Total RNA from whole blood was extracted from PAXgene® Blood RNA Tubes using MagMAX™ for Stabilized Blood Tubes RNA Isolation Kit (Life Technologies, CA, USA) according to the manufacturer’s protocol, including a TURBO™ DNase and protease step. RNA was isolated using MagMAX™ for Stabilized Blood Tubes RNA Isolation Kit, compatible with Tempus™ Blood RNA tubes (Life Technologies, CA, USA) following the manufacturer’s protocol with TURBO™ DNase treatment. All extracted RNA samples were stored at −80 °C. The technical characteristics of each extraction method are summarised in Table S1 .
Magmax for stabilized blood tubes rna isolation kit
Manufactured by Thermo Fisher Scientific
Sourced in United States, Germany
The MagMAX for Stabilized Blood Tubes RNA Isolation Kit is a lab equipment product designed for the isolation and purification of RNA from stabilized blood samples. The kit utilizes magnetic bead technology to efficiently extract and purify RNA from the samples.
Automatically generated - may contain errors
Lab products found in correlation
Tempus blood rna tube, by Thermo Fisher Scientific (10 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (6 mentions)
2100 bioanalyzer, by Agilent Technologies (5 mentions)
Nanodrop, by Thermo Fisher Scientific (5 mentions)
Paxgene blood rna tubes, by Thermo Fisher Scientific (4 mentions)
Agilent rna 6000 nano kit, by Agilent Technologies (3 mentions)
High capacity cdna reverse transcription kit, by Thermo Fisher Scientific (3 mentions)
Nanodrop 2000, by Thermo Fisher Scientific (3 mentions)
2100 bioanalyzer instrument, by Agilent Technologies (2 mentions)
Nanodrop 1000 uv vis spectrophotometer, by Thermo Fisher Scientific (2 mentions)
Quant it picogreen dsdna assay kit, by Thermo Fisher Scientific (2 mentions)
Chemagen kit, by PerkinElmer (2 mentions)
Tempus tube, by Thermo Fisher Scientific (2 mentions)
Agilent rna 6000 pico kit, by Agilent Technologies (2 mentions)
Agilent bioanalyzer, by Agilent Technologies (2 mentions)
Paxgene blood mirna kit, by Preanalytix (2 mentions)
Tempus rna tubes, by Thermo Fisher Scientific (2 mentions)
Truseq stranded mrna kit, by Illumina (2 mentions)
Beadstation 500, by Illumina (2 mentions)
Human ht 12 v4 beadchip arrays, by Illumina (2 mentions)
38 protocols using magmax for stabilized blood tubes rna isolation kit
1
RNA Isolation from Whole Blood
2
RNA Isolation from Stabilized Blood
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Total RNA was isolated from Tempus™ Blood RNA Tubes using the MagMAX™ for Stabilized Blood tubes RNA Isolation Kit (Thermo Fischer Scientific) following the manufacturer protocol. The quality of the total RNA extracted was assessed with the Agilent RNA 6000 Nano Kit (Agilent Technologies) using the 2100 Bioanalyzer Instrument (Agilent Technologies). Samples with an RNA integrity number (RIN) less than 6 were discarded from the subsequent profiling experiments.
3
RNA Extraction from Blood Samples
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Peripheral blood (3 ml) samples from all participants were collected using Tempus Blood RNA Tubes (Applied Biosystems, Foster City, CA, United States), mixed vigorously for 10 s immediately and stored at –80°C until use. Total RNA was isolated using the MagMAX for Stabilized Blood Tubes RNA Isolation Kit (Thermo Fisher Scientific, Waltham, MA, United States) according to the manufacturer’s instructions. The concentration of RNA samples was determined using a NanoDrop ND-2000. The integrity of the RNA samples was assessed using a Bioanalyzer 2100.
4
RNA Isolation from Blood Samples
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Total RNA was isolated from Tempus™ Blood Tubes using the MagMAX™ for Stabilized Blood Tubes RNA Isolation Kit (Thermo Fischer Scientific, USA) following the manufacturer protocol. Small RNAs, were isolated from 300 uL of serum using the miRCURY™ RNA Isolation Kits (Exiqon, Vedbaek Denmark) and quantified with the Qubit microRNA Assay Kit (Thermo Fischer Scientific, USA). The quality of the total RNA and miRNAs extracted was assessed with the Agilent RNA 6000 Nano Kit (agilent Technologies, USA) and Agilent Small RNA kit (Agilent Technologies, USA) respectively, by using the 2100 Bioanalyzer Instrument (Agilent Technologies, USA). Samples with RIN less than 6 were discarded from the subsequent profiling experiments.
5
Blood RNA Isolation and Sequencing
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Total RNA from blood samples was extracted using the MagMAX for Stabilized Blood Tubes RNA Isolation Kit (ThermoFisher Scientific, Vilnius, Lithuania). Quality control of isolated RNA was assessed using the Agilent RNA 6000 Pico Kit (Agilent Technologies, Waldbronn, Germany). Library preparation was performed using TruSeq Stranded Total RNA with Ribo-Zero Globin Kit (Illumina, San Diego, CA). Libraries were sequenced using the Illumina Hiseq 2500 (Illumina, San Diego, CA) at a target depth of about 20 million 51-nucleotide single-end reads per sample.
6
RNA Extraction from Whole Blood and Cell Suspensions
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RNA was purified from whole-blood cultures using MagMax for Stabilized Blood Tubes RNA Isolation Kit (Thermo Fisher Scientific). Manufacturer’s instructions were followed, but the initial homogenization volumes were proportionally adjusted to the sample input volume. RNA from cell suspensions was purified using the RNAqueous-Micro Total RNA Isolation Kit (Thermo Fisher Scientific). RNA integrity was assessed using RNA 6000 Pico or RNA 6000 Nano assay on Agilent 2100 Bioanalyzer (Agilent Technologies). RNA concentration and purity were assessed using the NanoDrop 8000 (Thermo Fisher Scientific). All procedures were performed according to the manufacturer’s instructions.
