G2565ba microarray scanner system

Manufactured by Agilent Technologies

Sourced in Canada

The G2565BA Microarray Scanner System is a high-performance microarray scanner designed for gene expression and genotyping applications. The system provides accurate and reliable data capture for a wide range of microarray platforms.

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12 protocols using g2565ba microarray scanner system

Integrative Glioma Biomarker Analysis

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The mRNA sequencing data, miRNA sequencing data, SNV data and corresponding clinical data used to construct the critical gene network and Cox/Lasso model in malignant glioma were obtained from the TCGA in the National Institutes of Health ‐ National Cancer Institute GDC data portal (https://portal.gdc.cancer.gov/, data release 26.0—8 September 2020). Data included 698 malignant glioma samples (529 GBM samples and 169 LGG samples) and 5 normal brain samples. The sequencing data were derived from the Illumina HiSeq platform (Illumina, Inc.) The research was conducted by the guidelines provided by the TCGA (http://cancergenome.nih.gov/publications/publicationguidelines).
The mRNA sequencing data, miRNA sequencing data and corresponding clinical data for validating the Cox and Lasso models were obtained from the CGGA (http://www.cgga.org.cn/, data release—14 June 2020). Data included 1211 malignant glioma samples (733 LGG samples and 478 GBM samples) and 5 normal brain samples. The mRNA sequencing data were derived from the Illumina HiSeq 2,000/2,500/4,000 Sequencing System, while the miRNA sequencing data were derived from the Agilent G2565BA Microarray Scanner System.

Huang Y., Gao X., Yang E., Yue K., Cao Y., Zhao B., Zhang H., Dai S., Zhang L., Luo P, & Jiang X. (2022). Top‐down stepwise refinement identifies coding and noncoding RNA‐associated epigenetic regulatory maps in malignant glioma. Journal of Cellular and Molecular Medicine, 26(8), 2230-2250.

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Genomic Structural Variant Analysis

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Agilent SurePrint G3 Mouse Microarray 4 × 180 K array technology (Agilent Technology, Inc., Palo Alto, USA) was used to analyse genomic structural variants62 (link). Genomic DNA was isolated from tumour cells by chloroform/phenol extraction followed by ethanol precipitation (Sigma). Test and reference genomic DNAs (500 ng per sample) were fluorescently labelled with Cy5 (test samples) and Cy3 (reference: original cells that inoculated into the mice) with a Genomic DNA Enzymatic Labeling Kit (Agilent Technologies). All array hybridizations were performed according to the manufacturer's methods, immediately scanned with a G2565BA Microarray Scanner System (Agilent), and processed by Feature Extraction Software Ver. 10.7.3.1 (Agilent). All regions of statistically significant copy-number change were determined using Aberration Detection Method-2 (ADM2) algorithms on Agilent Genomic Workbench software version 6.5 Lite software (Agilent Technology)63 (link). The ADM2 algorithms identify genomic regions with copy-number differences between the test and the reference based on log2 ratios of fluorescent signals from probes in the interval. Results were analysed under conditions that fuzzy zero was ON and Moving Average was set at 60 pt.

Takeda K., Nakayama M., Hayakawa Y., Kojima Y., Ikeda H., Imai N., Ogasawara K., Okumura K., Thomas D.M, & Smyth M.J. (2017). IFN-γ is required for cytotoxic T cell-dependent cancer genome immunoediting. Nature Communications, 8, 14607.

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Profiling IDH Mutations and Transcriptome

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Isocitrate dehydrogenase (IDH) mutations were detected by pyrosequencing in the CGGA training cohort (Zhang et al., 2014 (link)) and the molecular profiles (Ceccarelli et al., 2016 (link)) of patients in the validation cohort were collected from the TCGA database. Microarray analysis was performed for 47 patients in the training cohort using the Agilent Whole Human Genome Array (Agilent Technologies Inc., Santa Barbara, CA, USA) in accordance with the manufacturer's protocol (Yan et al., 2012 (link)). The integrity of total RNA was examined using a 2100 Bioanalyzer (Agilent). Biotinylated cRNA and cDNA were synthesized and hybridized to the array. The data were obtained using the Agilent Feature Extraction Software (version 9.1) and the Agilent G2565BA Microarray Scanner System. Probe intensities were normalized using GeneSpring GX 11.0.

