Abstract：
:Although enveloped retroviruses bud from the cell surface of T lymphocytes, they use the endocytic pathway and the internal membrane of multivesicular bodies for their assembly and release from macrophages and dendritic cells (DCs). Exosomes, physiological nanoparticles produced by hematopoietic cells, egress from this same pathway and are similar to retroviruses in terms of size, density, the molecules they incorporate and their ability to activate immune cells. Retroviruses are therefore likely to contaminate in vitro preparations of exosomes and vice versa and sucrose gradients are inefficient at separating them. However, we have found that their sedimentation velocities in an iodixanol (Optiprep) velocity gradient are sufficiently different to allow separation and purification of both vesicles. Using acetylcholinesterase as an exosome marker, we demonstrate that Optiprep velocity gradients are very efficient in separating exosomes from HIV-1 particles produced on 293T cells, primary CD4(+) T cells, macrophages or DCs, with exosomes collecting at 8.4-12% iodixanol and HIV-1 at 15.6%. We also show that immunodepletion with an anti-acetylcholinesterase antibody rapidly produces highly purified preparations of HIV-1 or exosomes. These findings have applications in fundamental research on exosomes and/or AIDS, as well as in clinical applications where exosomes are involved, more specifically in tumour therapy or in gene therapy using exosomes generated from DCs genetically modified by transfection with virus.
journal_name
J Immunol Methodsjournal_title
Journal of immunological methodsauthors
Cantin R,Diou J,Bélanger D,Tremblay AM,Gilbert Cdoi
10.1016/j.jim.2008.07.007subject
Has Abstractpub_date
2008-09-30 00:00:00pages
21-30issue
1-2eissn
0022-1759issn
1872-7905pii
S0022-1759(08)00216-0journal_volume
338pub_type
杂志文章abstract：:A sensitive method for staining proteins after transfer from polyacrylamide gels to nitrocellulose paper is described. Transferred proteins are first derivatized by reaction of the nitrocellulose replica with sulfosuccinimidobiotin and are then reacted sequentially with streptavidin, rabbit anti-streptavidin, and hors...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(86)90504-1
更新日期：1986-08-21 00:00:00
abstract：:Ethanol-fixed paraffin-embedded specimens of human tissues were studied whether the surface antigens of leukocytes in these tissues can be stained and analyzed. Three-layer indirect immunoperoxidase staining was performed on the ethanol-fixed paraffin-embedded sections by the use of several monoclonal antibodies for w...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(90)90189-3
更新日期：1990-08-07 00:00:00
abstract：:A simple and highly sensitive analytical isoelectric focusing (IEF) technique using immunofixation with anti-C2 serum followed by silver staining has been developed in order to study simultaneously the structural polymorphism of both native C2 and C2 activation fragments (C2a MW 74,000 and C2b MW 34,000). Structural C...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(89)90112-9
更新日期：1989-02-08 00:00:00
abstract：:We have developed specific immunoassays for secretory peroxidase (SP) and for myeloperoxidase (MP) (polymorphonuclear leukocyte-derived peroxidase) in human saliva. Antibodies against SP and MP were produced using bovine milk lactoperoxidase (LP) and human MP as the immunogens, respectively. The methods developed are ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(91)90154-8
更新日期：1991-08-09 00:00:00
abstract：:Leukemia inhibitory factor (LIF), oncostatin-M (OSM), ciliary neurotrophic factor (CNTF) and cardiotrophin-1 (CT1) act through transmembrane receptors which share the gp190 glycoprotein chain. The understanding of its involvement in the biology of these cytokines is of importance since these systems have recently been...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(97)00074-4
更新日期：1997-07-14 00:00:00
abstract：:We have developed a large scale (10 g/batch) non-chromatographic method for the collection of IgM directly from plasma obtained from therapeutic plasmapheresis of patients. The technique uses sequential precipitations from ammonium sulfate and polyethylene glycol, and gives high yields of undenatured IgM antibodies. T...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(93)90252-3
更新日期：1993-01-14 00:00:00
abstract：:Serologically and biochemically pure HLA antigens and other beta 2-microglobulin-associated proteins were isolated using a rapid immunoprecipitation technique with a monoclonal anti-beta 2-microglobulin reagent A88. Elution of the immunoprecipitate with beta 2-microglobulin isolated from kidney patients proved to be e...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(81)90237-4
