Hybridization oven 640

Manufactured by Thermo Fisher Scientific

Sourced in United States

The Hybridization Oven 640 is a laboratory equipment designed for incubating samples during hybridization processes. It provides a controlled environment with precise temperature and air circulation for optimal sample processing.

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Close spelling variants of this product

GeneChip Hybridization Oven 640 (55 citations) Hybridization Oven (12 citations) Affymetrix Hybridization Oven 640 (3 citations)

34 protocols using hybridization oven 640

Rat Genome 230 2.0 GeneChip analysis

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Total RNA was isolated using Trizol (Sigma) followed by RNeasy Mini Elute Kit (Qiagen, Hilden, Germany) purification. 1 µg of total RNA was used to prepare targets by One-Cycle Target labeling kit (Affymetrix, Santa Clara, CA, USA) following the instructions of the manufacturer. Hybridization cocktails were hybridized onto Rat Genome 230 2.0 GeneChips, containing 31,099 rat gene probes, at 45°C for 17 h (60 Rpm) in a Hybridization Oven 640 (Affymetrix). GeneChips were rinsed and stained in a GeneChip fluidics station 450 using the fluidics protocol “EukGE-WS2v5_450” (Affymetrix). Chips were scanned in a GeneChip scanner 3000 (Affymetrix). For all conditions, three separate experiments were analyzed on separate arrays; that is, a total of 36 arrays were used. Array data is available at Arrayexpress [31 (link)] (https://www.ebi.ac.uk/arrayexpress), accession number E-MTAB-3146.

Prause M., Mayer C.M., Brorsson C., Frederiksen K.S., Billestrup N., Størling J, & Mandrup-Poulsen T. (2016). JNK1 Deficient Insulin-Producing Cells Are Protected against Interleukin-1β-Induced Apoptosis Associated with Abrogated Myc Expression. Journal of Diabetes Research, 2016, 1312705.

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Affymetrix Mouse Gene Expression Profiling

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RNA expression was profiled by the Boston University Microarray and Sequencing Resource using Affymetrix Mouse Gene 2.0 ST microarrays. Samples were processed in two batches of nearly identical size and representation of experimental groups to reduce any batch effect. Biotin labeling was performed using the WT Plus reagent kit (Affymetrix, Santa Clara, CA) according to the manufacturer’s protocol. The labeled, fragmented DNA was hybridized to the Affymetrix Mouse Gene 2.0 ST Array for 18 h in a GeneChip Hybridization oven 640 at 45 °C with rotation (60 rpm). The hybridized samples were washed and stained using an Affymetrix fluidics station 450. After staining, microarrays were immediately scanned using an Affymetrix GeneArray Scanner 3000 7G Plus. Raw and processed microarray data have been deposited in the Gene Expression Omnibus (GEO), Series GSE113962.

Francis I.P., Islam E.A., Gower A.C., Shaik-Dasthagirisaheb Y.B., Gray-Owen S.D, & Wetzler L.M. (2018). Murine host response to Neisseria gonorrhoeae upper genital tract infection reveals a common transcriptional signature, plus distinct inflammatory responses that vary between reproductive cycle phases. BMC Genomics, 19, 627.

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Microarray-based Gene and miRNA Expression Analysis

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For Affymetrix PrimeView Human Gene Expression Chip:
The total RNA was amplified, labeled and purified using the GeneChip 3’IVT Express Kit (Affymetrix, Santa Clara, CA, US) following the manufacturer’s instructions to obtain biotin-labeled cRNA. After hybridization on Human PrimeView Arrays for 16 h at 45°C and 60 rpm in the Hybridization Oven 640 (Affymetrix, Santa Clara, CA, US), slides were washed and stained with a Fluidics Station 450 (Affymetrix, Santa Clara, CA, US). Scanning was performed on a seventh-generation GeneChip Scanner 3000 (Affymetrix, Santa Clara, CA, US). Affymetrix GCOS software was used for image analysis to generate raw intensity data.
For Agilent Human miRNA v19.0 Chip (Catalog ID: G4872A-046064):
Each miRNA v19.0 slide was hybridized with 100 ng Cy3-labeled RNA using the miRNA Complete Labeling and Hyb Kit (Agilent technologies, Santa Clara, CA, US) in the Hybridization Oven (Agilent technologies, Santa Clara, CA, US) at 55°C, 20 rpm for 20 h according to the manufacturer’s instructions. After hybridization, slides were washed with the Gene Expression Wash Buffer Kit (Agilent technologies, Santa Clara, CA, US). Slides were scanned by Agilent Microarray Scanner (Agilent technologies, Santa Clara, CA, US) using the Feature Extraction software 10.7 (Agilent technologies, Santa Clara, CA, US) with default settings.

