Video articles in JoVE about centrifugation density gradient include Characterization and Isolation of Mouse Primary Microglia by Density Gradient Centrifugation, DNA Extraction from 0.22 μM Sterivex Filters and Cesium Chloride Density Gradient Centrifugation, Protocol for Isolation of Primary Human Hepatocytes and Corresponding Major Populations of Non-parenchymal Liver Cells, A New Approach for the Comparative Analysis of Multiprotein Complexes Based on 15N Metabolic Labeling and Quantitative Mass Spectrometry, Isolation of Intact Mitochondria from Skeletal Muscle by Differential Centrifugation for High-resolution Respirometry Measurements, Preparation of Liquid-exfoliated Transition Metal Dichalcogenide Nanosheets with Controlled Size and Thickness: A State of the Art Protocol, Isolation of Cellular Lipid Droplets: Two Purification Techniques Starting from Yeast Cells and Human Placentas, Cloning and Large-Scale Production of High-Capacity Adenoviral Vectors Based on the Human
TY - JOUR. T1 - Hydrocarbon molecular markers as indicators of the Late Cenozoic sedimentation on the Amerasian continental margin (Arctic Ocean). AU - Petrova, V. I.. AU - Batova, G. I.. AU - Kursheva, A. V.. AU - Litvinenko, I. V.. AU - Morgunova, I. P.. PY - 2020/4. Y1 - 2020/4. N2 - The main factors controlling the bulk sedimentation in the region of the Mendeleev Rise and the adjacent part of the Arctic Ocean during the late Cenozoic were studied using a complex of geomorphological, lithological, and organic geochemical data. Samples for the study were collected during the cruises of the R/V Akademik Fedorov in 2000, 2005, and 2007 and icebreaker Kapitan Dranitsyn in 2012. Analysis of the group and molecular compositions of the dispersed organic matter (DOM) in bottom sediments has shown that the input of terrigenous sediments enriched with abrasion products of lithified rocks from the eastern source province determines the Holocene-Pleistocene sedimentation on the continental slope of the ...
An investigation of the distribution of immunoglobulins in the external secretions of the hamster revealed the predominance of γA. Only γA was found in saliva, whereas colostrum and urine both contained γ2 in addition to γA. Hamster γA was present in the serum at concentrations that were insufficient to allow its visualization by routine immunoelectrophoresis. No differences in electrophoretic mobility were found between serum and secretory γA regardless of derivation. The secretory γA from saliva, colostrum and urine had sedimentation characteristics intermediate between serum γA and 19S markers in sucrose density gradient ultracentrifugation studies. Similar results were obtained on Sephadex G-200 gel filtration. Isolated secretory γA had an s020,w of 15.2S. Mild reductive procedures did not appear to alter the sedimentation characteristics of this molecule. No antigenic differences were found between serum, salivary, colostral and urinary γA.. ...
Sigma-Aldrich offers Sigma-D1556, OptiPrep™ Density Gradient Medium for your research needs. Find product specific information including CAS, MSDS, protocols and references.
Matthew Meselson, Franklin Stahl, and Jerome Vinograd, developed cesium chloride, or CsCl, density gradient centrifugation in the 1950s at the California Institute of Technology, or Caltech, in Pasadena, California. Density gradient centrifugation enables scientists to separate substances based on size, shape, and density. Meselson and Stahl invented a specific type of density gradient centrifugation, called isopycnic centrifugation that used a solution of cesium chloride to separate DNA molecules based on density alone.. Format: Articles Subject: Technologies ...
Matthew Meselson, Franklin Stahl, and Jerome Vinograd, developed cesium chloride, or CsCl, density gradient centrifugation in the 1950s at the California Institute of Technology, or Caltech, in Pasadena, California. Density gradient centrifugation enables scientists to separate substances based on size, shape, and density. Meselson and Stahl invented a specific type of density gradient centrifugation, called isopycnic centrifugation that used a solution of cesium chloride to separate DNA molecules based on density alone.. Format: Articles Subject: Technologies ...
