Sw41 tube

Manufactured by Beckman Coulter

Sourced in United States

The SW41 tube is a laboratory centrifuge tube designed for use in ultracentrifuge applications. It is a conical-shaped tube with a capacity of approximately 13.5 milliliters. The SW41 tube is constructed from polycarbonate material and is intended for single-use.

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Lab products found in correlation

Nycodenz, by Beckman Coulter (2 mentions) Optiprep, by Merck Group (2 mentions) Optiprep, by Beckman Coulter (2 mentions) Sw41ti rotor, by Beckman Coulter (2 mentions) Carboxyfluorescein succinimidyl ester (cfse), by Beckman Coulter (1 mentions) Carboxyfluorescein succinimidyl ester (cfse), by Thermo Fisher Scientific (1 mentions) Sw28 tube, by Beckman Coulter (1 mentions) 410 electron microscope, by Philips (1 mentions) Sw41 rotor, by Beckman Coulter (1 mentions) Dulbecco phosphate buffered saline (dpbs), by Thermo Fisher Scientific (1 mentions) Gradient master ip, by BioComp Instruments (1 mentions) Optima l 80 xp, by Beckman Coulter (1 mentions) Complete mini, by Roche (1 mentions)

Spelling variants of this product

SW41 ultracentrifuge tube (10 citations)

14 protocols using sw41 tube

Purification and Labeling of Extracellular Vesicles from Babesia Parasites

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EVs were purified from culture supernatant of uRBCs or iRBCs from both B. divergens and B. microti by sequential centrifugations at 1600 × g, 3600 × g, and 12,000 × g each for 15 min each at 4°C. The supernatant was then filtered with 0.2-μm filter to remove any cellular debris and then spun at 100,000 × g for 2 hr at 4°C in Beckman Coulter SW28 tube to get a pellet enriched in EVs. This was resuspended in serum-free RPMI, layered on top of 8 ml 60% sucrose, and centrifuged for 16 h at 4°C in a Beckman SW41 tube at 100 000 × g. Purified EVs were collected from interface (500 ml) and washed with 10 ml phosphate-buffered saline (PBS) by spinning at 100,000 × g for 1.5 h at 4°C in a Beckman Coulter SW41 tube.
For carboxyfluorescein succinimidyl ester (CFSE) staining of EVs, 500 μl CFSE (V12883 Invitrogen) was added at a final concentration of 10 μM. The labeling was done for 15 min at 37°C, following which 30% bovine serum albumin (BSA) stock (Sigma-Aldrich, St. Louis, MO) was added at a final concentration of 1% BSA to stop the labeling. CFSE-labeled EV pellet was layered on 60% sucrose, spun for 16 h in a Beckman SW41 tube at 100 000 × g. EVs collected from interface were washed with 8 ml PBS by spinning 1.5 h in a Beckman SW41 tube at 100 000 × g.

Beri D., Rodriguez M., Singh M., Liu Y., Rasquinha G., An X., Yazdanbakhsh K, & Lobo C.A. (2022). Identification and characterization of extracellular vesicles from red cells infected with Babesia divergens and Babesia microti. Frontiers in Cellular and Infection Microbiology, 12, 962944.

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Subcellular Fractionation of Cell Lysates

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Subcellular fractionation was performed as described previously. In brief, cells were cultured in the presence or absence of 50 μg mL^–1 of β-cycloheximide for 5 h and then scraped into media with the use of a disposable cell scraper. Cells were spun down at 4 °C and 500× g for 5 min and washed sequentially with cold PBS and ice-cold homogenization buffer (25 mM Tris/Cl pH7.5 and 130 mM KCl). Cell pellets were resuspended in homogenization buffer containing protease inhibitors and homogenized by passing through a 25-gauge needle 20 times on ice. Lysates were cleared by a centrifugation at 4 °C and 1000× g for 5 min. Post-nuclear supernatants were overlaid on a discontinuous Nycodenz gradient pre-made in an SW41 tube (Beckman Coulter, Indianapolis, IN, USA) as 0.66 mL of 40% and 5 mL each of 25% and 5% Nycodenz (Axis-Shield/Alere Technologies AS, Oslo, Norway). After gradients were centrifuged at 4 °C and 30,000 rpm for 1 h, 13 fractions at 1 mL each, were collected from top to bottom of each gradient. Equal volumes of each fraction were used for precipitating proteins with 3 volumes of methanol/chloroform (2:1, v/v). After being dried at 37 °C for 1 h, precipitated proteins in each fraction were used for Western blot.

