3 mm tungsten carbide bead

Manufactured by Qiagen

Sourced in Germany, United States

The 3-mm tungsten carbide bead is a laboratory equipment product used for sample preparation. It is a small, hard sphere made of tungsten carbide, a durable material commonly used in industrial applications. The core function of this bead is to facilitate the efficient homogenization and disruption of various sample types during the sample preparation process.

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Lab products found in correlation

Tissuelyser 2, by Qiagen (13 mentions) Revertaid first strand cdna synthesis kit, by Thermo Fisher Scientific (6 mentions) Pierce biotech bca assay kit, by Thermo Fisher Scientific (3 mentions) High pure viral nucleic acid kit, by Roche (2 mentions) Dneasy blood and tissue kit, by Qiagen (2 mentions) Trizol reagent, by Thermo Fisher Scientific (2 mentions) Phosphate buffered saline (pbs), by Merck Group (2 mentions) Collection microtubes, by Qiagen (2 mentions) Tissuelyser system, by Qiagen (2 mentions) Quick rna miniprep kit, by Zymo Research (2 mentions) Zirconia silica bead, by Biospec (1 mentions) Lightcycler 1.5 detection system, by Roche (1 mentions) Lightcycler faststart dna masterplus sybr green 1 kit, by Roche (1 mentions) M mlv reverse transcriptase kit, by Thermo Fisher Scientific (1 mentions) Nucleospin rna isolation kit, by Macherey-Nagel (1 mentions) Tissuelyser 2 system, by Qiagen (1 mentions) Trizoltm reagent, by Thermo Fisher Scientific (1 mentions) Protein assay dye, by Bio-Rad (1 mentions) T per buffer, by Thermo Fisher Scientific (1 mentions) Edta solution, by Thermo Fisher Scientific (1 mentions)

24 protocols using 3 mm tungsten carbide bead

Comprehensive Plant DNA and RNA Extraction

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For DNA extraction, two leaf discs (d = 12.7 mm) were collected from expanded leaves below the meristem at 12, 20, 28 and 36 dpi. Leaf tissue was flash frozen in liquid nitrogen and homogenised with 3 mm tungsten carbide beads (Qiagen) using the Qiagen TissueLyser II system (Qiagen) at 20 Hz for 1 min twice. CTAB extraction was done in accordance with Doyle [54 (link)] with the following modifications: an additional chloroform-isoamyl alcohol extraction step, two ethanol wash steps (95% first wash, 75% second wash), and incubation of samples at -80°C for 30 min after the addition of isopropanol.
For total RNA extraction, ~100 mg of homogenized leaf material collected at 20 dpi was flash frozen in liquid nitrogen and homogenized with DEPC-treated 3 mm tungsten carbide beads (Qiagen) using the Qiagen TissueLyser II system (Qiagen) at 20 Hz for 1 min twice. RNA was extracted using TRIzol^TM reagent (Thermo Fisher Scientific) as per the manufacturer’s protocol with an additional chloroform separation step. All nucleic acids were quantified using the NanoDrop^TM One spectrophotometer (Thermo Fisher Scientific) and purity was also assessed using gel electrophoresis. Extracted DNA and RNA was stored at -80°C for downstream analysis.

Zwolinski A.M., Brigden A, & Rey M.E. (2023). Differences in the 3’ intergenic region and the V2 protein of two sequence variants of tomato curly stunt virus play an important role in disease pathology in Nicotiana benthamiana. PLOS ONE, 18(5), e0286149.

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Mosquito Pool Homogenization and RNA Extraction

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Mosquito pools were homogenized by shaking with 3 mm tungsten carbide beads (QIAgen, Hilden, Germany) in 500–600 μl of Eagle’s least essential medium, added to 5% fetal bovine plasma 1% penicillin–streptomycin and 1% l-glutamine. They were purified by centrifugation at 4,000 × rpm for 4 min, divided into tubes and preserved at −80 °C. Nucleic acids were clarified from one aliquot of each pool using High Pure Viral Nucleic Acid Kit (Roche Diagnostics, Mannheim, Germany), Then reverse transcription polymerase chain reaction conducted (TaqMan® one-step RT-PCR master mix reagents kit). The One-Step EZ RT-PCR TaqMan® reagent kit (Applied Biosystems, Sweden) reaction performed according to the manufacturers’ guidelines (Jupp et al., 2002 (link)). All collected samples were classified into their species using available morphological keys (The ten mosquito pools were grinded in a varying volume of Virus Transport Media (VTM) depending on the pool size according the method of (Zakhia et al., 2018 ). The supernatants were clarified through a 3.5 µm filter and centrifuged at 3000 cycle/30 min.

Ali El Hadi Mohamed R., Abdelgadir D.M., Bashab H.M., Al-Shuraym L.A., Sfouq Aleanizy F., Alqahtani F.Y., Ahmed Al-Keridis L, & Mohamed N. (2020). First record of West Nile Virus detection inside wild mosquitoes in Khartoum capital of Sudan using PCR. Saudi Journal of Biological Sciences, 27(12), 3359-3364.