AXIS-SHIELD Density Gradient Media - Macromolecule indexes - Isolation of cells, cell organelles, subcellular membranes, viruses and macromolecules using centrifugation techniques
mRNAs surrounded by polysomes are ready for translation into proteins (Warner et al., 1963); these mRNAs are defined as polysomal-mRNAs (Mustroph et al., 2009). The process is affected by various growth conditions or surrounding situations. Microarray analysis is a powerful tool for detecting genome-wide gene expression. Therefore, using polysomal-mRNAs for microarray analysis can reflect the gene translation information (the translatome) under different developmental stages or environmental conditions from eukaryotes. Polysomal-mRNAs can be collected from the polysomal fraction by sucrose-gradient separation for further quantitative PCR or microarray assay. We modified a protocol (Mustroph et al., 2009) for collecting polysomal-mRNAs via sucrose-gradient separation to eliminate monosomal-mRNA contamination from pLAT52:HF:RPL18 Arabidopsis. This transgenic Arabidopsis uses a pollen-specific promoter (ProLAT52) to generate epitope-tagged polysomal-RNA complexes that could be purified with a specific
Transcriptional regulation has been the main focus for gene regulation in the past. However, a tremendous amount of evidence from recent studies also indicates that translational regulation plays a key role during development, cell cycle control, and mechanisms related to acute drug resistance. Gene expression analysis on actively translated mRNA transcripts provides a unique approach to study post transcriptional regulation. Previous studies have relied on a traditional sucrose gradient ultracentrifugation procedure to isolate polysome complexes and requires a large amount of cells (up to 500 million cells). As a result, this still remains a major bottleneck for the investigation of post transcriptional regulation with limited quantities of clinical samples. Therefore, there is an urgent need to develop a novel approach to isolate actively translated polysomes from a small number of cells (10 to 500 cells). The new approach will allow us to systematically study potential translational ...
Blinded analyses of the concentrations of binding proteins for retinol and retinoic acid (CRABP) in homogenates of cancer and normal tissue aliquots obtained from human cervix, endometrium, ovary, breast, and lung were carried out by the sucrose gradient ultracentrifugation technique. In carcinomas of the cervix and endometrium, CRABP mean values of 50.4 and 123.2 pmol/g tissue, respectively, were detected. Such concentrations represent a 3- and 4-fold increase over the mean values of CRABP in the normal cervix (16.9 pmol/g) and normal endometrium (30.8 pmol/g), respectively. In carcinomas of the ovary, the mean CRABP level was 128.6 pmol/g compared to a maximal mean value of ≦0.46 pmol/g in the normal ovary. Elevated levels of CRABP were also found in breast and lung carcinomas compared to the amounts detected in the same patient in normal tissue aliquots of the same organ.. The differences between CRABP concentrations in cervical, endometrial, ovarian, and breast carcinomas and those in ...
Specialized tube for rapid PBMC isolation by density gradient centrifugation. Manufactured under cGMP and can be used for IVD applications.
In order to study the release of ATP from a neuronal preparation thought to be purinergic, isolated varicosities were prepared from myenteric plexus by mincing and homogenizing the longitudinal muscle of guinea pig ileum in 0.32 M sucrose. The presence of varicosities within the crude preparation (P2), isolated by differential centrifugation, was confirmed by electron microscopy and by the presence of occluded lactate dehydrogenase as a cytoplasmic marker. Varicosities were purified further from the P2 fraction on a discontinuous sucrose density gradient and characterized morphologically. Varicosities resembling cholinergic, purinergic, and adrenergic axonal terminals were identified. The release of ATP from both crude and purified varicosities was detected by monitoring the light produced when the released ATP reacted with firefly luciferin-luciferase which was present in the incubation medium. Elevated extracellular K+ or Rb+ caused the release of ATP, whereas elevated Na+ and Li+ did not. ...
Assays detecting antigen (Ag)-specific T-cell responses in immune-mediated processes are increasingly employed to understand disease pathogenesis and immune staging. The quantity and quality of starting peripheral blood mononuclear cell (PBMC) preparations are important factors in the performance of such assays. We therefore compared final PBMC yield and function by modifying parameters at the blood drawing, storage and processing steps. While drawing blood in vacuum-driven tubes or syringes and separating PBMCs on density gradients using standard or membrane (Leucosep (R)) tubes made no difference, storing tubes for 18 h without any agitation led to PBMC preparations contaminated with granulocytes and decreased interferon (IFN)-gamma enzyme-linked immunospot (ELISpot) responses. Even agitated blood showed a trend towards reduced ELISpot responses and increased human leukocyte Ag (HLA) multimer readouts when stored for 18 h compared to 3 h. These changes were reduced by diluting blood prior to ...
