Abstract：
:The study of organogenesis, tissue-homeostasis and regeneration requires the precise assessment of in vivo cell proliferation. To this end a host of methods have been developed to detect and quantify DNA synthesis in proliferating cells. These include cell labeling with various nucleotide analogues and fluorescence reporter-based animal models with each method presenting its idiosyncratic shortcomings. Quantitative assessment of epithelial cell turnover has been partly hampered due to their variable and limited in vivo accessibility and the requirement for harsher isolation procedures to procure single cells for FACS analysis. Here, we report a reliable protocol to study in vivo cell proliferation of epithelial cells in mice by repeatedly injecting EdU intravenously for an extended 12-day period. EdU incorporation was quantitated ex vivo by FACS after tissue dissociation in order to obtain single epithelial cell suspensions. As a lead population, we analyzed thymic epithelial cells (TECs), where we were able to label compartmentalized TEC subsets to saturation without apparent toxic effects on the thymus architecture or stress-sensitive TEC lineage differentiation. The data is in concordance with the prevailing model of medullary TEC terminal differentiation that includes the post-Aire stage. The same protocol was successfully applied to epithelial cells of various other organs - skin, lymph node, kidney and small intestine - tissues with widely varying frequencies and rates of proliferating epithelial cells.
journal_name
J Immunol Methodsjournal_title
Journal of immunological methodsauthors
Michel C,Küchler R,Martin I,Kyewski B,Pinto Sdoi
10.1016/j.jim.2018.03.015subject
Has Abstractpub_date
2018-06-01 00:00:00pages
82-86eissn
0022-1759issn
1872-7905pii
S0022-1759(17)30539-2journal_volume
457pub_type
杂志文章abstract：:In studies of the relation between structure and function of proteins of the immune system, there is a continuous need for screening of a large number of protein variants. To optimise the yield following transient gene expression in small or medium culture volumes, several parameters were investigated. First, secretio...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2005.01.002
更新日期：2005-03-01 00:00:00
abstract：:The metaphase index assay (MIA) for thyroid growth stimulators, as originally described, used FRTL-5 thyroid cells cultured in Bellco culture chambers and glass microscope slides. The metaphases were observed using the nuclear strain aceto-lacto orcein. However the surface properties of the glass proved to be variable...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(88)90067-1
更新日期：1988-06-28 00:00:00
abstract：:Cytotoxicity assays are widely used to evaluate the functional activity of NK and T cells against tumour target cells and the release of radioactive sodium chromate from labelled target cells is still the most commonly used marker of target lysis in culture supernatants. We describe here the standardization of a micro...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(94)90104-x
更新日期：1994-06-24 00:00:00
abstract：:Macrophages are an important part of the cellular immune system and play a key role during immune responses. Thus, macrophages are interesting targets in basic and clinical research. Primary monocytes or monocyte-derived macrophages do not proliferate on a suitable scale so that their use for functional studies in vit...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2009.03.014
更新日期：2009-05-31 00:00:00
abstract：:For antibody production, the O-phosphorylated derivative of tyrosine, threonine, or serine was covalently linked to succinylated bovine albumin via the carbodiimide reaction. Each conjugate was then complexed with methylated bovine albumin for immunization of rabbits. To determine binding, the corresponding O-phosphor...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(89)90360-8
更新日期：1989-11-30 00:00:00
abstract：BACKGROUND:Dendritic cells (DCs) are the most potent antigen-presenting cells. They can be generated in vitro from CD14+ cells, and also from CD34+ progenitor cells. Although T cell proliferation using [3H] thymidine incorporation assay has been used widely to check DC function, this technique only provides limited inf...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(03)00002-4
更新日期：2003-04-01 00:00:00
abstract：:This paper describes a modification of the peroxidase technique by which immunophenotyping may be carried out on routinely air-dried blood and bone marrow (BM) smears. The method is simple and quick, requires no special equipment, can be performed on fresh or stored specimens and gives a standard of morphological deta...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(86)90428-x
更新日期：1986-10-23 00:00:00
