Kingfisher duo prime magnetic particle processor

Manufactured by Thermo Fisher Scientific

Sourced in United States

The KingFisher™ Duo Prime Magnetic Particle Processor is a laboratory instrument designed for automated sample preparation using magnetic particles. The core function of this product is to perform rapid and reproducible extraction, purification, and transfer of various biomolecules, such as nucleic acids, proteins, and cells, from a wide range of sample types.

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KingFisher Duo Prime (30 citations) KingFisher Duo (24 citations) KingFisher magnetic particle processor (23 citations)

5 protocols using kingfisher duo prime magnetic particle processor

Plasma miRNA Isolation and Quantification

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Total RNA was isolated from 100 µL plasma with MagMAX™ mirVana™ Total RNA Isolation Kit (ThermoFisher Scientific, Waltham, MA, USA) using KingFisher™ Duo Prime Magnetic Particle Processor (ThermoFisher Scientific, Waltham, MA, USA) according to the user guidelines. Spike-in miRNA C. elegans 39 was added during the RNA purification at a concentration of 15 fmol per sample. Total RNA was eluted with elution buffer in final volumes of 50 μL, and samples were stored at −20 °C until further analysis.
After RNA purification, miRNA was transcribed into cDNA using the TaqMan™ Advanced miRNA cDNA Synthesis Kit (ThermoFisher). A 1:10 dilution of the cDNA was taken for the analysis of 48 miRNAs (C. elegans spike-in control, Supplementary Table S3) using prespotted Taqman adv. miRNA 96 well plates (ThermoFisher) on a QuantStudio 3 Real-Time PCR System (ThermoFisher) in a final reaction volume of 10µL. For data normalization global mean of all analyzed miRNAs were used as previously described [24 (link)].

Tomeva E., Switzeny O.J., Heitzinger C., Hippe B, & Haslberger A.G. (2022). Comprehensive Approach to Distinguish Patients with Solid Tumors from Healthy Controls by Combining Androgen Receptor Mutation p.H875Y with Cell-Free DNA Methylation and Circulating miRNAs. Cancers, 14(2), 462.

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Plasma Cell-Free DNA Isolation

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Cell-free DNA (cfDNA) was isolated with MagMAX™ Cell-Free DNA Isolation Kit (ThermoFisher Scientific, Waltham, MA, USA) using KingFisher™ Duo Prime Magnetic Particle Processor (ThermoFisher Scientific, Waltham, MA, USA) according to the user guidelines. CfDNA was isolated from 4 mL plasma and eluted with elution solution in a final volume of 80 µL. The purified cfDNA samples were stored at −20 °C until further analysis. DNA was quantified using the dsDNA HS Assay Kit on Qubit 4 Fluorometer (Invitrogen, ThermoFisher Scientific, Waltham, MA, USA) according to the standard kit protocol.

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Linked-read Sequencing for N. phaeopus Genome

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We obtained a sample of N. phaeopus (ZMUC 112728) from the Natural History Museum of Denmark, Copenhagen, for reference genome assembly. Its genomic DNA was extracted using the KingFisher Duo Prime Magnetic Particle Processor (Thermo Fisher Scientific, USA) and the KingFisher Cell and Tissue DNA Kit (Thermo Fisher Scientific). A linked-read sequencing library was prepared using the Chromium Genome library kits (10 X Genomics) and sequenced on one Illumina Hiseq X lane at SciLifeLab Stockholm (Sweden). The de novo assembly analysis was performed using 10 X Chromium Supernova (v. 2.1.1). Reads were filtered for low quality and duplication, while assemblies were checked for accuracy and coverage and the best assembly was selected based on the highest genome coverage with the fewest errors. The final genome had a size of 1.12 Gb at a coverage of 50 X with N50=3504.2 kbp.

Tan H.Z., Jansen J.J., Allport G.A., Garg K.M., Chattopadhyay B., Irestedt M., Pang S.E., Chilton G., Gwee C.Y, & Rheindt F.E. (2023). Megafaunal extinctions, not climate change, may explain Holocene genetic diversity declines in Numenius shorebirds. eLife, 12, e85422.

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Plasma cfDNA Isolation and Quantification

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Peripheral venous blood samples (n = 115) were collected for patients at three time points: two days before surgery (pre-op), 10 days after surgery (post-op), and at the end day of the last chemotherapy (post-chemo). At least 20 mL of blood was taken into EDTA-containing tubes. Plasma was separated within 4 h after sample collection. Obtained plasma was centrifuged at 2000 g for 5 min and at 16,000 g for 10 min, immediately aliquoted, and stored at −80 °C. cfDNA was isolated from 4 mL of plasma using a MagMAX Cell-Free DNA Isolation Kit (Applied Biosystems, Foster City, CA, USA) with a KingFisher Duo Prime Magnetic Particle Processor (Thermo Fisher Scientific, Waltham, MA, USA), according to each manufacturer’s instructions. The concentration of the purified plasma cfDNA was measured using a Qubit 2.0 fluorometer (Thermo Fisher Scientific, Waltham, MA, USA) in combination with a Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific, Waltham, MA, USA) according to the manufacturer’s instructions.

Lee C.S., Kim H.S., Schageman J., Lee I.K., Kim M, & Kim Y. (2021). Postoperative Circulating Tumor DNA Can Predict High Risk Patients with Colorectal Cancer Based on Next-Generation Sequencing. Cancers, 13(16), 4190.

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Quantifying mRNA expression in breast cancer cells

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Total RNA was isolated from MCF-7:WS8 and MCF-7:5C cells using MagMAX-96 Total RNA Isolation Kit (Applied Biosystems), and processed using Kingfisher Duo Prime magnetic particle processor (Thermo Scientific). The cDNA was synthesized using High Capacity cDNA Reverse transcription kit (Applied Bioscience). Quantitative real-time PCR assays were performed using Power SYBR Green PCR Master Mix (Applied Biosystems) and a QuantStudio 6 Flex real-time PCR System (Applied Biosystems). All primers were synthesized by Integrated DNA Technologies Inc. All data were normalized using reference gene 36B4.

Abderrahman B., Maximov P.Y., Curpan R.F., Fanning S.W., Hanspal J.S., Fan P., Foulds C.E., Chen Y., Malovannaya A., Jain A., Xiong R., Greene G.L., Tonetti D.A., Thatcher G.R, & Jordan V.C. (2020). Rapid Induction of the Unfolded Protein Response and Apoptosis by Estrogen Mimic TTC-352 for the Treatment of Endocrine-Resistant Breast Cancer. Molecular cancer therapeutics, 20(1), 11-25.

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