Constitutive secretory vesicles carrying heparan sulfate proteoglycan (HSPG) were identified in isolated rat hepatocytes by pulse-chase experiments with [35S]sulfate and purified by velocity-controlled sucrose gradient centrifugation followed by equilibrium density centrifugation in Nycodenz. Using this procedure, the vesicles were separated from plasma membranes, Golgi, trans-Golgi network (TGN), ER, endosomes, lysosomes, transcytotic vesicles, and mitochondria. The diameter of these vesicles was approximately 100-200 nm as determined by electron microscopy. A typical coat structure as described for intra-Golgi transport vesicles or clathrin-coated vesicles could not be seen, and the vesicles were not associated with the coat protein beta-COP. Furthermore, the vesicles appear to represent a low density compartment (1.05-1.06 g/ml). Other constitutively secreted proteins (rat serum albumin, apolipoprotein E, and fibrinogen) could not be detected in purified HSPG-carrying vesicles, but banded in ...
The dynamics of the flow inside an evaporating sessile droplet of water with polystyrene micro-spheres of 1.0 μm in diameter in suspension is described. The initial volume of the droplets is in the ra
Separation of particles according to density by employing a gradient of varying densities. At equilibrium each particle settles in the gradient at a point equal to its density. (McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed ...
Cytochrome P-450 in rat liver nuclear membranes of untreated, PB-pretreated, or 3-MC-pretreated animals was spectrally similar to corresponding liver microsomes. Epoxide hydrase activity of liver nuclear membrane and microsomes was induced by PB administration; 3-MC did not cause any induction. The same results were obtained when either the DNase digestion method or sucrose density centrifugation technique for preparation of nuclear membranes was used.. ...
Total CD4+ Th cells from IRF4+/+ or −/− mice were purified using the first steps of the multisort kit (Miltenyi Biotec) as described previously (5). 106/ml of purified Th cells were stimulated in vitro for 72 h with 5 μg/ml of immobilized anti-CD3 mAb, 2.5 μg/ml of soluble anti-CD28 mAb (BD Biosciences), or 100 U/ml of recombinant human IL-2 as described previously (5). After a resting period of 48-72 h in the absence of anti-CD3 mAb, but in the presence of IL-2, viable cells were purified on a Ficoll density gradient and restimulated in the presence of IL-2 either with anti-CD3 (immobilized at 5 μg/ml) or with 1 μg/ml of soluble anti-Fas mAb Jo2 (Becton Dickinson) together with 2 μg/ml of protein G (Sigma-Aldrich). Apoptosis was analyzed by annexin V and propidium iodide (PI) staining as described previously (12) at the indicated times after secondary stimulation. Viable as well as dead cells were included in this analysis according to forward and side scatter characteristics. The role ...
Equilibrium sedimentation uses a gradient of a solution to separate particles based on their individual densities (mass/volume). A pivotal aspect about this type of sedimentation is that it is completely independent of the shape of the molecule. It is used to purify the differential centrifugation. A solution is prepared with the densest portion of the gradient at the bottom. Particles to be separated are then added to the gradient and centrifuged. Each particle proceeds until it reaches an environment of comparable density. Such a density gradient may be continuous or prepared in an incremental fashion. For instance, when using sucrose to prepare density gradients, one can carefully float a solution of 40% sucrose onto a layer of 45% sucrose and add further less dense layers above. The homogenate, prepared in a dilute buffer and centrifuged briefly to remove tissue and unbroken cells, is then layered on top. After centrifugation typically for an hour at about 100,000 x g, disks of cellular ...
We investigated the function of Cav1. 4 MgCl2. The intracellular alternative included (in millimeter focus) 130 beliefs < .05 were considered significant. Outcomes Sucrose-density lean fractionation of protein involved in depolarization-induced insulin release in Inches-1 Cav1 and cells.2/II-III cells The KATP funnel, made up of Kir6.2 and SUR1 subunits, has a central function in the insulin release stimulated by blood sugar and sulfonylureas. The localization was examined by us of Kir6.2, SUR1, and EPAC2 in lipid rafts by fractionating the Triton A100-insoluble part of Cav1 and Inches-1.2/II-III cell lysates in discontinuous sucrose gradients. EPAC2 is normally reported to interact straight with both Piccolo (21) and SUR1 (19), and we discovered that both EPAC2 and SUR1 are extremely focused in lipid number fractions of sucrose gradients (the user interface of 5% and 30% sucrose) in both Inches-1 cells and Cav1.2/II-III cells (Figure 1). We assessed the localization of the KATP funnel subunit ...