7
Stabilizing and Isolating RNA from Peripheral Blood
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Peripheral blood (3 mL) from patients and HDs was drawn directly in Tempus Blood RNA Tubes (ThermoFischer Scientific, Germany) to stabilize and isolate total RNA. After overnight shipment at room temperature, samples were frozen and stored at − 80 °C. Total RNA was isolated using the MagMAX™ for Stabilized Blood Tubes RNA Isolation Kit (ThermoFischer Scientific) according to the manufacturer’s instructions. In brief, frozen samples were thaw on ice for 30 min, centrifuged at 4500 g for 10 min at 4 °C, pellets were then treated with Tempus Proteinase and TURBO DNase and finally RNA purification was performed using RNA binding beads and a magnet stand. After removing the supernatant and washing twice with the provided washing buffer, beads were left drying at room temperature and total RNA was finally eluted in 40 μL elution buffer. The protocol allowed the recovery of approximately 3-25 μg total RNA. Quality of RNA was checked by 2% agarose gel electrophoresis (SYBR Safe E-Gel 2%, ThermoFischer Scientific) and RNA yield was determined spectrophotometrically (NanoDrop, Implen, Germany).
8
Genome-wide Expression Profiling from Blood
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Total RNA was extracted from human whole blood collected in Tempus Blood RNA tubes (ThermoFisher) using the MagMAX™ for Stabilized Blood Tubes RNA Isolation Kit (ThermoFisher). Total RNA was subjected to globin mRNA depletion using the Ambion GLOBINclear kit (ThermoFisher). Agilent Bioanalyzer was used to evaluate total RNA integrity and the RNA Integrity Number (RIN) was calculated. All samples had RIN in the range of 4.7–8.8; however, the median value was 7. For genome-wide gene expression profiling, fluorescent-labeled PCR products were prepared according to Illumina Human Whole-Genome Gene Expression DASL Assay Guide. The labeled products were hybridized onto the Illumina HumanHT-12-v4 Expression Bead ChIP for 16 h at 58 °C. These arrays were washed, coated based on the Assay Guide, and scanned using Bead Array Scanner 500GX at BSF Microarray Facility.
9
RNA Extraction and qRT-PCR Analysis
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RNA was extracted using MagMAX™ for Stabilized Blood Tubes RNA Isolation Kit, compatible with PAXgene™ Blood RNA Tubes (ThermoFisher). RNA concentration and purity were determined with the NanoDrop 1000 spectrophotometer (NanoDrop, Wilmington, DE), and RNA quality was assessed with the Fragment Analyzer (Agilent, Santa Clara, CA). cDNA was prepared using the High-Capacity cDNA Reverse Transcription kit (ThermoFisher Scientific) following manufacturer’s recommendations. Quantitative RT-PCR was performed on all samples in triplicate using 5 ng of cDNA per 10-ul reaction in 384-well plates with TaqMan™ assays and TaqMan™ Universal Master Mix II, no UNG on the QuantStudio 12K Flex™ Real-Time PCR System (Applied Biosystems, Foster City, CA) with the following cycling parameters: −50 °C for 2 m, 95 °C for 10 m, 40 cycles of 95 °C for 15 s and 60 °C for 1 m. Following amplification, CT values were determined using default settings of the QuantStudio 12 K Flex Software v1.2.3. TaqMan assays used for this analysis were 18S (Hs99999901_s1), HPRT (Hs02800695_m1), and TNFa (Hs00174128_m1).
10
DNA and RNA Extraction from Whole Blood Samples
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DNA was obtained from buffy coats collected in EDTA tubes at mean age 8.1 years old. Briefly, DNA was extracted using the Chemagen kit (Perkin Elmer), in batches by cohort. DNA concentration was determined in a NanoDrop 1,000 UV-Vis Spectrophotometer (Thermo Fisher Scientific) and with Quant-iT PicoGreen dsDNA Assay Kit (Life Technologies).
RNA was extracted from whole blood samples collected in Tempus tubes (Applied Biosystems) using the MagMAX for Stabilized Blood Tubes RNA Isolation Kit (Thermo Fisher Scientific), in batches by cohort. The quality of RNA was evaluated with a 2100 Bioanalyzer (Agilent) and the concentration with a NanoDrop 1000 UV-Vis Spectrophotometer (Thermo Fisher Scientific). Samples classified as good RNA quality had an RNA Integrity Number (RIN) > 5, a similar RNA integrity pattern at visual inspection, and a concentration >10 ng/µl. Mean values for the RIN, concentration (ng/ul) and Nanodrop 260/230 ratio were: 7.05, 109.07, and 2.15, respectively.
RNA was extracted from whole blood samples collected in Tempus tubes (Applied Biosystems) using the MagMAX for Stabilized Blood Tubes RNA Isolation Kit (Thermo Fisher Scientific), in batches by cohort. The quality of RNA was evaluated with a 2100 Bioanalyzer (Agilent) and the concentration with a NanoDrop 1000 UV-Vis Spectrophotometer (Thermo Fisher Scientific). Samples classified as good RNA quality had an RNA Integrity Number (RIN) > 5, a similar RNA integrity pattern at visual inspection, and a concentration >10 ng/µl. Mean values for the RIN, concentration (ng/ul) and Nanodrop 260/230 ratio were: 7.05, 109.07, and 2.15, respectively.
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