Agilent dna microarray scanner

Sevoflurane-treated PC12 and HAPI cells administrated with or without Table (40 µg/mL) were collected, and the total RNA, including miRNAs, was extracted using a miRNeasy Mini Kit. The RNA samples were pooled to perform miRNA microarray analysis, which were firstly labeled by the Agilent miRNA Complete Labeling and Hybridization Kit as per the manufacturer’s instructions, then were hybridized to the Agilent miRNA Microarray Release at 65℃ for 20 h. The results were scanned on an Agilent DNA Microarray Scanner with the Scan Control software (Agilent Technologies), and the results were imported into GenePix Pro 6.0 software for data extraction.

The hippocampal RNA was analyzed using Arraystar RNA Flash Labeling Kit. After RNA labeling and hybridization, slides were scanned by Agilent DNA Microarray Scanner. Raw data were normalized and analyzed using GeneSpring GX v12.1 software (Agilent Technologies, Santa Clara, CA). Differentially expressed mRNA between two groups were identified through p value (cut–off: 0.05) and FC (cut–off: 1.5) sifting. Pathway and gene ontology (GO) analysis were applied to determine the roles of these differentially expressed mRNAs on biological pathways or GO terms. Hierarchical clustering and combined analysis were performed using in-house scripts (Kangcheng Biotechnology Company). The microarray data were deposited in Gene Expression Omnibus (GEO accession: GSE166311).

Fluorescent signal intensities were measured on an Agilent DNA Microarray Scanner (Cat # G2565BA; Agilent Technologies) using the Scan Control A.8.4.1 Software (Agilent Technologies), and images were extracted using the Feature Extraction 10.7.3.1 Software (Agilent Technologies). Detailed methodology was from a previous publication [20 (link)].

ESC lines carrying a tetracycline-regulatable TF were derived from MC1 (129.3) cell line, which was obtained from the expanded frozen stock at Johns Hopkins University, as described previously2 3 (link). ESCs of passage 25 were cultured in the standard LIF+ medium with added Dox+ on a gelatin-coated dish through the experiments. Cells from each cell line were split into six wells and the media was changed 24 hours after cell plating: three wells with Dox+ medium, and three wells with Dox− medium to induce transgenic TFs. Dox was removed via washing three times with PBS at three-hour intervals. The proportion of Venus-p;ositive cells was evaluated by FACS (Canto II, Becton Dickinson). Total RNA was isolated by TRIzol (Invitrogen) after 48 hours, and two replications were used for microarray hybridization. RNA samples were labeled with total RNA by Low RNA Input Fluorescent Linear Amplification Kit (Agilent). We hybridized Cy3-CTP labeled sample from Dox− medium together with Cy5-CTP labeled sample from Dox+ medium (i.e., control) to the NIA Mouse 44K Microarray v3.0 (Agilent, design ID 015087)28 (link). Slides were scanned with Agilent DNA Microarray Scanner. All DNA Microarray data are available in Table S2, at GEO/NCBI (http://www.ncbi.nlm.nih.gov/geo; GSE72350), and at NIA Array Analysis, http://lgsun.grc.nia.nih.gov/ANOVA29 (link).

Total RNA samples (100 ng) were extracted from P-HUVECs and M-HUVECs. Hybridization and signal acquisition of Agilent human miRNA microarray (Release 10.1) (Agilent Technologies, Santa Clara, CA) carrying 723 human and 76 human viral miRNAs was performed using an Agilent DNA Microarray Scanner with Agilent ScanControl version 7.0 software. The analysis and background correction were performed using Agilent Feature Extraction Software ver. 9.5. All the microarray data in this work is deposited at GEO with an accession number of GSE56663

Arraystar Mouse LncRNA Microarray version 3.0 was designed for the global profiling of mouse lncRNAs and protein-coding transcripts, with ~35,923 lncRNAs and 24,881 coding transcripts detected. Sample preparation and microarray hybridization were performed based on manufacturer standard protocols with minor modifications. An Arraystar RNA Flash Labeling Kit (Arraystar, Rockville, MD, USA) was used for sample labeling. Hybridization was performed in SureHyb Hybridization Chambers (Agilent Technology). After washing, the arrays were scanned using an Agilent DNA Microarray Scanner (Agilent Technology).

Three sets of biological replicates of untreated HeLa and DIA-treated HeLa were prepared. HeLa cells were seeded in 6 well plates at a density of 2.4 × 10⁵ cells/well overnight. After 24 h, 15 μg/mL of DIA was added into the designated wells and was left to incubate for 48 h in a humidified 37°C CO₂ incubator. After the incubation time, the cells were harvested and RNA was extracted using the QiagenRneasy Mini Kit (Qiagen, Germany). The quality of the RNA extracted was measured using the 2100 Bioanalyzer using a RNA Pico chip (Agilent, USA). In order to proceed to microarray, the RIN (RNA Integrity Number) should be more than eight. After all of the samples have passed the minimum requirement for microarray analysis the samples were then used for microarray. All of the samples were subjected to the SurePrint G3 Human Gene Expression 8x60K v2 microarray kit (Agilent Technologies, USA) according to manufacturer protocol, and scanned with Agilent DNA microarray scanner. The results from the microarray study has already been uploaded on the Gene Expression Omnibus with the accession number GSE72974.