Hou J., Wang F., Kong P., Yu P.K., Wang H, & Han W. (2015). Gene Profiling Characteristics of Radioadaptive Response in AG01522 Normal Human Fibroblasts. PLoS ONE, 10(4), e0123316.

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miRNA Microarray Data Analysis

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Total RNA from 3 replicates of snc886-3p (hsa-miR-886-3p) mimic and negative controls (NC, CN-001000-01-05) was extracted after 48 h of transfection using the Qiagen^TM miRNAeasy kit according to the manufacturer’s protocol. The total RNA of the three replicates was pooled and labeled according to Affymetrix (Affymetrix, Santa Clara, CA, USA). Hybridization, staining and washing of the Affymetrix^® HG-U133 Plus 2.0 Arrays were performed with the Affymetrix Fluidics Station 450 and Hybridization Oven 640 under standard conditions (Affymetrix, Santa Clara, CA, USA). Quality control analysis and Pre-processing of the CEL-files microarray expression data was done using a graphical user interface, Chipster (v1.4.3, CSC, Finland, http://chipster.csc.fi/) following the manufacturer’s guidelines [88 (link)]. Normalization was performed using RMA algorithm [89 (link)] and annotation using the specific Affymetrix^® HG-U133 Plus 2.0 Arrays probe set library in Chipster. Normalized log2 expression values were used to determine the fold change expression of the snc886-3p (hsa-miR-886-3p) mimic and negative controls (NC, CN-001000-01-05) (Table S1).

Fort R.S., Garat B., Sotelo-Silveira J.R, & Duhagon M.A. (2020). vtRNA2-1/nc886 Produces a Small RNA That Contributes to Its Tumor Suppression Action through the microRNA Pathway in Prostate Cancer. Non-Coding RNA, 6(1), 7.

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Quantification of mRNA Expression in Atrial Fibroblasts

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To determine mRNA expression levels relative to glyceraldehyde 3-phosphate dehydrogenase (GAPDH) by quantitative polymerase chain reaction (qPCR), mRNA isolation from immortalized human atrial fibroblasts (Künzel et al., 2020 (link)) was performed using a commercial RNA isolation kit (RNeasy Micro Kit; Qiagen, Germany). Complementary DNA was amplified in TaqMan Fast Advanced Master Mix (4444556, ThermoFisher) for a total of 40 cycles. A more detailed protocol can be found in Emig et al. (2021) (link).
KCNMA1 expression in primary atrial fibroblasts from patients with SR and AF was assessed based on the Affymetrix GeneChip array (Affymetrix, Santa Clara, CA, United States) analysis described in Poulet et al. (2016) (link). Briefly cells were cultured for 3 weeks, replated at a density of 2.5 × 10³ cells per cm² and kept two more weeks in culture before analysis. Hybridization at 45°C and 60 revolutions per minute for 16 h (hybridization oven 640; Affymetrix), staining and washing, processing (Fluidics Station 450, Affymetrix) and scanning (GeneChip Scanner 3000 G7) were all carried out as recommended by Affymetrix.

Jakob D., Klesen A., Darkow E., Kari F.A., Beyersdorf F., Kohl P., Ravens U, & Peyronnet R. (2021). Heterogeneity and Remodeling of Ion Currents in Cultured Right Atrial Fibroblasts From Patients With Sinus Rhythm or Atrial Fibrillation. Frontiers in Physiology, 12, 673891.