更新日期：1981-01-01 00:00:00
abstract：:Effector-target conjugation between different cell populations of human NK cells and K562 tumor cells has been studied from binding isotherms obtained from data of effector (alpha) and target (beta) conjugate frequencies measured by flow cytometry analysis at different effector-to-target ratios. Non-linear and linear ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(97)00153-1
更新日期：1997-12-01 00:00:00
abstract：:An easy rapid method for layering mammalian blood on Ficoll-Isopaque gradients is described. ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(75)90036-8
更新日期：1975-11-01 00:00:00
abstract：:We propose a new data analysis approach for reactive oxygen species detection using Dihydrorhodamine 123 in blood monocytes and neutrophils. This approach, based on data transformation using lymphocytes as internal standard, allows to appreciate free radical production by monocytes also without Phorbol 12-myristate 13...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2012.02.015
更新日期：2012-05-31 00:00:00
abstract：:Fungi represent emerging infectious threats to human populations worldwide. Mice and other laboratory animals have proved invaluable in modeling clinical syndromes associated with superficial and life-threatening invasive mycoses. This review outlines salient features of common vertebrate animal model systems to study...
journal_title：Journal of immunological methods
pub_type： 杂志文章,评审
doi：10.1016/j.jim.2014.03.022
更新日期：2014-08-01 00:00:00
abstract：:A flow cytometric NK assay was developed in which the K562 targets were labelled with the fluorogenic substrate, carboxyfluorescein diacetate (c'FDA). This new assay compared favourably with results obtained using the conventional 51Cr-release assay. c'FDA was not toxic to target cells and did not inhibit lysis. The a...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(86)90258-9
更新日期：1986-01-22 00:00:00
abstract：:To maximize safety in the setting of an increasing number of requests for flow cytometric analysis of specimens potentially contaminated with the AIDS retrovirus, we evaluated some commonly used fixatives for their ability to inactivate the infectious potential of the virus. We found that both formaldehyde (0.37% v/v)...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(86)90278-4
更新日期：1986-01-22 00:00:00
abstract：:We describe a stable chelating linkage for the reversible immobilization of oligohistidine tagged proteins in the flow cell of the 'BIAcore' surface plasmon resonance (SPR) biosensor. The carboxymethylated dextran surface of the flow cell was covalently derivatized with N-(5-amino-1-carboxypentyl)iminodiacetic acid (N...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(95)00032-6
更新日期：1995-06-14 00:00:00
abstract：:Natural Killer cells are cells of the innate immune system that are important for the recognition and clearance of virally infected cells or tumors. Examination of the development and signaling of these cells has been severely hampered due to an inability to over-express proteins in these cells. We developed a novel t...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2008.10.014
更新日期：2009-01-30 00:00:00
abstract：:The Cellular Antigen Stimulation Test (CAST), a recent innovation in the laboratory detection of allergen sensitivity, is a functional assay based on the release of sulphidoleukotrienes (LTs) from allergen-activated circulating basophils. The current study was undertaken to establish optimum determinants of this proce...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2003.12.014
更新日期：2004-05-01 00:00:00
abstract：:A competitive enzyme immunoassay for rat prolactin using an immunogen and a tracer obtained by recombinant DNA technology is described. Polyclonal antibodies were raised in rabbits immunized with a purified chimeric protein consisting of rat prolactin fused with two synthetic immunoglobulin G binding domains derived f...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(94)90264-x
更新日期：1994-03-10 00:00:00
abstract：:A high-capacity manifold support with immobilized antigen was developed for the analysis of IgE-mediated immune reactivity in allergic subjects. Using this 96-pronged support, specific antibodies were trapped and detected from large sets of serum samples. We describe the binding of large amounts of antigen onto the ex...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(94)90218-6
更新日期：1994-01-12 00:00:00
abstract：:A highly sensitive fluorimetric enzyme immunoassay was developed for prostaglandin H (PGH) synthase, an intrinsic membrane protein, using n-octyl beta-D-glucopyranoside for solubilization of the synthase from the endoplasmic reticulum membrane. An anti-PGH synthase IgG-coated polystyrene ball was reacted with a PGH sy...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(93)90403-t