We investigated the function of Cav1. 4 MgCl2. The intracellular alternative included (in millimeter focus) 130 beliefs < .05 were considered significant. Outcomes Sucrose-density lean fractionation of protein involved in depolarization-induced insulin release in Inches-1 Cav1 and cells.2/II-III cells The KATP funnel, made up of Kir6.2 and SUR1 subunits, has a central function in the insulin release stimulated by blood sugar and sulfonylureas. The localization was examined by us of Kir6.2, SUR1, and EPAC2 in lipid rafts by fractionating the Triton A100-insoluble part of Cav1 and Inches-1.2/II-III cell lysates in discontinuous sucrose gradients. EPAC2 is normally reported to interact straight with both Piccolo (21) and SUR1 (19), and we discovered that both EPAC2 and SUR1 are extremely focused in lipid number fractions of sucrose gradients (the user interface of 5% and 30% sucrose) in both Inches-1 cells and Cav1.2/II-III cells (Figure 1). We assessed the localization of the KATP funnel subunit ...
Results. Western blot analysis of PKCα and PKCγ translocation. N/N 1003A cells were treated with 200 nM TPA (a conventional PKC activator) for 60 min, 10 ng/ml EGF, or 15 ng/ml bFGF. Confluent cells were taken up in ice cold 50 mM Tris, 20 mM MgCl2, 25 μg/ml aprotinin, and 25 μg/ml leupeptin, pH 7.5 and sonicated. The lysate was spun down for 60 min at 35,000 rpm (100,000x g). Western blot analysis was performed on the pellet that contained plasma membrane and the supernatant that contained cytosolic components.. Upon activation by TPA, PKCα and PKCγ should translocate from the cytosol to the plasma membrane [25]. In order to determine if PKCα and PKCγ translocate to the plasma membrane in N/N 1003A cells upon activation, TPA was added to the cell culture media for 60 min. PKCα did translocate to the membrane (pellet fraction) upon activation of the enzyme by TPA (Figure 1A, lane 2 versus lane 4). PKCγ also translocated to the membrane (pellet fraction, Figure 1B, lane 2 versus lane ...
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The present invention relates to methods of enriching for desired cell population from cell sources, such as body fluids, dispersed tissue specimens and cultured cells. In particular, the present inve
As of today, there are 39 hydrogen fueling stations in the U.S., all but four of them in the state of California. They serve roughly 39 million Californians, whereas 284 million Americans outside the state have no access to hydrogen fuel-cell cars nor stations at which to refill them. Germany, on the other hand, has 83 million residents-and 45...
Sucrose Density Gradient Centrifugation Separation of Gold - Sensor basics: Types, functions and applications Martin Rowe while the application of sensors and their associated algorithms has multiplied and changed the
Gradient Makers, C.B.S. Scientific - Each (10 ML) : Designed for use in sucrose density gradient centrifugation, gradient gel electrophoresis, and li
Sucrose density gradient centrifugation& recover fractions-,SDS-PAGE(10% gel) (GM1 would migrate with the dye front)-,electro-transfer to nitrocellulose (or PVDF) membrane -,blocking with 4% skimmilk for 30min-, incubate with Ctx-HRP in 4% skimmilk for 1h-,wash PBS-Tween x 3times-,Detection with ECL. ...
Thus, if a mixture of minerals with different densities can be placed in a liquid with an intermediate density, the grains with densities less than that of the liquid will float and grains with densities greater than the liquid will sink. Typical mineral densities range from about 2.2 g/cc to as much as 8 g/cc, but are generally between 2.5 and .... ...
Gravity currents impact significantly the environment and humans life. Understanding their internal structure and their dynamics is fundamental for modelling purposes. In this study we observe experimentally how bed porosity affects the dynamics of density currents. We will focus on density and velocity profiles of currents traveling over porous bed, sinking and entraining water from its cavities. This abstract presents the preliminary results of this study which are relative to density currents travelling over the reference substratum, a smooth impermeable bed. The tests presented will be used as a reference to observe how the water entrained from the bottom cavities can change the structure of the current and are herein used to validate the experimental techniques. Brine water is injected at constant discharge in a 7.5 m long, 0.3 m wide and 0.3 m deep flume while instantaneous velocity and density profiles are simultaneously acquired 3 m downstream of the inlet. Initial excess densities and inlet
pluriSelect separation tubes have been developed for optimal separation of leukocytes and peripheral blood mononuclear cells (PBMC) from whole blood and bone marrow. pluriMate and TwinSpin prevents you from time-consuming and laborious overlaying of the sample material. During centrifugation, Leukocytes, lymphocytes and PBMCs are separated from unwanted erythrocytes and granulocytes, depending on the density gradient media ...