Ding Y., Li Y., Chhetri G., Peng X., Wu J., Wang Z., Zhao B., Zhao W, & Li X. (2021). Parkinson’s Disease Causative Mutation in Vps35 Disturbs Tetherin Trafficking to Cell Surfaces and Facilitates Virus Spread. Cells, 10(4), 746.

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Exosome Purification via Sucrose Gradient

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Sucrose density gradients were performed to purify exosomes. Exosomes
were resuspended in 2 ml of HEPES/sucrose stock solution (2.5M sucrose, 20mM
HEPES/NaOH solution, pH 7.4). The exosomes suspension was overlaid with a linear
sucrose gradient (2.0-0.25M sucrose, 20mM HEPES/NaOH, pH 7.4) in a SW41 tube
(Beckman). The gradients were ultracentrifuged for 16 hrs at 210,000g at
4°C. Gradient fractions of 1 ml were collected from top to bottom and
densities of each fractions were evaluated using a refractometer. Next, the
exosomes pellets were washed in 1X PBS followed by a second step of
ultracentrifugation at 150,000g at 4°C for 2 hrs. Exosomes pellets were
resuspended in Laemmli buffer and/or PBS for further immunoblot and flow
cytometry analysis.

Melo S.A., Luecke L.B., Kahlert C., Fernandez A.F., Gammon S.T., Kaye J., LeBleu V.S., Mittendorf E.A., Weitz J., Rahbari N., Reissfelder C., Pilarsky C., Fraga M.F., Piwnica-Worms D, & Kalluri R. (2015). Glypican1 identifies cancer exosomes and facilitates early detection of cancer. Nature, 523(7559), 177-182.

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Sucrose Gradient Fractionation of Plasma Membranes

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δ-OR-G_i1α (C³⁵¹I)-HEK293 cells were washed in PBS, harvested from totally 60 flasks (80 cm²) by centrifugation for 10 min at 300 × g (0–4°C). Homogenization was performed in TME buffer containing 1 mM fresh PMSF plus Complete protease inhibitors cocktail in Teflon–glass homogenizer for 7 min at 1360 rpm on ice. The cell homogenate was divided into four identical portions having exactly the same volume. The first portion was diluted with 1/10 volume of water (no detergent). The second, third and fourth portion was diluted with 1/10 volume of water plus 10% w/v Brij-58 to a final concentration of 0.025, 0.05 and 0.1% w/v Brij-58, respectively. Exactly 2 ml volumes of these four samples were transferred into Beckman SW41 tube, mixed with 2 ml of 80% w/v sucrose, overlaid with 35%, 30%, 25%, 20% and 15% w/v sucrose (1.5 ml each) and centrifuged for 24 hours at 187,000 × g (0–4°C). Centrifugation resulted in the separation of two clearly visible layers. The lower, optically dense layer represented the plasma membrane fraction (PM). The low-density membranes (LDM) were visible as a hazy area in the upper part of sucrose gradient. Sucrose fractions 1–12 (1 ml) were collected from top to bottom of the centrifuge tube, snap frozen in liquid nitrogen and stored at -80°C.

Roubalova L., Vosahlikova M., Brejchova J., Sykora J., Rudajev V, & Svoboda P. (2015). High Efficacy but Low Potency of δ-Opioid Receptor-G Protein Coupling in Brij-58-Treated, Low-Density Plasma Membrane Fragments. PLoS ONE, 10(8), e0135664.

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Purification and Visualization of Extracellular Vesicles

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Sucrose-purified EVs or iRBCs were washed with 1× PBS and resuspended in fixative with 1% paraformaldehyde and 0.1% glutaraldehyde in 0.1 M cacodylate buffer for 1 h at 4°C, washed in 0.1 M buffer (pH 7.4). They were then treated with 50 mM ammonium chloride to quench the remaining aldehydes and spun for 1.5 h in a Beckman SW41 tube at 100,000 × g and resuspended in 50 ml PBS. Negative staining of purified vesicles from a sucrose gradient interface was performed by using uranyl acetate (1%) in water. After sections were stained with uranyl acetate, they were observed using a Philips 410 electron microscope (Holland).