Liu X., Li Y., Qian Z., Sun Z., Xu K., Wang K., Liu S., Fan X., Li S., Zhang Z., Jiang T, & Wang Y. (2018). A radiomic signature as a non-invasive predictor of progression-free survival in patients with lower-grade gliomas. NeuroImage : Clinical, 20, 1070-1077.

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miRNA Microarray Analysis Protocol

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Three control samples and three experimental samples were sent to the LC-Bio Company (Beijing, China) for miRNA microarray hybridization and analysis. The miRCURY™ RNA Isolation Kit and the miRCURY™ LNA Array power labeling kit (Exiqon) were used to extract total RNA and label the samples. The samples were subsequently scanned using the Agilent G2565BA Microarray Scanner System (Santa Clara, CA). ImaGene 8.0 software was used for the image analysis (BioDiscovery, Inc., Hawthorne, CA). Normalization was performed using the global Lowess regression algorithm, the original data was shown in Supplementary Table 1.

Zhou Y., Wang Y., Su J., Wu Z., Wang C., Zhong W., Liu X., Cui L., Zhou X., Ma Y., Xin Y., Zhang J., Wu L., Hu X., Chen X., Peng C, & Gao M. (2017). Integration of microRNAome, proteomics and metabolomics to analyze arsenic-induced malignant cell transformation. Oncotarget, 8(53), 90879-90896.

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Integrative Glioma Transcriptome Analysis

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Microarray data from CGGA (http://www.cgga.org.cn/portal.php), GSE16011 (http://www.ncbi.nlm.nih.gov/geo/) and the Rembrandt databases (https://caintegrator.nci.nih.gov/rembrandt/) were gathered from published studies [5] (link), [7] (link), [8] (link). The CEL files for GSE16011 and the Rembrandt data set (Affymetrix GeneChip Human Genome U133 Plus 2.0 Array) were separately merged and computed with Matlab software. The expression data were normalized according to theRobust Multi-array Average (RMA) normalization and expressed in a natural scale. A microarray analysis of CGGA glioma samples was performed with the Agilent Whole Human Genome Array, according to the manufacturer’s instructions. Data were acquired on the Agilent G2565BA Microarray Scanner System, with Agilent Feature Extraction Software (v9.1). Probe intensities were normalized with GeneSpring GX 11.0.

Lin N., Yan W., Gao K., Wang Y., Zhang J, & You Y. (2014). Prevalence and Clinicopathologic Characteristics of the Molecular Subtypes in Malignant Glioma: A Multi-Institutional Analysis of 941 Cases. PLoS ONE, 9(4), e94871.

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Tumor Cell Proportion Analysis and Gene Expression Profiling

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A rapid hematoxylin & eosin stain for frozen sections was applied to each sample to assess the tumor cell proportion before RNA extraction. RNA was extracted only from the samples with > 80% tumor cells. Total RNA was extracted from frozen tumor tissues with the Ambion mirVana miRNA Isolation kit (Cat No. AM1560, Austin, TX) as described previously [31] (link). The ND-1000 spectrophotometer (NanoDrop, Wilmington, DE) was applied to evaluate the quality and concentration of the extracted total RNA, and the Agilent 2100 Bioanalyzer was used to assess RNA integrity. Then, the qualified RNA was collected for further procedures. cDNA and biotinylated cRNA were synthesized and hybridized to the Agilent Whole Human Genome Array according to the manufacturer's instructions. Finally, the array-generated data were analyzed by the Agilent G2565BA Microarray Scanner System and Agilent Feature Extraction software (V9.1). GeneSpring GX11.0 was applied to calculate probe intensity.

Zhao Z., Zhang K.N., Wang Q., Li G., Zeng F., Zhang Y., Wu F., Chai R., Wang Z., Zhang C., Zhang W., Bao Z, & Jiang T. (2021). Chinese Glioma Genome Atlas (CGGA): A Comprehensive Resource with Functional Genomic Data from Chinese Glioma Patients. Genomics, Proteomics & Bioinformatics, 19(1), 1-12.

