Techniques to improve the direct ex vivo detection of low frequency antigen-specific CD8+ T cells with peptide-major histocompatibility complex class I tetramers.
AuthorsChattopadhyay PK, Melenhorst JJ, Ladell K, Gostick E, Scheinberg P, Barrett AJ, Wooldridge L, Roederer M, Sewell AK, Price DA,
JournalCytometry A
PubMed ID18836993
'The ability to quantify and characterize antigen-specific CD8+ T cells irrespective of functional readouts using fluorochrome-conjugated peptide-major histocompatibility complex class I (pMHCI) tetramers in conjunction with flow cytometry has transformed our understanding of cellular immune responses over the past decade. In the case of prevalent CD8+ T cell populations that ... More
Easily reversible desthiobiotin binding to streptavidin, avidin, and other biotin-binding proteins: uses for protein labeling, detection, and isolation.
AuthorsHirsch JD, Eslamizar L, Filanoski BJ, Malekzadeh N, Haugland RP, Beechem JM, Haugland RP
JournalAnal Biochem
PubMed ID12419349
'The high-affinity binding of biotin to avidin, streptavidin, and related proteins has been exploited for decades. However, a disadvantage of the biotin/biotin-binding protein interaction is that it is essentially irreversible under physiological conditions. Desthiobiotin is a biotin analogue that binds less tightly to biotin-binding proteins and is easily displaced by ... More
Nonradioactive methods for the assay of phosphoinositide 3-kinases and phosphoinositide phosphatases and selective detection of signaling lipids in cell and tissue extracts.
AuthorsGray A, Olsson H, Batty IH, Priganica L, Peter Downes C
JournalAnal Biochem
PubMed ID12605860
'We describe a novel approach to quantitation of phosphoinositides in cell extracts and in vitro enzyme-catalyzed reactions using suitably tagged and/or labeled pleckstrin homology (PH) domains as probes. Stable complexes were formed between the biotinylated target lipid and an appropriate PH domain, and phosphoinositides present in samples were detected by ... More
Development of a ubiquitin transfer assay for high throughput screening by fluorescence resonance energy transfer.
'An assay based on fluorescence resonance energy transfer (FRET) has been developed to screen for ubiquitination inhibitors. The assay measures the transfer of ubiquitin from Ubc4 to HECT protein Rsc 1083. Secondary reagents (streptavidin and antibody to glutathione-S-transferase [GST]), pre-labeled with fluorophores (europium chelate, Eu(3+), and allophycocyanin [APC]), are noncovalently ... More
Cell surface markers in human embryonic stem cells.
AuthorsRao RR, Johnson AV, Stice SL,
JournalMethods Mol Biol
PubMed ID18453248
'The pluripotent nature of human embryonic stem cells (hESCs) is based on their potential to form every cell type in the body. Prior to use in directed differentiation strategies, these cells need to be thoroughly characterized. The large number of glycoproteins and carbohydrates that exist on the cell surface provide ... More
Signaling alterations in activation-induced nonresponsive CD8 T cells.
AuthorsTham EL, Mescher MF
JournalJ Immunol
PubMed ID11489986
'Costimulation-dependent production and autocrine use of IL-2 by activated CD8 T cells results in initial clonal expansion, but this is transient. The cells quickly become anergic, unable to produce IL-2 in response to Ag and costimulation, irrespective of the form of costimulation. This activation-induced non-responsiveness (AINR) differs from "classical" anergy ... More
Quantitative measurement of mast cell degranulation using a novel flow cytometric annexin-V binding assay.
AuthorsDemo SD, Masuda E, Rossi AB, Throndset BT, Gerard AL, Chan EH, Armstrong RJ, Fox BP, Lorens JB, Payan DG, Scheller RH, Fisher JM
JournalCytometry
PubMed ID10404150
'BACKGROUND: Mast cells are primary mediators of allergic inflammation. Antigen-mediated crosslinking of their cell surface immunoglobulin E (IgE) receptors results in degranulation and the release of proinflammatory mediators including histamine, tumor necrosis factor-alpha, and leukotrienes. METHODS: Mast cells were stimulated to degranulate by using either IgE crosslinking or ionophore treatment. ... More
'Two homogeneous proximity assays for tyrosine kinases, scintillation proximity assay (SPA) and homogeneous time-resolved fluorescence (HTRF), have been developed and compared. In both formats, the kinase assay was performed using biotinylated peptide substrate, ATP ([33P]ATP in the case of SPA), and tyrosine kinase in a 96-well assay format. After the ... More
Characterization of plasmablasts in the blood of HIV-infected viremic individuals: evidence for nonspecific immune activation.
