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Following dye incorporation and sample cleanup, sample cDNA and control cDNA were mixed together with spiked controls that aid in hybridization calibration, and then the mixed sample was hybridized to the microarray surface using a Maui Hybridization System (Biomicro, Salt Lake City, UT).
Then, 50 g of the mixed sample was transferred into the enrichment medium.
The mixed sample was centrifuged (10,000×g at 4°C) for 5 min to obtain the RNA pellet.
A volume of 25 µl (corresponding to ∼13 µl of plasma) of the mixed sample was then transferred onto filter paper and left to dry overnight.
The 10 μL mixed sample was used for 12% SDS-PAGE loading.
Five minutes later, the absorbance at 545 nm of the final mixed sample was measured.
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The basic principle of the approach is that a fluorescence pattern of a mixed sample is a superposition of the fluorescent patterns for each target.
On the other hand, novel mass peaks only observed in the mixed sample were discovered and may serve as biomarkers for mixtures or contaminations [83, 86].
Heavy and light samples were then mixed 1 1, and 4 μg of each mixed sample were in-solution reduced, alkylated and digested with trypsin.
CaMKII is activated by Ca2+/CaM and ATP in one mixed sample (blue squares) while the other mixed sample is unactivated (red circles).
The flowability of the mixed samples was tested immediately after mixing following ASTM D6103 (2004).
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