Find out all of the information about the Deltalab product: sedimentation analysis microplate 141746. Contact a supplier or the parent company directly to get a quote or to find out a price or your closest point of sale.
try using percol gradient centrifugation. ive forgotten the manufacturer. they do have protocols for separating live/dead cells. you will have to optimize the percol density(s) for your type of cells. you may also have to use more than one density gradient of percol to separate the layers of dead and live cells.good luck. john. ...
Adenovirus are efficient gene delivery systems. The standard method for purification of adenoviral vectors is based on using a cesium chloride (CsCl) density gradient combined with ultracentrifugation. This method is suitable for small-scale purification and is less expensive than column chromatography or commercial purification kits.
Studies on the composition and structure of plant viruses can only be attempted after their purification from infected tissue homogenates. Owing to their intrinsic differences, however, the ease with which different viruses can be purified varies considerably. A few relatively stable viruses, e.g. tobacco mosaic virus or turnip yellow mosaic virus, can be treated with high concentrations of salt and precipitated by acid or alcohol without inactivation. With less stable viruses such methods are usually unsuccessful. The particles of such viruses, however, can be sedimented by ultracentrifugation and procedures developed frequently involve two treatments: (a) differential centrifugation and concentration after initial clarification of buffered homogenates such as with organic solvents (Steere, 1956; Tomlinson, Shepherd & Walker, 1959; Wetter, 1960), (b) further fractionation by rate or equilibrium density gradient centrifugation as developed by Brakke (1960). Because of the small capacity of the
Technetium-99m-Exametazime (HMPAO) is widely used for radiolabelling leucocytes for localization of infection. The subcellular distribution of radionuclide in the labelled cells and the distribution of radioactivity among the leucocyte population are incompletely understood. Frozen section autoradiography was used to determine quantitatively the distribution of Tc-99m in leucocytes labelled with Tc-99m-Exametazime. Sections of rapidly frozen suspensions of labelled leucocytes in plasma were autoradiographed on Ilford K2 emulsion and stained with haematoxylin and eosin. Neutrophils, eosinophils and mononuclear cells were separated by Percoll density gradient centrifugation. Cell nuclei were isolated by a rapid cell-breakage and fractionation method. In a typical experiment mean grain densities [grains/100 mu m(2) (ESD)] over cells were: eosinophils 31.2 (18.4), neutrophils 3.5 (3.5), mononuclear cells 4.2 (5.1). Mean grain numbers per cell (ESD) were: eosinophils 13 (6.8), neutrophils 1.3 (1.3), ...
The molecular nature and replicative behavior of R factor 222 was examined in Proteus mirabilis . In deoxyribonucleic acid (DNA} from R+ P. mirabilis , R factor 222 was identified by CsCl density gradient centrifugation as 2 satellite DNA bands at densities corresponding to 50 and 58 moles percent guanine plus cytosine (% GC) . Replication of the 50 and 58% GC components of R factor 222 in P. mirabilis was analyzed during growth in the presence and absence of chloramphenicol (CAM} and after shifting exponentialand stationary- phase cells to conditions which inhibit host protein or DNA synthesis . CAM reduced the cellular growth rate but increased the amount of both R factor components relative to host chromosomal DNA . However, the 58% GC component showed a larger proportionate increase. This was inferred to indicate reduced synthesis of an inhibitor that acts on both R factor components and an initiator required for replication of the 50% GC component . Replicative patterns observed after shifting
Molecular Cloning, also known as Maniatis, has served as the foundation of technical expertise in labs worldwide for 30 years. No other manual has been so popular, or so influential.
On Wed, 30 Sep 1998, Patrick wrote: , Hi all. , , A request from first-time user of CsCl/EtBr for plant DNA purification: , Upon addition of ethidium bromide to my phenol/chloroform purified , Plant-DNA solution in CsCl, I observe a change of color from normal , orange-red to light cherry red. If I remember correctly this was also the case, when I have purified DNA with CsCl density gradient centrifugation. , Check http://www.biologie.uni-ulm.de/bio2/knoop/images/ethidium.jpg , for visual impression... Unfortunatly my display doesnt show the correct colours. , After centrifugation color accumulated COMPLETELY with a sluggish , material on top of the gradient. Considering the possibility that some , remaining protein/polysaccharides may be responsible for this, I have , taken the remainder of clear solution and repeated the EtBr addition. No , improvement - same color change (even after repeating this for the 4th , time!) , , Anybody ideas what is going on? , I dont know, what is going on, but ...