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Transcriptomic Analysis of Vascular Endothelium

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In order to explore the mechanism of the protective effect of the combination of Uncaria and Semen Raphani on vascular endothelium, RT-PCR chip for biological function of endothelium was performed to observe mRNA expression of the thoracic aorta [10 (link)]. The total RNA of the thoracic aorta was isolated using the TRIzol method and then reverse-transcribed into cDNA with the PrimeScript RT reagent kit (RR047A, TaKaRa, Dalian, China). Following quantitative detection, the cDNA was hybridized in an Affymetrix Hybridization Oven 640, with subsequent elution inside a clean workstation (GeneChip Fluidics Station 450, Affymetrix). The chips were then scanned with a Gene Array Scanner 3000 7G (Affymetrix) to trace the detection signal. A GeneSpring GX 7.3.1 was used for the gene expression analysis and transcripts with low expression levels (<25% of the median gene expression value) and frequently missed hybridizations (>2 absent flags among the samples) were not analyzed further. Transcripts that showed a >2-fold expression reduction in tissue from drug-treatment groups relative to SHR groups were extracted. The transcript list contained 84 transcripts. Both the test group and control group included three biological replicates [11 (link)].

Li Y.L., Jiang Y.H., Yang C.H., Sun J.C., Wang M.M, & Yang W.Q. (2015). Enhanced Protective Effect of the Combination of Uncaria and Semen Raphani on Vascular Endothelium in Spontaneously Hypertensive Rats. Evidence-based Complementary and Alternative Medicine : eCAM, 2015, 358352.

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PBMC RNA Extraction and Microarray Analysis

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After PMBC isolation, 1 × 10⁶ PBMCs were immediately lysed in 1 ml of TRIzol (Life Technologies, Carlsbad, CA, USA) and stored at −80°. The samples were thawed and RNA extracted following the manufacturer’s instructions. The quality of the RNA was assessed using a Nanodrop 2000 Spectrometer (Thermo Scientific, MA, USA) as well as by visualization of the integrity of the 28S and 18S bands on an Agilent Bioanalyzer 2100 instrument (Agilent Technologies, Santa Clara, CA, USA).
Qualified total RNA was further purified using an RNeasy micro kit (QIAGEN, GmBH, Germany). Contaminating genomic DNA was removed using an RNase-Free DNase Set (QIAGEN, GmBH, Germany). The purified RNA was stored at −80° C.
To obtain biotin-labeled cRNA, total RNA was amplified, labeled, and purified using the GeneChip 3’IVT Express Kit (Affymetrix, Santa Clara, CA, USA) following the manufacturer’s instructions. After hybridization on Human PrimeView Arrays for 16 h at 45°C and rotation at 60 rpm in a Hybridization Oven 640 (Affymetrix), slides were washed and stained with a Fluidics Station 450 system (Affymetrix). Scanning was performed on a seventh-generation GeneChip Scanner 3000 (Affymetrix). Affymetrix GCOS software was used to perform image analysis and generate raw intensity data.

Hou J., Wang S., Jia M., Li D., Liu Y., Li Z., Zhu H., Xu H., Sun M., Lu L., Zhou Z., Peng H., Zhang Q., Fu S., Liang G., Yao L., Yu X., Carpp L.N., Huang Y., McElrath J., Self S, & Shao Y. (2017). A systems vaccinology approach reveals temporal transcriptomic changes of immune responses to the yellow fever 17D vaccine. Journal of immunology (Baltimore, Md. : 1950), 199(4), 1476-1489.

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RNA Extraction and Microarray Analysis

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RNA was extracted from control or Hnf6/Oc2 double-mutant spinal cords. The tissue was manually dissociated in Tripur isolation reagent (Roche, 11 667 165 001). After dissociation, chloroform (Merck Millipore, 1 02445 1000) was added to the sample, incubated at room temperature for 10 min and centrifugated for 10 min at 4°C. The aqueous phase was collected and the RNA was precipitated with isopropanol (VWR, 20880.310) and centrifugated for 15 min at 4°C. The pellet was washed in ethanol (Biosolve, 06250502) and centrifugated for 10 min at 4°C. The dried pellet was resuspended in RNAse free water. The integrity of the RNA was assessed using an Agilent RNA 6000 Nano assay procedure. For microarray analyzes, the RNA was converted in single-strand cDNA, labeled using the GeneChip^® WT PLUS Reagent Kit (Affymetrix) and hybridized on the GeneChip^® MoGene 2.0 ST array (Affymetrix, 90 2118) using Affymetrix devices: Genechip^® Fluidics Station 450, Genechip^® Hybridization oven 640, Affymetrix Genechip^® scanner and the Expression Consol software. The analyzes were performed using the R software. Microarray data have been deposited in the GEO repository (accession number: GSE117871).