AuthorsBuckner CM, Moir S, Ho J, Wang W, Posada JG, Kardava L, Funk EK, Nelson AK, Li Y, Chun TW, Fauci AS
JournalJ Virol
PubMed ID23487459
'Terminal differentiation of B cells and hypergammaglobulinemia are hallmarks of B-cell hyperactivity in HIV disease. Plasmablasts are terminally differentiating B cells that circulate transiently in the blood following infection or vaccination; however, in HIV infection, they arise early and are maintained at abnormally high levels in viremic individuals. Here we ... More
Mature dendritic cells prime functionally superior melan-A-specific CD8+ lymphocytes as compared with nonprofessional APC.
AuthorsSalio M, Shepherd D, Dunbar PR, Palmowski M, Murphy K, Wu L, Cerundolo V
JournalJ Immunol
PubMed ID11466333
'Priming of melan-A(26/27-35)-specific CTL occurs only in a fraction of late stage melanoma patients, whereas during the early stages of the disease and in healthy volunteers, melan-A CTL have functional and phenotypic markers consistent with a naive phenotype. To study the requirements for expansion of naive melan-A CTL from healthy ... More
Visualization and characterization of respiratory syncytial virus F-specific CD8(+) T cells during experimental virus infection.
AuthorsChang J, Srikiatkhachorn A, Braciale TJ
JournalJ Immunol
PubMed ID11591747
'CTL play a major role in the clearance of respiratory syncytial virus (RSV) during experimental pulmonary infection. The fusion (F) glycoprotein of RSV is a protective Ag that elicits CTL and Ab response against RSV infection in BALB/c mice. We used the strategy of screening a panel of overlapping synthetic ... More
Visible diode lasers can be used for flow cytometric immunofluorescence and DNA analysis.
AuthorsDoornbos RM, De Grooth BG, Kraan YM, Van Der Poel CJ, Greve J
JournalCytometry
PubMed ID8187585
'This report describes a feasibility study concerning the use of a visible diode laser for two important fluorescence applications in a flow cytometer. With a 3 mW 635 nm diode laser, we performed immunofluorescence measurements using the fluorophore allophycocyanin (APC). We have measured CD8 positive lymphocytes with a two-step labeling ... More
Naive CD4 T cells constitutively express CD40L and augment autoreactive B cell survival.
AuthorsLesley R, Kelly LM, Xu Y, Cyster JG
JournalProc Natl Acad Sci U S A
PubMed ID16815973
'Chronic engagement of the B cell receptor by soluble autoantigen leads to reduced B cell survival. Using the Ig and hen egg lysozyme double transgenic mouse model, we demonstrate that the survival of soluble autoantigen-engaged B cells is further reduced in mice lacking CD4 T cells or deficient in CD40. ... More
Organ-specific regulation of the CD8 T cell response to Listeria monocytogenes infection.
AuthorsPope C, Kim SK, Marzo A, Masopust D, Williams K, Jiang J, Shen H, Lefrançois L
JournalJ Immunol
PubMed ID11207297
'The intestinal mucosal CD8 T cell response to infection with Listeria monocytogenes was measured using MHC class I tetramers and was compared with the response in peripheral blood, secondary lymphoid tissue, and liver. To assess the vaccination potential of Listeria and to analyze responses in C57BL/6 mouse strains, a recombinant ... More
Diabetes is predicted by the beta cell level of autoantigen.