abstract：:Paraneoplastic neurological syndromes (PNS) are devastating neurological disorders secondary to cancer, associated with onconeural autoantibodies. Such antibodies are directed against neuronal antigens aberrantly expressed by the tumor. The detection of onconeural antibodies in a patient is extremely important in diag...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2013.02.017
更新日期：2013-05-31 00:00:00
abstract：:The microenvironment of the thymus consists of functionally discrete niches composed of distinct stromal cell subsets. Clinically relevant changes affecting T-cell differentiation occur within these niches with age and injury caused by irradiation and chemotherapy treatments. The study of thymic stromal cells has been...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2007.09.010
更新日期：2008-01-01 00:00:00
abstract：:To date, the applications of bispecific antibodies in immunoassay and immunocytochemical procedures have been directed at uniting two different biomolecules through the binding of epitopes on each respective substance. In this study, bispecific antibodies were constructed in which both binding sites were directed to t...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(94)90042-6
更新日期：1994-05-16 00:00:00
abstract：:In this paper, we describe a fluorimeter-based, closed-tube, polymerase chain reaction (PCR) assay for the detection and quantification of the mRNA of porcine interleukin 1alpha (IL1alpha) and interleukin 2 (IL2) cytokines in peripheral blood leukocytes (PBLs) using melting curve analysis and compare it to a standard ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(99)00114-3
更新日期：1999-11-19 00:00:00
abstract：:Exosomes are nanometer-sized vesicles, secreted from most cell types, with documented immune-modulatory functions. Exosomes can be purified from cultured cells but to do so effectively, requires maintenance of cells at high density in order to obtain sufficient accumulation of exosomes in the culture medium, prior to ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2008.03.001
更新日期：2008-06-01 00:00:00
abstract：:We have developed a modified microtiter plate which has following advantageous features and functions to both conventional microtiter plate and protein array, such as 1) use of conventional microtiter plate reader and washer, and 2) allowance of simultaneous reaction in the same liquid for all wells. Four proteins of ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2007.04.007
更新日期：2007-06-30 00:00:00
abstract：:We have observed that the red reaction product of alkaline phosphatase immuno-conjugates and certain substrate preparations produces a brilliant red fluorescence that is visible by fluorescence microscopy using both fluorescein and rhodamine filter combinations. This provides a level of sensitivity greater than that o...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(90)90396-d
更新日期：1990-08-28 00:00:00
abstract：:The chemiluminescence (CL) assay has been used to measure the reactive oxygen species (ROS)-generating capacity of phagocytes. To achieve more optimal measurement conditions for a multi-channel microplate photon-counting CL analyzer with the cooled charge-coupled device (CCD) camera which offers enhanced sensitivity, ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(97)00159-2
更新日期：1997-12-15 00:00:00
abstract：:Many normal and pathological processes have been shown to be influenced by the quantitative and qualitative nature of cytokine production. Cytokines have also been shown to be central in modulating host responses to invasion by pathogens. Therefore detection and analysis of cytokine production are critical in the unde...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(97)00124-5
更新日期：1998-03-01 00:00:00
abstract：:A new T(H1)/T(H2) in vitro model for mechanistic studies and drug screening in human T cells was established working with ficoll-separated PBMCs or elutriated lymphocytes cultured in serum-free medium. Human T cells could be kept viable and reactive in this medium for several months. In this model, superantigens (SAs)...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(98)00142-2
更新日期：1998-10-01 00:00:00
abstract：:The circumsporozoite (CS) protein is the major surface protein of Plasmodium sporozoites. Antibodies to the immunodominant repeat domain of CS immobilize sporozoites and prevent infection of hepatocytes. Plasmodium falciparum vaccines containing CS repeats are undergoing human trials in endemic areas, and proof of eff...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2004.06.017
更新日期：2004-09-01 00:00:00
abstract：:An enzyme-linked immunosorbent assay (ELISA) was developed for screening production of monoclonal antibodies with specificity for HBsAg. Mouse hybridoma IgG were firstly extracted from assay medium with goat anti-mouse IgG adsorbed on polystyrene beads. The specific antibody was revealed by saturation with HBs antigen...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(82)90361-1
