Ncounter flex instrument

Manufactured by NanoString

The NCounter FLEX instrument is a versatile tool designed for high-throughput gene expression analysis. It utilizes a unique digital counting technology to precisely measure the expression levels of multiple target genes simultaneously. The instrument's core function is to provide accurate and reliable data on the abundance of specific RNA molecules within a sample, enabling researchers to gain valuable insights into gene expression patterns.

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Lab products found in correlation

Nanodrop one, by Thermo Fisher Scientific (2 mentions) Infinite 200 pro nanoquant reader, by Tecan (1 mentions) Qubit system, by Thermo Fisher Scientific (1 mentions) Nsolver 3, by NanoString (1 mentions) Tapestation 4200, by Agilent Technologies (1 mentions) Ncounter digital analyzer, by NanoString (1 mentions) Agilent 2100 bioanalyzer, by Agilent Technologies (1 mentions) Ncounter human immunology v2 panel, by NanoString (1 mentions) Rnaeasy plus micro kit, by Qiagen (1 mentions) Rlt lysis buffer, by Qiagen (1 mentions) Ionomycin, by Merck Group (1 mentions) Phorbol myristate acetate (pma), by Merck Group (1 mentions)

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NCounter instrument (7 citations) NCounter Flex (5 citations)

5 protocols using ncounter flex instrument

PAXgene Blood RNA Extraction and Analysis

Cited 1 time

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PAXgene blood RNA tubes were shipped to Inflammatix, Inc. (Burlingame, CA), under a sponsored research agreement where RNA was extracted using a protocol previously described (29 (link)), and 29 host mRNAs were quantified using the nCounter FLEX instrument (Nanostring, Seattle, WA).

Ram-Mohan N., Rogers A.J., Blish C.A., Nadeau K.C., Zudock E.J., Kim D., Quinn J.V., Sun L., Liesenfeld O, & Yang S. (2022). Using a 29-mRNA Host Response Classifier To Detect Bacterial Coinfections and Predict Outcomes in COVID-19 Patients Presenting to the Emergency Department. Microbiology Spectrum, 10(6), e02305-22.

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RNA Extraction and Expression Profiling

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Total RNA was extracted and purified using the High Pure formalin-fixed paraffin-embedded tissue RNA Isolation Kit (Roche, Indianapolis, IN) according to the manufacturer protocol. RNA was quantified and its quality was assessed using the Infinite 200 PRO NanoQuant reader (Tecan, Mannedorf, Switzerland) according to the manufacturer protocol.
Expression profiling of RNA samples (Table S2) was performed on the nCounter FLEX Instrument using the NanoPCIP panel of 770 genes (NanoString Technologies, Seattle, WA). Briefly, purified RNA was quantitated using the Qubit System (Life Technologies, Carlsbad, CA) and quality-checked using the NanoDrop One (Thermo Scientific, Waltham, MA) and TapeStation 4200 (Agilent, Santa Clara, CA). Fifty nanograms of RNA was hybridized to gene-specific, fluorescent-labeled probes that were then purified on the nCounter Prep Station. The fluorescent-labeled products were then scanned on the nCounter Digital Analyzer.
Gene expression data were analyzed by nSolver 3.0 software (NanoString Technologies), providing differential expression of all 770 genes in the NanoPCIP panel and immune cell profiling. Using expression of genes previously shown to be characteristic of various immune cell subsets, we estimated the abundance of 17 different immune cell subsets.

Hailemichael Y., Johnson D.H., Abdel-Wahab N., Foo W.C., Bentebibel S.E., Daher M., Haymaker C., Wani K., Saberian C., Ogata D., Kim S.T., Nurieva R., Lazar A.J., Abu-Sbeih H., Fa-ak F., Matthew A., Wang Y., Falohun A., Trinh V., Zobniw C., Spillson C., Burks J.K., Awiwi M., Elsayes K., Soto L.S., Melendez B.D., Davies M., Wargo J., Curry J., Yee C., Lizee G.A., Singh S., Sharma P., Allison J.P., Hwu P., Ekmekcioglu S, & Diab A. (2022). Interleukin-6 blockade abrogates immunotherapy toxicity and promotes tumor immunity. Cancer cell, 40(5), 509-523.e6.

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NanoString Profiling of Interferon Pathway Genes

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An RNeasy FF Macro kit (Qiagen, Hilden, Germany) was used to extract RNA from the cell pellets following the manufacturer’s instructions. RNA quality and quantity were examined using the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). RNA concentration was corrected to include the percentage of the sample that was >300 base pairs. Because interferon pathway-related genes are included in the PanCancer Immune profile panel of NanoString (Nanostring Technologies, Seattle, WA, USA) the panel was used to profile the samples. A total of 200 ng of RNA samples were hybridized with the panel probes at 65 °C for 17 h. RNA expression levels were measured using the nCounter^® FLEX Instrument (Nanostring Technologies). The counting of the genes was performed by scanning 490 fields of view (FOV).

Mustafa D.A., Saida L., Latifi D., Wismans L.V., de Koning W., Zeneyedpour L., Luider T.M., van den Hoogen B, & van Eijck C.H. (2021). Rintatolimod Induces Antiviral Activities in Human Pancreatic Cancer Cells: Opening for an Anti-COVID-19 Opportunity in Cancer Patients?. Cancers, 13(12), 2896.

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NanoString Profiling of Immune Cells

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Gene expression profiling was performed by NanoString, using the pre-designed nCounter Human Immunology v2 Panel (NanoString Technologies). The four assessed immune cell subsets were flow sorted as described above, and 25,000 cells were collected into RLT lysis buffer (Qiagen) either: (i) directly ex vivo; or (ii) following in vitro stimulation for 165 min in the presence or absence of 50 ng/ml PMA (Sigma-Aldrich) and 500 ng/ml ionomycin (Sigma-Aldrich), without addition of protein transport inhibitors. RNA from the flow-sorted T cell subsets was extracted using the RNAeasy Micro Plus kit (Qiagen), with gDNA cleanup, following manufacturer's instructions. Total RNA samples were then hybridised to the NanoString CodeSets, following manufacturer's instructions. Expression levels were assessed using an nCounter Flex instrument (NanoString Technologies). Data were processed using the nSolver Analysis Software following normalisation of the raw read counts to the geometric mean of positive control spike-ins, and the gene expression of 15 selected housekeeping genes (ATG10, C14orf166, CD3E, CD46, G6PD, GPI, POLR1B, POLR2A, PSMB5, PSMB10, PTPRC, SDHA, SKI, TOLLIP and TUBB) that were found to have low variability on both the samples collected ex vivo and following in vitro stimulation.

Ferreira R.C., Rainbow D.B., Rubio García A., Pekalski M.L., Porter L., Oliveira J.J., Waldron-Lynch F., Wicker L.S, & Todd J.A. (2017). Human IL-6RhiTIGIT− CD4+ CD127lowCD25+ T cells display potent in vitro suppressive capacity and a distinct Th17 profile. Clinical Immunology (Orlando, Fla.), 179, 25-39.

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PAXgene Blood RNA Extraction and Analysis

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