AuthorsByersdorfer CA, Schweitzer GG, Unanue ER
JournalJ Immunol
PubMed ID16177075
'Two novel transgenic (Tg) strains were created expressing hen egg-white lysozyme (HEL) in a pancreas-specific fashion. RmHP.111 mice had levels of HEL per cell similar to that of the established ILK-3 strain, while RmHP.117 mice had 10-fold lower levels (50,000 molecules per cell). When bred to 3A9 TCR Tg mice, ... More
Expansion of mature thymocyte subsets before emigration to the periphery.
AuthorsPénit C, Vasseur F
JournalJ Immunol
PubMed ID9366410
'A small population of DNA-synthesizing mature thymocytes could be defined by analyzing cell surface markers and 5-bromo-2''-deoxyuridine (BrdUrd) labeling by four-color cytofluorometry. These cells have a completely mature phenotype (CD4- CD8+ or CD4+ CD8- TCR(high), HSA-, Qa-2(high)) and expand only weakly after BrdUrd incorporation. They recovered immediately in total number ... More
Cryptomonad algal phycobiliproteins as fluorochromes for extracellular and intracellular antigen detection by flow cytometry.
AuthorsTelford WG, Moss MW, Morseman JP, Allnutt FC
JournalCytometry
PubMed ID11309804
'BACKGROUND: Phycobiliproteins play an important role in fluorescent labeling, particularly for flow cytometry. The spectral properties of R-phycoerythrin (R-PE) and allophycocyanin (APC) have made them the dominant reagents in this class of fluorochromes. In this study, we evaluate a lesser-known but potentially important series of low-molecular weight cryptomonad-derived phycobiliproteins (commercially ... More
Parallel detection of antigen-specific T-cell responses by multidimensional encoding of MHC multimers.
AuthorsHadrup SR, Bakker AH, Shu CJ, Andersen RS, van Veluw J, Hombrink P, Castermans E, Thor Straten P, Blank C, Haanen JB, Heemskerk MH, Schumacher TN,
JournalNat Methods
PubMed ID19543285
'The use of fluorescently labeled major histocompatibility complex multimers has become an essential technique for analyzing disease- and therapy-induced T-cell immunity. Whereas classical major histocompatibility complex multimer analyses are well-suited for the detection of immune responses to a few epitopes, limitations on human-subject sample size preclude a comprehensive analysis of ... More
Apoptosis in hematopoietic cells is associated with an extensive decrease in cellular phosphotyrosine content that can be inhibited by the tyrosine phosphatase antagonist pervanadate.
AuthorsLund-Johansen F, Frey T, Ledbetter JA, Thompson PA
JournalCytometry
PubMed ID8891448
'In the present study, we investigated whether apoptosis in hematopoietic cells is associated with changes in cellular phosphotyrosine content. Murine thymocytes and B cells, human leukemia cells, and normal peripheral blood leukocytes were induced to undergo apoptosis by treatment with specific stimuli or by incubation in growth factor-deprived medium. Multiparameter ... More
Variable immunodominance hierarchies for H2-M3-restricted N-formyl peptides following bacterial infection.
AuthorsKerksiek KM, Busch DH, Pamer EG
JournalJ Immunol
PubMed ID11145694
'H2-M3-restricted presentation of N-formyl methionine (f-Met) peptides to CD8(+) T cells provides a mechanism for selective recognition of bacterial infection. In this report we demonstrate that Listeria monocytogenes infection induces distinct CD8(+) T cell populations specific for each of the known Listeria-derived formyl methionine peptides presented by M3. The sum ... More
A novel model of inflammatory bowel disease: mice deficient for the multiple drug resistance gene, mdr1a, spontaneously develop colitis.
AuthorsPanwala CM, Jones JC, Viney JL
JournalJ Immunol
PubMed ID9820555
'The murine multiple drug resistance (mdr) gene, mdr1a, encodes a 170-kDa transmembrane protein that is expressed in many tissues including intestinal epithelial cells, a subset of lymphoid cells and hematopoietic cells. We report that mdr1a knockout (mdr1a-/-) mice are susceptible to developing a severe, spontaneous intestinal inflammation when maintained under ... More
An absolute requirement for STAT4 and a role for IFN-gamma as an amplifying factor in IL-12 induction of the functional IL-18 receptor complex.
