What signal and fold-change concordance can I expect between Affymetrix miRNA 4.0 and miRNA 4.1 Arrays and miRNA 3.0 and miRNA 3.1 Arrays?
You should expect high signal and fold-change concordance for transcripts with identical probes on the miRNA 4.0 and miRNA 4.1 Arrays and the miRNA 3.0 and miRNA 3.1 Arrays. The snoRNA, scaRNA, and precursor miRNA probe sets were reselected, and for some probe sets in these categories, the probes are not identical between the two array designs. Because the probes selected may be different, one may observe changes in the probe set signal summary for a given target. This is possible as different probe sequences will have varying affinity to the target and may show a higher or lower absolute probe signal depending on the GC content of the probes. However, fold changes are typically well-correlated to the legacy probe set unless there is cross-hybridization in the legacy probe set. Our expectation is that more unique and specific probes result in more specific hybridization to the intended target, and therefore a better performing probe set. Target preparation replicates of human brain and lung samples were pooled and hybridized to four miRNA 3.0 Arrays and four miRNA 4.0 Arrays. The Pearson product moment correlation was calculated from the median signal of matched probe sets and from the median fold change of detected matched probe sets on the miRNA 3.0 Array compared to the miRNA 4.0 Array. Probe sets were defined as detected if the median DABG p-value of the four miRNA 3.0 Array brain replicates was less than 0.06. Two subsets of matched probe sets were used for miRNA 3.0 Array to miRNA 4.0 Array signal and fold change correlation: All human probe sets including those that do not necessarily share identical probe sequences and human probe sets for which the probe sequences are identical on both the miRNA 3.0 and miRNA 4.0 Arrays.
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Do we have to use any Affymetrix Reagents with our assay?
GeneChip Hybridization Control Kit 30 reactions
GeneTitan Hybridization, Wash, and Stain Kit for miRNA Array Plates 96 reactions
GeneTitan Hybridization Module for miRNA Plates 96 reactions
Affymetrix FlashTag Biotin HSR RNA Labeling Kit 10 reactions
Affymetrix FlashTag Biotin HSR RNA Labeling Kit 30 reactions
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What kind of dynamic range can I expect from Affymetrix miRNA Array Plates?
Affymetrix miRNA Array Plate provides a similar dynamic range to GeneChip miRNA Array.
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How do results from GeneChip miRNA Cartridge Array compare to Affymetrix miRNA Array Plates?
When evaluating non-control probe sets in common to both arrays, we observed very high signal and fold-change correlation for GeneChip miRNA Cartridge Arrays compared to Affymetrix miRNA Array Plates (Pearson correlation coefficients of 0.97-0.98 for signal correlation and Pearson correlation coefficients of 0.96-0.98 for fold-change correlation).
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What is the tolerance around hybridization time when working with Affymetrix miRNA Array strips?
We recommend maintaining consistency in array processing protocols to minimize introduction of variability. Our recommended hybridization time for Thermo Fisher Scientific miRNA Array Strip is 20 hrs.
Thermo Fisher Scientific miRNA Array Strip should be hybridized no less than 20 hrs and no longer than 24 hrs. We strongly encourage maintaining consistent hybridization times to avoid false positives with respect to differential expression due to hybridization time.
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