更新日期：1993-06-04 00:00:00
abstract：:A new compact and cell dense continuous culture system was developed. The system consisted of a culture bag, which had two compartments separated by a semipermeable membrane, and an external rotator. Cells were cultured in the inner compartment with medium containing serum or the necessary growth factors. The outer co...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(89)90049-5
更新日期：1989-03-10 00:00:00
abstract：:The identification and characterization of human mast cell (MC) functions are hindered by the shortage of MC populations suitable for investigation. Here, we present a novel technique for generating large numbers of well differentiated and functional human MCs from peripheral stem cells (=peripheral stem cell-derived ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2014.07.003
更新日期：2014-11-01 00:00:00
abstract：:We describe an ex vivo rat infection model to study protective immunity against Fasciola hepatica at the gut level. An exact number of newly excysted juveniles (NEJs) was injected into a gut segment with an intact blood supply and which was still attached to a live anaesthetized rat. NEJs that penetrated the gut wall ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(98)00026-x
更新日期：1998-04-15 00:00:00
abstract：:The fluorescence activated cell sorter (FACS) was used with an indirect membrane immunofluorescence technique to detect antibody against the Forssman antigen, a glycosphingolipid. Sheep erythrocytes, which contain Forssman antigen as a major membrane glycosphingolipid, were used as the target antigen. Detection of the...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(83)90098-4
更新日期：1983-02-25 00:00:00
abstract：:Hapten-carrier protein conjugates were made using five different small haptens (MW < 1000), two carrier proteins and two methods of conjugation. Nondenaturing agarose gel electrophoresis was used to demonstrate that when as few as two molecules of a small hapten are attached to the carrier, the conjugate band migrates...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(93)90317-z
更新日期：1993-09-15 00:00:00
abstract：:A cell membrane permeabilizing treatment is described which involves the use of lysolecithin at low concentration in acidic acetate buffer and paraformaldehyde fixation. It preserved well-separated scatter cytograms of small and large lymphocytes. The accuracy of the immunochemical detection of internal antigens by fl...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(91)90162-9
更新日期：1991-04-25 00:00:00
abstract：:Six monoclonal and two polyclonal antibodies to fluorescein (FLU) were affinity purified and immobilized on Immulon 2 polystyrene as capture antibodies (CAbs): (a) by passive adsorption at pH 9.6, (b) via a streptavidin bridge to a biotinylated carrier molecule, and (c) via an antiglobulin which had been previously ad...
journal_title：Journal of immunological methods
pub_type： 杂志文章,评审
doi：10.1016/0022-1759(92)90066-3
更新日期：1992-06-24 00:00:00
abstract：:A new system for assaying monoclonal antibodies consisting of an 8 x 12 array of sticks which fits into a 96-well microtiter plate is described. Tests using virus specific monoclonal antibodies and virus proteins demonstrated sensitivity equivalent to the conventional microtiter plate assay. Antibody production, antig...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(83)90356-3
更新日期：1983-03-25 00:00:00
abstract：:Immunoglobulins of the IgE and IgG classes have been causally associated with hypersensitivity reactions in man and in numerous animal species including mice, rats and guinea pigs. The use of the guinea pig as an animal model for both pulmonary and dermal hypersensitivity reactions, and the recent recognition of the i...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(91)90359-n
更新日期：1991-05-17 00:00:00
abstract：:Macrophages can be separated from other cell types by their ability to readily attach and spread on glass or on plastic surfaces which are treated for optimal growth of cultured cells (tissue culture-treated plastic). To detach macrophages from these surfaces, techniques must be used which require prior preparation of...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(84)90142-x
更新日期：1984-03-30 00:00:00
abstract：:We describe the setting up and validation of a reporter gene assay for type I IFN based on monkey Vero cells transfected with pMx-Luc, a plasmid carrying the luciferase gene under the control of the type I IFN inducible mouse Mx1 promoter. Vero cells were stably transfected with pMx-Luc and clone 3-143/5 was selected ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(96)00168-8
更新日期：1996-11-29 00:00:00