更新日期：1982-01-01 00:00:00
abstract：:Antibody-dependent cell-mediated cytotoxicity (ADCC) is an important mechanism of action implicated in the clinical efficacy of several therapeutic antibodies. In vitro ADCC assays employing effector cells capable of inducing lysis of target cells bound by antibodies are routinely performed to support the research and...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2014.03.021
更新日期：2014-05-01 00:00:00
abstract：:Enzyme-linked antibody was used to treat unfixed herpesvirus-infected human fetal lung cell cultures in a mode which permitted the visualizing of local sites of infection. Foci containing as few as 20 herpesvirus-infected cells produced sufficient viral mass to be easily detectable by this method. 'Clouds' or 'plumes'...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(81)90361-6
更新日期：1981-01-01 00:00:00
abstract：:An improved sensitivity to low levels of hepatitis B surface antigen (HBsAg) was demonstrated when ELISA results were interpreted by a statistical method. The absorbance of each test well was compared with the mean absorbance of all low-colour test samples. Those wells with an absorbance of more than two standard devi...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(88)90316-x
更新日期：1988-11-25 00:00:00
abstract：:Conditions for desorbing a DNP carrier from its immunoadsorbent were studied in various hydro-organic media at different pHs and temperatures, including the sub-zero range. It appeared that DNP--anti-DNP interaction is the result of a balance between various bonds and the best result (95% yield) was obtained in hydro-...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(78)90105-9
更新日期：1978-01-01 00:00:00
abstract：:Design of automated image processing systems to determine migration characteristics of individual cells is not trivial. Every test sample requires separate recording and the analysis of individual cell tracks in two- or three-dimensional migration systems by time-lapse microscopy is extremely laborious. Here, we descr...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(03)00262-x
更新日期：2003-09-01 00:00:00
abstract：:Apoptosis and primary necrosis are the two modes of cell death induced by a lethal injury. The majority of structural and biochemical events occurring during cell death can be analysed by flow cytometry. The 7-aminoactinomycin D (7-AAD) assay can be used to detect the loss of membrane integrity during apoptosis of mur...
journal_title：Journal of immunological methods
pub_type： 杂志文章,评审
doi：10.1016/s0022-1759(02)00072-8
更新日期：2002-07-01 00:00:00
abstract：:We evaluated an adjuvant, TiterMax, as an alternative to complete Freund's adjuvant (CFA) for producing antisera in animals. TiterMax, consists of a microparticulate stabilized water-in-oil emulsion of a metabolizable oil, squalene, with the adjuvant block copolymer CRL89-41. This paper reports two evaluations of Tite...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(92)90302-a
更新日期：1992-08-30 00:00:00
abstract：:A monoclonal antibody to human IgG was tested with myeloma proteins of the four IgG subclasses. When tested by immunofluorometric assay, enzyme-linked immunosorbent assay, hemagglutination and hemagglutination inhibition assays, the antibody reacted with IgG3 but not with the other three IgG subclasses. When tested by...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(87)90317-6
更新日期：1987-02-11 00:00:00
abstract：:A quantitative, minimized version of the slide immunoenzymatic assay (SIA) is described, which is performed with a 5 microliters drop of each reagent on a circular glass surface (3 mm in diameter) delimited by a thin layer of a hydrophobic material. Measurements are done directly, without disturbing the drop of substr...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(83)90143-6
更新日期：1983-04-15 00:00:00
abstract：:Radioimmunoassays are usually developed empirically since there have been few established rules which cover general RIA behaviour. Evidence is presented which enables the delineation of 2 types of RIA with distinct group characteristics. (1) Simple RIA, conforming to the law of mass-action kinetics, is a result of uni...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(85)90441-7
更新日期：1985-11-28 00:00:00
abstract：:IL-21 is a pleiotropic cytokine that plays a key role in modulating inflammatory responses, including the promotion of autoimmune diseases. Several groups have quantitated circulating levels of IL-21 in plasma and serum samples using various commercial ELISAs. We determined, however, that the most commonly used commer...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2018.12.005
更新日期：2019-03-01 00:00:00
