Abstract：
OBJECTIVE:The central mechanism for establishing a self-tolerant and functional T cell repertoire includes the promiscuous expression of otherwise tissue-restricted proteins by medullary thymic epithelial cells (TEC). We here demonstrate a novel and highly efficient method for isolating this rare key cell type. METHODS:We combined the enrichment of medullary TEC via UEA-1 MicroBeads with the subsequent depletion of residual CD45+ hematopoietic cells via specific size exclusion and compared our results to the standard Percoll enrichment method and isolation procedure via flow cytometric cell sorting. RESULTS:The addition of 2 μl UEA-1 MicroBeads per 108 thymus cells turned out best for optimal enrichment (an average of 22% purity compared to 1.2% for Percoll) and yield (an average of 1.73 × 105 medullary TEC per thymus compared to 5.16 × 104 for Percoll). After depletion of residual CD45+ cells, our method not only reached a purity of 75.5% but also turned out less stressful for the cells as compared to flow cytometric cell sorting. CONCLUSION:We here provide a fast and versatile procedure for enriching medullary TEC that yields higher purity and recovery rates than the standard Percoll enrichment method Our enrichment procedure in combination with CD45+ depletion via specific size exclusion is comparable to the current gold standard flow cytometric cell sorting method. SIGNIFICANCE STATEMENT:We developed a fast and versatile procedure to isolate a high number medullary TEC to investigate the biochemical processes of medullary TEC in more depths.
journal_name
J Immunol Methodsjournal_title
Journal of immunological methodsauthors
Dohr D,Engelmann R,Müller-Hilke Bdoi
10.1016/j.jim.2019.02.001subject
Has Abstractpub_date
2019-04-01 00:00:00pages
12-18eissn
0022-1759issn
1872-7905pii
S0022-1759(18)30281-3journal_volume
467pub_type
杂志文章abstract：:The proinflammatory cytokine IL-18 mediates IFN-gamma production as well as the induction of Th1 polarized immune responses in synergy with IL-12. In this study, we describe the production of isogeneic monoclonal antibodies (Mabs) directed against murine IL-18 (mIL-18). Immunization of IL-18-deficient mice with recomb...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(01)00505-1
更新日期：2002-01-01 00:00:00
abstract：:Embryonic stem cells provide a potentially convenient source of macrophages in the laboratory. Given the propensity of macrophages for plasticity in phenotype and function, standardised culture and differentiation protocols are required to ensure consistency in population output and activity in functional assays. Here...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2012.06.008
更新日期：2012-11-30 00:00:00
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journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2007.01.020
更新日期：2007-04-30 00:00:00
abstract：:The large scale preparation of two second generation immunotoxins containing murine monoclonal antibodies and deglycosylated ricin A chain is described. The procedure for the preparation of immunotoxins consists of the derivatization of antibody with SMPT and reduction of dgA with DTT followed by their reaction to est...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(91)90110-2
更新日期：1991-09-13 00:00:00
abstract：:The antigen capture capacity of antibodies covalently immobilized on injection-molded polystyrene beads was evaluated. Bromoacetyl groups on the bead surfaces rendered them reactive to protein nucleophilic groups. The bromoacetyl surface exhibited up to a ten-fold greater capacity for protein compared to unmodified po...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(88)90136-6
更新日期：1988-07-22 00:00:00
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journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2017.04.005
更新日期：2017-08-01 00:00:00
abstract：:Two commercially available bioreactor systems, CELLine and miniPERM, were evaluated for their ability to support the production of monoclonal antibody (mAb) from a variety of murine hybridoma cell lines. Production and purity of mAbs were compared between the two systems and with mouse ascites tumour fluid generation....