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Sardinian Newt Skin Sampling for Chytrid Fungus

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Euproctus platycephalus samples, comprising skin swabs (Müller, Lenhardt, & Theissinger, 2013), toe clips (Arntzen, Smithson, & Oldham, 1999) or larva tail tips (Polich, Searcy, & Shaffer, 2013), were collected between 2005 and 2012 as part of a survey of B. dendrobatidis infection (Bielby et al., 2013; Bovero et al., 2008). DNA was extracted using DNeasy Blood and Tissue kit (Qiagen), with the addition during the tissue lysis step of 0.5‐mm Zirconia/silica beads (BioSpec Products) for swabs and 3‐mm tungsten carbide beads (Qiagen) for toe clips. A total of 227 DNA samples were included in the study, from 13 GPS‐defined sampling sites, spanning the three main mountain ranges of Sardinia to ensure coverage of the geographical range of the species. Results were analyzed for 168 successfully genotyped individuals (detailed in Table 1).

Ball S.E., Bovero S., Sotgiu G., Tessa G., Angelini C., Bielby J., Durrant C., Favelli M., Gazzaniga E, & Garner T.W. (2017). Islands within an island: Population genetic structure of the endemic Sardinian newt, Euproctus platycephalus. Ecology and Evolution, 7(4), 1190-1211.

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Quantitative RT-PCR Analysis of Mouse Brain Regions

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Each sample consisted of the tissue deriving from one mouse brain. Mice were sacrificed by cervical dislocation. The dorsal striatum (DS) and the NAc were quickly dissected out and placed on ice. Each tissue sample was homogenized in TRIzol with 3 mm tungsten carbide beads (Qiagen cat.No. 69997). Total RNA was extracted with TRIzol Reagent (Life Technologies) according to manufacturer’s instructions. The RNA was quantified by using the NanoDrop 1000 spectrophotometer. Between 85 and 500 ng of mRNA from each sample were used for retro-transcription, performed with the Reverse Transcriptase M-MLV (Life Technologies) following the manufacturer’s instructions. Quantitative real time PCRs, were performed in a LightCycler 1.5 detection system (Roche, Meylan France) using the LightCycler FastStart DNA Master plus SYBR Green I kit (Roche) in 384-well plates according to the manufacturer’s instruction. All reactions were carried out in duplicate (Striatum) or triplicate (Hypothalamus). Results are presented as normalized to the house-keeping gene and the delta-CT method was used to obtain a FC.

Berland C., Montalban E., Perrin E., Di Miceli M., Nakamura Y., Martinat M., Sullivan M., Davis X.S., Shenasa M.A., Martin C., Tolu S., Marti F., Caille S., Castel J., Perez S., Salinas C.G., Morel C., Hecksher-Sørensen J., Cador M., Fioramonti X., Tschöp M.H., Layé S., Venance L., Faure P., Hnasko T.S., Small D.M., Gangarossa G, & Luquet S. (2020). Circulating triglycerides gate dopamine-associated behaviours through dopamine receptor type 2 (DRD2)-expressing neurons. Cell metabolism, 31(4), 773-790.e11.

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Mosquito Pool RNA Extraction Protocol

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Mosquito pools were homogenised by vortexing with 3 mm tungsten carbide beads (QIAgen, Hilden, Germany) in 500–600 μl of Eagle’s minimal essential medium, supplemented with 5% fetal bovine serum, 1% penicillin-streptomycin and 1% l-glutamine. They were clarified by centrifugation at 4,000× rpm for 4 min, aliquoted and stored at -80 °C. Nucleic acids were purified from one aliquot of each pool using High Pure Viral Nucleic Acid Kit (Roche Diagnostics, Mannheim, Germany), followed by a reverse transcription reaction, using random hexamers and the RevertAid First Strand cDNA Synthesis Kit (Thermo Fisher Scientific, Tokyo, Japan), performed according to the manufacturers’ guidelines.

Ergünay K., Litzba N., Brinkmann A., Günay F., Sarıkaya Y., Kar S., Örsten S., Öter K., Domingo C., Erisoz Kasap Ö., Özkul A., Mitchell L., Nitsche A., Alten B, & Linton Y.M. (2017). Co-circulation of West Nile virus and distinct insect-specific flaviviruses in Turkey. Parasites & Vectors, 10, 149.

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Quantifying Nosema ceranae Spores

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Bee abdomens were individually placed in 1.5 mL tubes and homogenized in 1 mL of distilled water with 3 mm tungsten carbide beads (Qiagen, Germany) in a TissueLyser II (Qiagen, Germany) for 1 min at 25 Hz. N. ceranae spore was estimated for each bee using a haemocytometer according to Cantwell [74 ] and OIE [75 ].