AuthorsNakahira M, Tomura M, Iwasaki M, Ahn HJ, Bian Y, Hamaoka T, Ohta T, Kurimoto M, Fujiwara H
JournalJ Immunol
PubMed ID11466347
'IL-12 and IL-18 are both proinflammatory cytokines that contribute to promoting Th1 development and IFN-gamma expression. However, neither IL-12R nor IL-18R is expressed as a functional complex on most resting T cells. This study investigated the molecular mechanisms underlying the induction of an IL-18R complex in T cells. Resting T ... More
Identification of HLA-B27-restricted peptides from the Chlamydia trachomatis proteome with possible relevance to HLA-B27-associated diseases.
AuthorsKuon W, Holzhütter HG, Appel H, Grolms M, Kollnberger S, Traeder A, Henklein P, Weiss E, Thiel A, Lauster R, Bowness P, Radbruch A, Kloetzel PM, Sieper J
JournalJ Immunol
PubMed ID11591805
'The association of HLA-B27 with ankylosing spondylitis and reactive arthritis is the strongest one known between an MHC class I Ag and a disease. We have searched the proteome of the bacterium Chlamydia trachomatis for HLA-B27 binding peptides that are stimulatory for CD8(+) cells both in a model of HLA-B27 ... More
Two distinct stages in the transition from naive CD4 T cells to effectors, early antigen-dependent and late cytokine-driven expansion and differentiation.
AuthorsJelley-Gibbs DM, Lepak NM, Yen M, Swain SL
JournalJ Immunol
PubMed ID11046030
'Efficient peptide presentation by professional APC to naive and effector CD4 T cells in vitro is limited to the first 1-2 days of culture, but is nonetheless optimum for effector expansion and cytokine production. In fact, prolonging Ag presentation leads to high levels of T cell death, decreased effector expansion, ... More
Competition between CTL narrows the immune response induced by prime-boost vaccination protocols.
AuthorsPalmowski MJ, Choi EM, Hermans IF, Gilbert SC, Chen JL, Gileadi U, Salio M, Van Pel A, Man S, Bonin E, Liljestrom P, Dunbar PR, Cerundolo V
JournalJ Immunol
PubMed ID11970981
'Recombinant vaccines encoding strings of virus- or tumor-derived peptides and/or proteins are currently being designed for use against both cancer and infectious diseases. These vaccines aim to induce cytotoxic immune responses against several Ags simultaneously. We developed a novel tetramer-based technique, based on chimeric HLA A2/H-2K(b) H chains, to directly ... More
Application of homogeneous time-resolved fluorescence (HTRFTM) to monitor poly-ubiquitination of wild-type p53.
AuthorsYabuki N, Watanabe S, Kudoh T, Nihira S, Miyamato C
JournalComb Chem High Throughput Screen
PubMed ID10539989
'Rapid degradation of wild-type p53 in the human uterine cervix is induced by the infection of high-risk human papilloma virus (HPV) types 16 and 18. HPV-E6 protein plays a critical role in the poly-ubiquitination of wild-type p53 by mediating the association of p53 with E6-associated protein (E6AP). As a result, ... More
Central nervous system microglial cell activation and proliferation follows direct interaction with tissue-infiltrating T cell blasts.
AuthorsSedgwick JD, Ford AL, Foulcher E, Airriess R
JournalJ Immunol
PubMed ID9605131
'Central nervous system (CNS)-resident macrophages (microglia) normally express negligible or low level MHC class II, but this is up-regulated in graft-vs-host disease (GvHD), in which a sparse CNS T cell infiltrate is observed. Relative to microglia from the normal CNS, those from the GvHD-affected CNS exhibited a 5-fold up-regulation of ... More
Expression of alpha(4)beta(7) integrin defines a distinct pathway of lymphoid progenitors committed to T cells, fetal intestinal lymphotoxin producer, NK, and dendritic cells.