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(02)00081-9
更新日期：2002-06-01 00:00:00
abstract：:We describe here a spot test to detect antibodies to non-erythroid cell surface antigens, using ethidium bromide staining to visualize areas of target cell lysis. This assay allows rapid screening of large numbers of samples of culture supernatants, antisera, or ascitic fluids. Antibody titers determined by this spot ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(82)90367-2
更新日期：1982-01-01 00:00:00
abstract：:Nontyphoidal isolates of Salmonella (NTS), particularly Salmonella Typhimurium, are a major cause of invasive bacteremia in Africa. Despite this, no vaccine against NTS is currently available for use in humans. If a NTS vaccine is to be developed in a timely manner, there is a need to develop assays to assess its in v...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2012.10.005
更新日期：2013-01-31 00:00:00
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journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(97)90008-9
更新日期：1977-01-01 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
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journal_title：Journal of immunological methods
pub_type： 杂志文章,评审
doi：10.1016/0022-1759(92)90078-8
更新日期：1992-06-24 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：:A simple and rapid method was developed for immunofluorescence measurements of cells by flow cytometry which does not require washing procedures, permitting absolute enumeration of cell subpopulations. Peripheral blood cells were labeled with fluorescein and phycoerythrin conjugated monoclonal antibodies and the nucle...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(89)90034-3
更新日期：1989-09-29 00:00:00
abstract：:Conventional culture conditions for GM-CSF expanded murine bone marrow derived dendritic cells (BMDCs) uses ambient (hyperoxic) oxygen pressure (20% v/v, 152 Torr) and medium supplemented with the thiol 2-mercaptoethanol (2-Me). Given the redox activities of O2 and 2-Me, the effects of 2%, 5%, 10%, and 20% v/v O2 atmo...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2005.10.012
更新日期：2006-01-20 00:00:00
abstract：:As much as one percent of antibodies in human plasma are reported to be specific for the non-human disaccharide structure αGal. Various microbes express αGal. However, the implications of anti-αGal antibodies for the anti-microbial defenses are poorly established. With the perspective of studying the biological import...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2011.07.017
更新日期：2011-10-28 00:00:00
abstract：:A competition radioimmunoassay for human superoxide dismutase (SOD-1) is described. Radioiodinated SOD-1 is incubated with unlabeled competitor and limiting antibody, then immune complexes are isolated using Staphylococcus aureus. The assay is sensitive (detecting 5 ng enzyme) and reproducible. Average values for SOD-...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(79)90313-2
更新日期：1979-01-01 00:00:00
abstract：:Studies on the kinetics of antigenic peptide binding to major histocompatibility complex class II molecules have been used extensively to probe major histocompatibility complex (MHC) structure as well as to investigate the molecular mechanism of peptide recognition. Previous experiments have frequently been carried ou...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(94)90290-9
更新日期：1994-07-12 00:00:00
abstract：:A panel of 18 monoclonal antibodies was raised to the human calcitonin gene related peptide (CGRP). Of these mabs, seven were specific for alpha CGRP and five for beta CGRP, while the remainder reacted with both alpha and beta CGRP. Nine different epitopes on CGRP were defined with these mabs. In addition, the mabs we...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(90)90115-c
更新日期：1990-11-06 00:00:00
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journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(02)00183-7
更新日期：2002-09-15 00:00:00
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journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2012.01.004
更新日期：2012-03-30 00:00:00
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journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(96)00030-0
更新日期：1996-06-10 00:00:00
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journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(99)00037-x
更新日期：1999-05-27 00:00:00
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journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(94)90093-0
更新日期：1994-01-03 00:00:00
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journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2015.03.005
更新日期：2015-05-01 00:00:00
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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：:In the guinea pig system, there are no T lymphocyte tumors available. In addition, in this species the production of T cell lines in vitro for analysis of T cell markers or function proved to be difficult. As an alternative approach, guinea pig X mouse and guinea pig X rat T cell hybridomas were obtained by fusion of ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(89)90003-3
更新日期：1989-03-31 00:00:00
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journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(87)90454-6
更新日期：1987-03-12 00:00:00
abstract：:Overlapping peptide scans prepared by Spot synthesis have been used to map interaction sites in several systems. Here we report our experience with this approach to identify peptides from the variable parts of anti-hapten, anti-peptide and anti-protein antibodies that retain their specific antigen-binding capacity in ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(02)00140-0
更新日期：2002-09-01 00:00:00
abstract：:The purpose of this study was to develop a flow cytometric method for the detection of adenosine deaminase (ADA) in a single cell suspension of mononuclear cells. Anti-human ADA antibody was purified by affinity chromatography on a column of Sepharose 4B to which calf ADA was covalently linked. This antibody was used ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(85)90017-1
更新日期：1985-06-25 00:00:00
