescence/expression curve to allow semi-quantitative analysis.
Hybridization your samples on the MI PathArray is readily accomplished using any commercially-available hybridization systems, or with MI-developed protocols for bench-top static hybridization methods. Additionally, the printed dimensions of the assay areas have been designed to allow for use in the Tecan HS Pro and other commercially available hybridization systems.
Each MI PathArray contains a collection of targeted pathway-specific probes from MI’s human gene probe collection. MI PathArray assays provide content that allow researchers to advance confidently from other technologies (including PCR arrays or smaller microarrays) that typically provide fewer probes.
Content is selected through a vigorous curation process which evaluates many factors for inclusion.
Current MI PathArray assays | Catalog # | # Probes | Genes Covered | transcripts covered | |
---|---|---|---|---|---|
Growth Factors | MPA010 | 190 | 142 | >390 | more details |
Inflammation | MPA020 | 737 | 677 | >2450 | more details |
Apoptosis | MPA030 | 1048 | 994 | >3600 | more details |
Cancer | MPA040 | 509 | 465 | >1950 | more details |
Transcription Factors | MPA050 | 946 | 859 | >3500 | more details |
Kinases | MPA060 | 1013 | 907 | >3600 | more details |
In addition to the MI PathArray array-specific probe content, 17 oligo probes for genes with high levels of constitutive expression (positive controls), eight RNA spike-in sense oligonucleotide probes, and six negative control spots enable a semi-quantitative analysis of gene expression. Both one- and two- color experiments are supported under these control oligonucleotides.
A set of positive control gene probes included on each MI PathArray was designed to include transcripts known to be of high, moderate, and low expression across a variety of tissue types. Expression of each positive control will be RNA-source dependent and can help to gauge expression levels of transcripts of interest in your sample. The following 70mer oligonucleotide probes for constitutively expressed genes are included on each MI PathArray in triplicate (Table 1).
ACTB | B2M |
---|---|
GAPDH | HPRT1 |
PPIA | RPL13A |
RPL37A | RPL41 |
PSMD4 | EIF3C |
EEF1A1 | EEF2 |
SRP14 | OAZ1 |
TPT1 | GUSB |
FTL |
A minimum of two housekeeping genes are included in each block of the array to demonstrate adequate hybridization of each sample.
Negative control probes are also included in triplicate in each block of the array. These spots are made of several 70mer DNA oligonucleotides with absence of complimentarity to known human DNA sequences. These negative control sequences are included in the array to gauge non-specific binding present after hybridization and post-hybridization washing.
RNA spike sense oligonucleotide probes are present in triplicate in each block of the array. These 70mer probe sequences, provided by Ambion Inc., are complimentary to the Ambion ArrayControlTM RNA spikes available through Life Technologies (Invitrogen, Ambion, and Applied Biosystems). MI protocols provide guidelines for RNA spike quantities to be included in the reverse transcription and labeling process. These quantities generate a fluorescence/expression curve by which semi-quantitative interpretation can be achieved for your transcripts of interest.
ASSAYS | Genelist | QC file | GAL file |
---|---|---|---|
MPA010 GROWTH FACTORS | MPA010-growth-factor.xls | MPA010-growth-factors-qc.xls | MPA010-growth-factors.gal |
MPA020 INFLAMMATION | MPA020-inflammation.xls | MPA020-inflammation-qc.xls | MPA020-inflammation.gal |
MPA030 APOPTOSIS | MPA030-apoptosis.xls | MPA030-apoptosis-qc.xls | MPA030-apoptosis.gal |
MPA040 CANCER | MPA040-cancer.xls | MPA040-cancer-qc.xls | MPA040-cancer.gal |
MPA050 TRANSCRIPTION FACTORS | MPA050-transcription-factors-genelist.xls | Contact MI for more info | MPA050-transcription-factors.gal |
MPA060 KINASES | MPA060-kinases-genelist.xls | Contact MI for more info | Contact MI for more info |