AuthorsYoshida H, Kawamoto H, Santee SM, Hashi H, Honda K, Nishikawa S, Ware CF, Katsura Y, Nishikawa SI
JournalJ Immunol
PubMed ID11509590
'During embryogenesis, the Peyer''s patch anlagen are induced by a cell population that produces lymphotoxin (LT) alpha(1)beta(2) following stimulation of IL-7Ralpha. In this study, we show that the LT-producing cell is localized within the IL-7Ralpha(+) and integrin alpha(4)beta(7) (alpha(4)beta(7))(+) population in the embryonic intestine. Lineage commitment to the LT producer ... More
Use of fluorogenic histocompatibility leukocyte antigen-A*0201/HPV 16 E7 peptide complexes to isolate rare human cytotoxic T-lymphocyte-recognizing endogenous human papillomavirus antigens.
AuthorsYoude SJ, Dunbar PR, Evans EM, Fiander AN, Borysiewicz LK, Cerundolo V, Man S
JournalCancer Res
PubMed ID10667589
'Cervical cancer (CaCx) is the second most common female malignancy worldwide and remains a clinical problem despite improvements in early detection and therapy. CaCx and preinvasive cervical intraepithelial neoplasia (CIN3) are strongly associated with infection by human papillomavirus (HPV), particularly types 16 and 18. Two nonstructural viral proteins, E6 and ... More
Monitoring CD8 T cell responses to NY-ESO-1: correlation of humoral and cellular immune responses.
AuthorsJäger E, Nagata Y, Gnjatic S, Wada H, Stockert E, Karbach J, Dunbar PR, Lee SY, Jungbluth A, Jäger D, Arand M, Ritter G, Cerundolo V, Dupont B, Chen YT, Old LJ, Knuth A
JournalProc Natl Acad Sci U S A
PubMed ID10781081
'NY-ESO-1, a member of the cancer-testis family of antigens, is expressed in a subset of a broad range of different human tumor types. Patients with advanced NY-ESO-1-expressing tumors frequently develop humoral immunity to NY-ESO-1, and three HLA A2-restricted peptides were defined previously as targets for cytotoxic CD8(+) T cells in ... More
In vivo self-renewal of c-Kit+ Sca-1+ Lin(low/-) hemopoietic stem cells.
AuthorsOsawa M, Nakamura K, Nishi N, Takahasi N, Tokuomoto Y, Inoue H, Nakauchi H
JournalJ Immunol
PubMed ID8617942
'The long term marrow-repopulating ability of mouse bone marrow c-Kit+ Sca-1+ Lin(low/-) cells was studied. Injection of as few as 100 c-Kit+ Sca-1+ Lin(low/-) cells could rescue a lethally irradiated recipient mouse and reconstitute both myeloid and lymphoid cells in their peripheral blood for at least 10 mo. When limiting ... More
Kinetics of dendritic cell activation: impact on priming of TH1, TH2 and nonpolarized T cells.
AuthorsLangenkamp A, Messi M, Lanzavecchia A, Sallusto F
JournalNat Immunol
PubMed ID11017102
'To prime immune responses, dendritic cells (DCs) need to be activated to acquire T cell stimulatory capacity. Although some stimuli trigger interleukin 12 (IL-12) production that leads to T helper cell type I (TH1) polarization, others fail to do so and favor TH2 polarization. We show that after activation by ... More
A new model for CD8+ T cell memory inflation based upon a recombinant adenoviral vector.
AuthorsBolinger B, Sims S, O'Hara G, de Lara C, Tchilian E, Firner S, Engeler D, Ludewig B, Klenerman P
JournalJ Immunol
PubMed ID23509359
'CD8(+) T cell memory inflation, first described in murine CMV (MCMV) infection, is characterized by the accumulation of high-frequency, functional Ag-specific CD8(+) T cell pools with an effector-memory phenotype and enrichment in peripheral organs. Although persistence of Ag is considered essential, the rules underpinning memory inflation are still unclear. The ... More
Cytolytic activity of human natural killer cell subpopulations isolated by four-color immunofluorescence flow cytometric cell sorting.
'The natural killer activity and phenotypic properties of six different subpopulations of normal human peripheral blood lymphocytes obtained by four-color immunofluorescence cell sorting were examined. Phycoerythrin-conjugated CD16 and CD56 were used simultaneously to identify (CD16 + CD56+) NK cells. The cells most effective in mediating NK cytolysis against K562 target ... More
Approaches to multicolor immunofluorescence measurements.
