reponse to therapy: Rapid response sample type: plasma disease state: hepatatis B individual: patient 4606
Extracted molecule
total RNA
Extraction protocol
Total RNA was extracted and purified using mirVana™ miRNA Isolation Kit (Cat#AM1560, Ambion, Austin, TX, US) following the manufacturer’s instructions and checked for a RIN number to inspect RNA integration by an Agilent Bioanalyzer 2100 (Agilent technologies, Santa Clara, CA, US).
Label
Cy3
Label protocol
Total RNA was amplified and labeled by Low RNA Input Linear Amplification kit (Cat#5184-3523, Agilent technologies, Santa Clara, CA, US), 5-(3-aminoallyl)-UTP (Cat#AM8436, Ambion, Austin, TX,US), Cy3 NHS ester (Cat#PA13105,GE healthcare Biosciences, Pittsburgh, PA,US) followed the manufacturer’s instructions. Labeled cRNA were purified by RNeasy mini kit (Cat#74106, QIAGEN, GmBH, Germany).
Hybridization protocol
Each Slide was hybridized with 1.65μg Cy3-labeled cRNA using Gene Expression Hybridization Kit (Cat#5188-5242, Agilent technologies, Santa Clara, CA, US) in Hybridization Oven (Cat#G2545A, Agilent technologies, Santa Clara, CA, US), according to the manufacturer’s instructions. After 17 hours hybridization, slides were washed in staining dishes (Cat#121, Thermo Shandon, Waltham, MA, US) with Gene Expression Wash Buffer Kit(Cat#5188-5327, Agilent technologies, Santa Clara, CA, US), Stabilization and Drying Solution (Cat#5185-5979, Agilent technologies, Santa Clara, CA, US)followed the manufacturer’s instructions。
Scan protocol
Slides were scanned by Agilent Microarray Scanner (Cat#G2565BA, Agilent technologies, Santa Clara, CA, US).
Description
miRNA expression in plamsa
Data processing
The scanned images were analyzed with Feature Extraction software 10.7(Agilent technologies, Santa Clara, CA, US) with default settings，Scan resolution=5μm, PMT 100%，10%.