Abstract: t had been reported that the GCHV(Gtass Carp Hemorrhage Virus)is a new member.in Reoviri-dae and has 11 polypeptides by our laboratory,We did further experiments on the viral polypeptides.The virions(GCHV-875)were purified from the medium of infection culture cells by CsCl density gradient centrifugation,dissolved in the buffer containing Tris-HCl 250mmol/L,glycerin 25%,SDS86mmol/L,2-ME 320mmol/L(pH6.9)and then heated at 100℃for 3 min.After that,by addingan equal volume of FITC solution(2mg/M)in 1 mol/L NaHCO3,the viral polypeptides were labelledwith FITC,and analysed by SDS-PAGE. All the viral structural polypeptides(11 bands),namelyVP130,VP120,VP113,VP107,VP75,VP65,VP60,VP52,VP42,VP39 and VP31,wete visible under. UV light.These polypeptides were retrieved by cutting down each band from the gel and demonstrated kept immunological property.The method can determine 500ng protein band in gel under UVlight.. ...
A simple technique is described for the isolation of Toxoplasma gondii tissue cysts from fetal ovine brain by centrifugation on a discontinuous density gradient of 30 per cent and 90 per cent Percoll (colloidal silica solution). Brain samples from 51 aborted ovine fetuses were examined by both the Percoll and mouse inoculation techniques; eight infections were detected by the Percoll technique compared to 12 by mouse inoculation. Possible reasons for this discrepancy and the scope for improving the Percoll technique are discussed.. ...
An analytical ultracentrifluge was used to characterize the behaviors of ultracentrifugal sedimentation of bovine serum albumin solutions. We obtained data both for the sedimentation velocity and for the growth rate of the sediment that accumulates during ultracentrifugation. Effects of the rotor speed, the solution concentration, and the volume of the solution contained in the cell on the sedimentation behaviors were investigated experimentally. The sedimentation behaviors and the properties of the compressible sediment were explained well in terms of the theory which has been applied with respect to particulate suspension. Finally, it was shown that such factors of the solution environment as the solution pH and the salt concentration have a large effect on the sedimentation behaviors.. ...
Mitochondria are essential for the manufacturing of major and secondary metabolites, which largely decide the standard of fruit. Nevertheless, a technique for isolating high-quality mitochondria is at present not out there in citrus fruit, stopping high-throughput characterization of mitochondrial capabilities. Right here, primarily based on differential and discontinuous Percoll density gradient centrifugation, we devised.... ...
Mitochondria are essential for the manufacturing of major and secondary metabolites, which largely decide the standard of fruit. Nevertheless, a technique for isolating high-quality mitochondria is at present not out there in citrus fruit, stopping high-throughput characterization of mitochondrial capabilities. Right here, primarily based on differential and discontinuous Percoll density gradient centrifugation, we devised.... ...
Outer-arm dynein purified from trout spermatozoa was disrupted by low-ionic-strength dialysis, and the resulting subunits were separated by sucrose density-gradient centrifugation. The intact 19 S dynein, containing the alpha- an beta-heavy chains, intermediate chains (ICs) 1-5 and light chains (LCs) 1-6, yielded several discrete particles: a 17.5 S adenosine triphosphatase (ATPase) composed of the alpha- and beta-chains ICs 3-5 and LC 1; a 9.5 S complex containing ICs 1 and 2 together with LCs 2, 3, 4, and 6; and a single light chain (LC 5), which sedimented at approximately 4 S. In some experiments, ICs 3-5 also separated from the heavy chain complex and were obtained as a distinct subunit. Further dissociation of the 17.5 S particle yielded a 13.1 S ATPase that contained the beta-heavy chain and ICs 3-5. The polypeptide compositions of the complexes provide new information on the intermolecular associations that occur within dynein. Substructural features of the trout dynein polypeptides also were
Harvest cowpea plants about 2 weeks after inoculation, then homogenize at 4°C in two volumes of 0.5 M citrate buffer (pH 7.0) containing 0.1% thioglycollic acid. Express juice through cheesecloth, and add 20 ml carbon tetrachloride to every 100 ml extract. Shake the extract for 15 min, and clarify by low-speed centrifugation. Concentrate the virus by three cycles of differential centrifugation. Resuspend the pellets from high speed centrifugation in 0.01 M citrate buffer. Purify further by sucrose density-gradient centrifugation (Tsuchizaki et al., 1971).. ...