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Subcellular Fractionation of Membrane Proteins

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Cells were washed twice with buffer (0.25 M sucrose in 20 mM Hepes-NaOH, pH 7.4), homogenized three times at 4°C in the same buffer using a tight-fitting Dounce pestle, and debris removed by centrifugation at 3,310 g for 10 min. The supernatant was placed on a 2-M sucrose cushion and ultracentrifuged at 200,000 g for 1 h. The pellet was resuspended and sucrose concentration was adjusted to 1.5 M. A discontinuous sucrose gradient was generated in an SW41 tube (Beckman Coulter) by overlaying the following sucrose solutions all in buffer consisting of 150 mM NaCl and 20 mM Hepes-NaOH, pH 7.4: 1.5 ml of 2 M, 2 ml of 1.7 M, 3 ml supernatant, 2 ml of 1.25 M, 2 ml of 0.9 M, and 1.5 ml of 0.5 M. The gradient was centrifuged at 110,000 g in an SW41 rotor (Beckman Coulter) for 2 h. Fractions were collected from the top. Equal aliquots of fractions were subjected to SDS-PAGE and proteins probed by Western blotting. Organelles in each fraction were identified by determining the PM marker Na/K ATPase 1, ER marker ribophorin I, and Golgi marker syntaxin 6. The His-Flag–tagged PGAP6 was determined by anti-PGAP6 and anti-Flag antibodies.

Lee G.H., Fujita M., Takaoka K., Murakami Y., Fujihara Y., Kanzawa N., Murakami K.I., Kajikawa E., Takada Y., Saito K., Ikawa M., Hamada H., Maeda Y, & Kinoshita T. (2016). A GPI processing phospholipase A2, PGAP6, modulates Nodal signaling in embryos by shedding CRIPTO. The Journal of Cell Biology, 215(5), 705-718.

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Subcellular Distribution of XK-EGFP Reporter

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To examine whether the XK-EGFP reporter mimicked endogenous XK in subcellular distribution, STHdhQ7/Q7 cells in 10-cm dishes were transfected with pcDNA3.1-XK-EGFP for 16 h and cultured further in the presence or absence of 50 μg/ml of β-cycloheximide for 5 h. Cells were then scraped into culture media and collected by centrifugation at 4°C 1,000 rpm for 5 min. After sequential washes with cold PBS and cold homogenization buffer (25 mM Tris/Cl pH 7.5 and 130 mM KCl), cells were resuspended in homogenization buffer containing protease inhibitors and homogenized by passing through a 25-gauge needle 20 times on ice. Lysates were cleared by centrifugation at 4°C and 2,000 rpm for 5 min. Postnuclear supernatants were overlaid on a discontinuous Nycodenz gradient premade in an SW41 tube (Beckman Coulter) as 0.66 ml of 40% and 5 ml each of 25 and 5% Nycodenz (Axis-Shield/Alere Technologies AS) and centrifuged at 4°C and 30,000 rpm for 1 h. Twelve fractions of 1 ml each were collected from top to bottom of each gradient. Equal volumes of each fraction were used for precipitating the proteins with three volumes of methanol/chloroform (2:1, v/v). After being dried at 37°C for 1 h, precipitated proteins in each fraction were resuspended in 1XSDS sample buffer and analyzed by SDS-PAGE and Western blot.

Chhetri G., Ke Y., Wang P., Usman M., Li Y., Sapp E., Wang J., Ghosh A., Islam M.A., Wang X., Boudi A., DiFiglia M, & Li X. (2022). Impaired XK recycling for importing manganese underlies striatal vulnerability in Huntington's disease. The Journal of Cell Biology, 221(10), e202112073.

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Isolating RNA Granules from Cell Lysate

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Immediately after lysis, K(OAc) in the lysate was adjusted to 150 mM and the nuclei and mitochondria were removed from the lysate by centrifugation at 2500 × g/4 o C for 2 min. The resulting post-mitochondrial supernatant was further clarified by centrifugation at 11 000 × g/4 o C for 9 min and the resulting supernatant (5 -6 mL) loaded on a 2 mL 60 % sucrose cushion in RNA Granule Buffer (20 mM Hepes-KOH, pH 7.5, 150 mM K(OAc), 4 mM Mg(OAc) 2 , 0.5 mM TCEP) in a Beckman SW41 tube. The tube was filled to top with 2 -3 mL ice-cold RNA Granule Buffer and centrifuged for 2 h in a Beckman SW41 Ti swing-out rotor at 38 000 rpm (= 246942 × g; ω 2 t =1.139 × 10 11 rad 2 /s). The supernatant liquid on top of the 60 % sucrose cushion was carefully removed with a pipet and the tube walls and the top of the cushion were washed with 1 -2 mL ice-cold RNA Granule Buffer. After removing the sucrose cushion, the pellet (Pellet 1) was redissolved in ca. 100 μL RNA Granule Buffer and centrifuged through a 700 μL bed-volume Sephacryl-S500 (Cytiva) spin column (equilibrated with RNA Granule Buffer) in a fixed-angle rotor at 1000 × g/4 o C for 1 min to obtain the RNA Granule fraction. The RNA granule preparation was aliquoted in 1/10 th original volume in "ProtLoBind" microcentrifuge tubes (Eppendorf), flash-frozen in liquid nitrogen and stored at -80 C.