Glavinic U., Stevanovic J., Ristanic M., Rajkovic M., Davitkov D., Lakic N, & Stanimirovic Z. (2021). Potential of Fumagillin and Agaricus blazei Mushroom Extract to Reduce Nosema ceranae in Honey Bees. Insects, 12(4), 282.

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Fecal Sample Preparation for Analysis

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The faeces were centrifuged to form pellets and the supernatant were discarded. The pellets were incubated at 56 °C for 2 h to evaporate moisture (i.e., ethanol), pooled according to collection week and homogenised using a TissueLyser II (Qiagen, Hilden, Germany) with 3 mm tungsten carbide beads (Qiagen, Hilden, Germany) for 4 min at 30 1/s.

Lim V.C., Ramli R., Bhassu S, & Wilson J.J. (2018). Pollination implications of the diverse diet of tropical nectar-feeding bats roosting in an urban cave. PeerJ, 6, e4572.

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Transcriptional Analysis of Stress Genes

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Plant tissue was disrupted mechanically using 3 mm tungsten carbide beads (Qiagen, Hilden, Germany), and a TissueLyser II (Qiagen) at 30 Hz for 2 min. RNA extraction was performed using NucleoSpin RNA isolation kit according to manufacturer’s protocol (Macherey-Nagel, Düren, Germany). One microgram of total RNA was used for cDNA synthesis using the M-MLV reverse Transcriptase Kit (Invitrogen Ltd, Carlsbad, CA, USA) according to the manufacturer’s protocol. Instead of RNAseOut, the equal amount of RNase free deionized H₂O was used. PCR was carried out on 1 μL of cDNA with gene-specific primers against ERO1 (P7/P8), ERO2 (P9/10), and ACTIN7 (At5g09810; P11/P12).

Ugalde J.M., Aller I., Kudrjasova L., Schmidt R.R., Schlößer M., Homagk M., Fuchs P., Lichtenauer S., Schwarzländer M., Müller-Schüssele S.J, & Meyer A.J. (2022). Endoplasmic reticulum oxidoreductin provides resilience against reductive stress and hypoxic conditions by mediating luminal redox dynamics. The Plant Cell, 34(10), 4007-4027.

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Tungsten-Carbide Bead Milling and CTAB DNA Extraction

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A 200 mg (dry weight) of each sample was transferred to a 2 ml microcentrifuge tube with two 3 mm tungsten-carbide beads (Qiagen) and ground to a fine powder on a Retsch mill MM 301 (Retsch GmbH, Haan Germany) (2X cycles for 45 s at 25 Hz). Total DNA was extracted from the homogenized contents using a modified CTAB extraction method as described by Doyle and Doyle (1990) , and adapted by Raclariu et al. (2017b) (link). The final elution volume was 100 μl.

Raclariu-Manolică A.C., Anmarkrud J.A., Kierczak M., Rafati N., Thorbek B.L., Schrøder-Nielsen A, & de Boer H.J. (2021). DNA Metabarcoding for Quality Control of Basil, Oregano, and Paprika. Frontiers in Plant Science, 12, 665618.

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Measuring Mucosal Inflammation Biomarkers

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Total 6 mm pieces of distal colon or terminal ileum were placed in T-per buffer (Thermo Fisher Scientific) containing 1× Halt Protease Inhibitor Cocktail (Thermo Fisher Scientific), 0.5 M EDTA solution (Thermo Fisher Scientific), and two 3 mm tungsten carbide beads (Qiagen). Tissues were disrupted in TissueLyser II (Qiagen) for 10 min at an oscillation frequency of 27.5 Hz. Generated tissue homogenates were centrifuged at 15000 rcf for 10 min at 4 °C and the supernatants were collected into deep 96-well plates. Protein assay dye (BioRad) was used to quantify total protein content using Bradford protein assay according to the manufacturer’s instructions. Cytokine concentrations were measured using V-PLEX Plus Proinflammatory Panel 1 mouse kit according to the manufacturer’s instructions (Meso Scale Diagnostics). Absorbance was measured on the Meso SECTOR S600 instrument (Meso Scale Diagnostics). Myeloperoxidase (MPO) activity was measured using a mouse MPO ELISA kit according to the manufacturer’s instructions (Hycult Biotech). Absorbance was measured on the SpectraMax i3x instrument (Molecular Devices). Data analysis was performed using GraphPad Prism™ (GraphPad Software, Inc.). Cytokine and MPO levels were normalized to total protein content.

Panea C., Zhang R., VanValkenburgh J., Ni M., Adler C., Wei Y., Ochoa F., Schmahl J., Tang Y., Siao C.J., Poueymirou W., Espert J., Lim W.K., Atwal G.S., Murphy A.J., Sleeman M.A., Hovhannisyan Z, & Haxhinasto S. (2021). Butyrophilin-like 2 regulates site-specific adaptations of intestinal γδ intraepithelial lymphocytes. Communications Biology, 4, 913.

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