Harris A., Masgutova G., Collin A., Toch M., Hidalgo-Figueroa M., Jacob B., Corcoran L.M., Francius C, & Clotman F. (2019). Onecut Factors and Pou2f2 Regulate the Distribution of V2 Interneurons in the Mouse Developing Spinal Cord. Frontiers in Cellular Neuroscience, 13, 184.

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Affymetrix Gene Expression Analysis

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RNA was extracted using the RNeasy Mini Kit (Qiagen) according to manufacturer’s instructions. Quality of RNA was analyzed using the Bioanalyzer platform (Agilent Technologies). Two micrograms of total RNA were treated according to standard Affymetrix eukaryotic RNA labeling protocols (Affymetrix, Santa Clara, CA). Fifteen micrograms of biotin labeled cRNA was fragmented according to Affymetrix eukaryotic sample protocol. Hybridization, staining, and washing of the Affymetrix HG-U133_Plus_2 Arrays were performed using the Affymetrix Fluidics Station 450 and Hybridization Oven 640 under standard conditions. The raw data were processed using the GCRMA-normalization method [18 (link)].

Ruuth M., Soronen J., Kaiharju E., Merikanto K., Perttilä J., Metso J., Lee-Rueckert M., Taskinen M.R., Kovanen P.T., Öörni K., Olkkonen V.M., Jauhiainen M.S, & Laurila P.P. (2018). USF1 deficiency alleviates inflammation, enhances cholesterol efflux and prevents cholesterol accumulation in macrophages. Lipids in Health and Disease, 17, 285.

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Microarray Analysis of Sorted IL-17+ Cells

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Sorted IL-17+ cell samples were lysed in 500 μl of TRIzol (Invitrogen). Chloroform (0.2 ml) was added to 1 ml TRIzol cell homogenate and whirl mixed for 15 sec. Homogenates were incubated 2-3 min at RT and centrifuged 15 min at 10,000g (4°C). The water phase containing the RNA was further purified using RNeasy MinElute Cleanup Kit (Qiagen). RNA integrity was confirmed on an Agilent 2100 Bioanalyzer using total RNA nano chips (Agilent Technologies, Santa Clara, CA, USA). An amount of 100 ng of total RNA was used to prepare targets by 3′ IVT Express kit (Affymetrix, Santa Clara, CA, USA) following manufacturer’s instructions. Hybridization cocktails were hybridized onto Human Genome U133 Plus 2.0 Gene Chips® (Affymetrix) at 45°C for 17 h (60 rpm) in a Hybridization Oven 640 (Affymetrix). GeneChips® were washed and stained in a GeneChip® fluidics station 450 using the fluidics protocol “EukGE-WS2v5_450” (Affymetrix). Chips were scanned in a GeneChip® scanner 3000 (Affymetrix). Microarray data were normalized and gene expression measures derived using the RMA algorithm and the Bioconductor package “Affy” (http://www.bioconductor.org). Custom CDF (chip definition file) from brainarray.mbni.med.umich.edu was used. Qlucore Omics Explorer 2.2 (Qlucore AB, Sweden) was used for the statistical analysis of the normalized data.

Evans H.G., Roostalu U., Walter G.J., Gullick N.J., Frederiksen K.S., Roberts C.A., Sumner J., Baeten D.L., Gerwien J.G., Cope A.P., Geissmann F., Kirkham B.W, & Taams L.S. (2014). TNF-α blockade induces IL-10 expression in human CD4+ T cells. Nature communications, 5, 3199.

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