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Microarray Analysis of Human Genome

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Microarray analysis was performed using the Agilent Whole Human Genome Array according to the manufacturer's instructions [18 (link)]. The integrity of total RNA was checked using an Agilent 2100 Bioanalyzer (Agilent). cDNA and biotiny-lated cRNA were synthesized and hybridized to the array. Data were acquired using the Agilent G2565BA Microarray Scanner System and Agilent Feature Extraction Software (version 9.1). Probe intensities were normalized using GeneSpring GX 11.0. The microarray data was deposited in the CGGA (http://www.cgga.org.cn). The REMBRANDT and GSE16011 microarray data were downloaded from the repository for molecular brain neoplasia data (https://caintegrator.nci.nih.gov/rembrandt/) and the Gene Expression Omnibus (http://www.ncbi.nlm.nih.gov/geo/).

Liu Y., Hu H., Zhang C., Wang H., Zhang W., Wang Z., Li M., Zhang W., Zhou D, & Jiang T. (2015). Co-expression of mitosis-regulating genes contributes to malignant progression and prognosis in oligodendrogliomas. Oncotarget, 6(35), 38257-38269.

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RNA Isolation and Microarray Analysis

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RNA isolation and microarray analysis were performed as previously described [18 (link)]. Briefly, first, total RNA was isolated by MirVana miRNA Isolation kit (Thermo Fisher Scientific, Waltham, USA). Then a NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, USA) was subsequently used to perform the quantification of exacted RNA. Total RNA integrity was checked by an Agilent 2100 Bioanalyzer (Agilent, California, USA), Messenger RNA expression microarray was performed on all samples with the Agilent Whole Human Genome Array (Agilent, California, USA). Data acquisition was carried out by the Agilent G2565BA Microarray Scanner System and Agilent Feature Extraction Software (version 9.1). The normalization of probe intensities was performed using GeneSpring GX 11.0. Normalized gene expression value were log-transformed for analysis.

Wu Z., Liang J., Wang Z., Li A., Fan X, & Jiang T. (2020). HLA-E expression in diffuse glioma: relationship with clinicopathological features and patient survival. BMC Neurology, 20, 59.

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Profiling Transcriptome Changes in Hybrid Cells

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Total RNA from ES cells, cancer cells, hybrid cells and differentiated hybrid cells (D7 and D14) were labeled with Cy5. Samples were hybridized to a Mouse Oligo Microarray (G4121B; Agilent, Shanghai, China) according to the manufacturer's protocol. Arrays were scanned with a G2565BA Microarray Scanner System (Agilent). Data were analyzed using GeneSpring GX software (Agilent). Microarray data have been deposited in the Gene Expression Omnibus database (GSE30965).

Yao J., Zhang L., Hu L., Guo B., Hu X., Borjigin U., Wei Z., Chen Y., Lv M., Lau J.T., Wang X., Li G, & Hu Y.P. (2016). Tumorigenic potential is restored during differentiation in fusion-reprogrammed cancer cells. Cell Death & Disease, 7(7), e2314-.

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miRNA Expression Profiling by Microarray

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We conducted three independent experiments using an Agilent Human 8×60 K miRNA array with the two pooled samples. The integrity of the total RNA was checked using an Agilent 2100 Bioanalyzer (Agilent, Santa Clara, USA). We synthesized the cDNA and biotinylated cRNA and hybridized to the array. Data were acquired using the Agilent G2565BA Microarray Scanner System and Agilent Feature Extraction Software. Probe intensities were normalized using Percentile Shift implemented in GeneSpring 12. Accession number(GSE45737)was obtained after submitting the dataset to Gene Expression Omnibus. Differentially expressed miRNAs were identified through fold- change filtering (fold change>2).

Dai D., Lu Q., Huang Q., Yang P., Hong B., Xu Y., Zhao W., Liu J, & Li Q. (2014). Serum miRNA Signature in Moyamoya Disease. PLoS ONE, 9(8), e102382.

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