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Exosomes purification and analyses comprise a fast evolving research area; more than 70% of published research on exosomes has been done within the last six years. Challenges to researchers working with exosomes include setting up density gradients by hand, because it is tedious, time-consuming and subject to user, lab, and method variability.
Previous studies in this laboratory have allowed the formulation of a model for the molecular arrangement of C5, C6, C7, C8, and C9 on the surface of cells undergoing immune cytolysis with an assigned cumulative m.w. of 995,000. To verify directly the existence of a C5-C9 complex, serum samples containing radiolabeled terminal components were activated at 37°C with EA, antigen-antibody complexes, CVF, inulin or zymosan. Subsequent sucrose density gradient ultracentrifugation showed that all treatments cited led to the formation, in varying degrees, of rapidly sedimenting material which incorporated C5, C6, C7, C8, and C9, but not C3. The reaction was inhibited by 0.01 M EDTA and 0°C. The complex had a sedimentation coefficient of 22.4S, a diffusion coefficient of 1.98 × 10-7 cm2 sec-1 and thus a calculated m.w. of 1.04 × 106.. ...
Purification of mitochondria and mitochondrial subfractionation. Mitochondria were purified from brain tissue using the discontinuous sucrose gradient method. Briefly, brain homogenate was made in ice-cold homo-buffer (0.32 M sucrose, 20 mM Tris-HCl, pH 7.4) and spun at 900 g, 4°C, for 10 minutes. The supernatant was transferred to another clean tube and spun at 10,000 g, 4°C, for 10 minutes. The resultant pellet, enriched for mitochondria, was resuspended in 2 ml homo-buffer, loaded on top of a sucrose gradient (1.2 M, 0.8 M, and 0.32 M sucrose; 20 mM Tris-HCl, pH 7.4) and spun at 53,000 g, 4°C, for 2 hours. The white band at the interface between medium (0.8 M) and heavy (1.2 M) solutions was collected as highly purified mitochondria. Mitochondria from cultured cells were isolated using a kit (catalog no. 89874) from Pierce. Mitochondrial subfractionation was carried out as described by Hovius et al. (40). Briefly, purified mitochondria (1 mg) were resuspended in 500 μl ice-cold buffer (10 ...
Cells. The Raji control cell line and the cell lines expressing either DC-SIGN (Raji-DC-SIGN) or L-SIGN (Raji-L-SIGN) were cultured as previously described (4). PBMCs were isolated from buffy coats by standard Ficoll-Hypaque density centrifugation, activated with phytohemagglutinin (3 μg/ml), and cultured in RPMI medium containing 10% FCS, penicillin (100 units/ml), and streptomycin (100 units/ml). On day 3 the cells underwent CD4+ enrichment by incubation with CD8 immunomagnetic beads (Dynal Biotech) and were negatively selected according to the manufacturers instructions and cultured with IL-2 (100 U/ml). DCs for the single-cycle transmission assay were generated from fresh PBMCs with cells layered on a standard Percoll gradient (Amersham Pharmacia). The light fraction with predominantly monocytes was collected, washed, and seeded in 24-well or 6-well culture plates at a density of 5 × 105 cells or 2.5 × 106 per well, respectively. After 60 minutes at 37°C, the adherent cells were ...
Cells. The Raji control cell line and the cell lines expressing either DC-SIGN (Raji-DC-SIGN) or L-SIGN (Raji-L-SIGN) were cultured as previously described (4). PBMCs were isolated from buffy coats by standard Ficoll-Hypaque density centrifugation, activated with phytohemagglutinin (3 μg/ml), and cultured in RPMI medium containing 10% FCS, penicillin (100 units/ml), and streptomycin (100 units/ml). On day 3 the cells underwent CD4+ enrichment by incubation with CD8 immunomagnetic beads (Dynal Biotech) and were negatively selected according to the manufacturers instructions and cultured with IL-2 (100 U/ml). DCs for the single-cycle transmission assay were generated from fresh PBMCs with cells layered on a standard Percoll gradient (Amersham Pharmacia). The light fraction with predominantly monocytes was collected, washed, and seeded in 24-well or 6-well culture plates at a density of 5 × 105 cells or 2.5 × 106 per well, respectively. After 60 minutes at 37°C, the adherent cells were ...