Kipper K., Mansour A, & Pulk A. (2022). Neuronal RNA granules are ribosome complexes stalled at the pre-translocation state. Journal of molecular biology, 434(20).

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Polysome Profiling Protocol with Modifications

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Polysome profiling was performed as described in Savant-Bhonsale and Cleveland, 1992 (link), with modifications. The day before harvesting the cells, continuous 15–45% (w/v) sucrose gradients were prepared in SW41 tubes (Beckman) in polysome gradient buffer (10 mM HEPES-KOH, pH 7.6; 100 mM KCl; 5 mM MgCl₂) employing the Gradient Master ip (Biocomp) and stored overnight at 4°C. All steps of the protein extraction were performed on ice. Exponentially growing cells were washed twice with ice-cold DPBS (Gibco) supplemented with 100 µg/ml f. c. cycloheximide, scraped in 300 µl polysome lysis buffer (10 mM HEPES-KOH, pH 7.6; 100 mM KCl; 5 mM MgCl₂; 0.5% IGEPAL CA-630; 100 µg/ml cycloheximide) supplemented with 0.1 U/µl murine RNase Inhibitor (NEB), and passed through a G25 needle 25 times. Nuclei were pelleted at 16,000 × g for 6 min at 4°C, and the supernatants were carefully layered onto the sucrose gradients. Samples were centrifuged at 35,000 rpm for 2 hr at 4°C (with brakes switched off) using an SW 41 Ti rotor (Beckmann). Twenty fractions of 0.6 ml were collected using a peristaltic pump P1 (Amersham Biosciences), and polysome profiles were generated by optical density measurement at 254 nm using optical unit UV-1 (Amersham Biosciences) and chart recorder Rec 111 (Amersham Biosciences).

Vietor I., Cikes D., Piironen K., Vasakou T., Heimdörfer D., Gstir R., Erlacher M.D., Tancevski I., Eller P., Demetz E., Hess M.W., Kuhn V., Degenhart G., Rozman J., Klingenspor M., Hrabe de Angelis M., Valovka T, & Huber L.A. (2023). The negative adipogenesis regulator Dlk1 is transcriptionally regulated by Ifrd1 (TIS7) and translationally by its orthologue Ifrd2 (SKMc15). eLife, 12, e88350.

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Purification and Characterization of Extracellular Vesicles

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For the proteomics analysis, platelet-free plasma samples were purified on a 10–70% sucrose gradient as previously described [27] (link), with some modifications. Approximately 2 mL of PFP isolated from CDAA- and CSAA-fed mice for 20 weeks were layered on top of the sucrose gradient in SW41 tubes (Beckman Coulter Inc., Brea, CA, USA) and ultracentrifuged at 100,000 g for 18 h at 10°C. Fractions comprised between 1.19 and 1.26 g/mL⁻¹were collected, resuspended in PBS and further ultracentrifuged at 100,000 rpm for 1 h at 10°C to wash out sucrose from EV preparation. For the EV characterization, the same fractions were ultracentrifuged at 20,000 g for 30 min at 10°C to pellet the microparticles. The supernatant was transferred to a new tube and centrifuged at 100,000 g for 1 h at 10°C to pellet exosomes. The resulting purified microparticles and exosomes were resuspended in PBS and processed for western blot, FACS and dynamic light scattering analyses.

Povero D., Eguchi A., Li H., Johnson C.D., Papouchado B.G., Wree A., Messer K, & Feldstein A.E. (2014). Circulating Extracellular Vesicles with Specific Proteome and Liver MicroRNAs Are Potential Biomarkers for Liver Injury in Experimental Fatty Liver Disease. PLoS ONE, 9(12), e113651.

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