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Molecular Cloning, also known as Maniatis, has served as the foundation of technical expertise in labs worldwide for 30 years. No other manual has been so popular, or so influential.
Centrifugation is the use of the centrifugal forces generated in a spinning rotor to separate biological particles, such as cells, viruses, sub‐cellular organelles, macromolecules (principally proteins and nucleic acids) and macromolecular complexes (such as ribonucleoproteins and lipoproteins)
The growth of a mouse leukemia virus in an established mouse cell line was examined after the line became contaminated with an unidentified Mycoplasma species. The contaminated cultures grew well in small plastic cultures dishes, but they could not be propagated in larger roller bottles unless the growth medium was changed frequently. Cells from Mycoplasma-contaminated and Mycoplasma-free cultures were exposed to 3H-labeled uridine for 24 hr. Culture fluids were harvested 2 or 24 hr after labeling and purified by centrifugation through discontinuous sucrose gradients. Considerably less uridine-3H-labeled virus was recovered from supernatant fluids of Mycoplasma-contaminated cultures than from Mycoplasma-free cultures. Equilibrium sedimentation in sucrose gradients of uridine-3H-labeled material from culture supernatants of contaminated cultures produced 3H peaks at buoyant densities of 1.20 to 1.24 and 1.16 to 1.18 g/ml. Virus titers in culture fluids from Mycoplasma-contaminated cultures were ...
The present invention generally encompasses the control of the release rate of agents from a polymeric matrix. This control over the release rate of agents provides for control over, inter alia, the therapeutic, prophylactic, diagnostic, and ameliorative effects that are realized by a patient in need of such treatment. In addition, the control of the release rate of agents also has an effect upon the mechanical integrity of the polymeric matrix, as well as a relationship to a subjects absorption rate of the absorbable polymers.
Ns by differential centrifugation. B and C. Immunoblot analysis of soluble/ insoluble fractions separated by differential centrifugation. FKIPS DCARD stable
Plasma membrane(PM) protein accounts for a small fraction of total cellular protein in plants but performs a very critical role in plant physiology. Isolation and purification of PM protein from plant tissues have been traditionally done by sucrose density ultracentrifugation and aqueous two-phase partitioning. These methods, while relatively effective, require ultracentrifugation and large amount of starting material. The procedures are usually tedious and time consuming.To overcome the shortcomings, we have developed this PM isolation kit. Plant tissues are first sensitized by buffer A, homogenized, and pass through a specialized filter cartridge that allows homogenates to pass through with a zigzag path. The cell membranes are ruptured into a range of predefined size during the process. Native plasma membranes are separated from a mixture of un-ruptured cells, nuclei, cytosol and organelles by subsequent differential centrifugation and density centrifugation without using ...
I am having trouble isolating mtDNA using CsCl gradients. Is there an easier way to isolate mtDNA? I was wondering if anyone could give me some help! This is becoming a really big problem!!!!! Thank oyu very much, Rodney Earl Pettway Rodney Earl Pettway Department of Plant Pathology and Microbiology Texas A&M University College Station, TX 77843 Pettway at ppserver.tamu.edu ...
Density marker beads are small colored microspheres of known density that are used for calibrating density gradients and determining density in gradient columns
StraightFrom® Whole Blood MicroBeads for cell isolation directly from whole blood without the need for density gradient centrifugation. - 日本
Find a potocol and data for the isolation of exosomes without density gradient centrifugation and fast screening of isolated exosomes by flow cytometry. | Ireland
JML-135-2007-105-114. This paper contains the results of a new experimental study of the effect of temperature on density, refractive index on mixing and ultrasonic velocity for a number of linear n-alkanes conbined with ethanol. A perusal of deviations between the experimental and calculated derived magnitudes shows that the predictive procedures give a qualitative estimation for the studied mixtures due to their high nonideality.. ...
2014.1-2015.12 Academy of Mathematics & Systems Science Postdoc within the research center NCMIS, working with Prof. Ping Zhang on density patch problem ...
View Notes - ps_2 from CHEMICAL E 20.410j at MIT. DOWNSTREAM PROCESSING Problem Set #2 Problem 1 A key to understanding centrifugation is understanding the equations that describe it. One can
The classical procedure for estimating the approximate size of a protein is by its sedimentation coefficient, determined either using an analytical ultr...
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