AuthorsHoffman RA
JournalCytometry Suppl
PubMed ID3076363
Tracking T cells with tetramers: new tales from new tools.
AuthorsKlenerman P, Cerundolo V, Dunbar PR
JournalNat Rev Immunol
PubMed ID12001997
To understand the success or failure of immune responses against pathogens or tumours requires the direct measurement of specific lymphocytes. Recently, there has been an explosion of data in this field through the use of several new tools for measuring the number and function of T cells. This has allowed ... More
A high-throughput homogeneous assay for reverse transcriptase using generic reagents and time-resolved fluorescence detection.
AuthorsZhang JH, Chen T, Nguyen SH, Oldenburg KR
JournalAnal Biochem
PubMed ID10870833
A homogeneous time-resolved fluorescence (HTRF) assay has been developed for determining the activity of HIV reverse transcriptase (HIV-RT). By using a sequential capping strategy, the assay has been configured to utilize only generic reagents such as biotinylated dUTP, streptavidin-allophycocyanin, and streptavidin-europium. The assay was optimized for a HIV-RT high-throughput screen. ... More
Polypyrrole DNA chip on a silicon device: example of hepatitis C virus genotyping.
AuthorsLivache T, Fouque B, Roget A, Marchand J, Bidan G, Téoule R, Mathis G
JournalAnal Biochem
PubMed ID9451503
We describe in this article an oligonucleotide array constructed on a silicon device bearing a matrix of addressable 50-microns microelectrodes. Each electrode was covered by a conducting polymer (polypyrrole) grafted by an oligonucleotide (ODN). The DNA chip was prepared by successive electrochemically addressed copolymerizations of 5' pyrrole-labeled ODN and pyrrole. ... More
Using the Microcyte flow cytometer to monitor cell number, viability, and apoptosis in mammalian cell culture.
AuthorsHarding CL, Lloyd DR, McFarlane CM, Al-Rubeai M
JournalBiotechnol Prog
PubMed ID11027173
The Microcyte is a novel, portable flow cytometer based on diode laser technology whose use has been established for yeast and bacterial analysis. We present data that demonstrate its suitability for routine mammalian cell counting and viability determination. To extend its range of applications in the field of animal cell ... More
A profound deficiency in thymic progenitor cells in mice lacking Jak3.
AuthorsBaird AM, Lucas JA, Berg LJ
JournalJ Immunol
PubMed ID11034372
Humans and mice with genetic deficiencies that lead to loss of signaling through common gamma-chain (gammac)-containing cytokine receptors have severe defects in B and T lymphocytes. In humans, these deficiencies lead to a complete absence of T cells, whereas in mice, small thymuses give rise to normal numbers of peripheral ... More
Cutting edge: influence of the TCR V beta domain on the avidity of CD1d:alpha-galactosylceramide binding by invariant V alpha 14 NKT cells.
AuthorsSchümann J, Voyle RB, Wei BY, MacDonald HR
JournalJ Immunol
PubMed ID12794105
CD1d tetramers loaded with alpha-galactosylceramide (alpha-GalCer) bind selectively to mouse invariant Valpha14 (Valpha14i) NKT cells and their human counterparts. Whereas tetramer binding strictly depends on the expression of a Valpha14-Jalpha18 chain in murine NKT cells, the associated beta-chain (typically expressing Vbeta8.2 or Vbeta7) appears not to influence tetramer binding. In ... More
Isolating and engineering human antibodies using yeast surface display.
This protocol describes the process of isolating and engineering antibodies or proteins for increased affinity and stability using yeast surface display. Single-chain antibody fragments (scFvs) are first isolated from an existing nonimmune human library displayed on the yeast surface using magnetic-activated cell sorting selection followed by selection using flow cytometry. ... More
Cutting edge: rapid cloning of tumor-specific CTL suitable for adoptive immunotherapy of melanoma.
AuthorsDunbar PR, Chen JL, Chao D, Rust N, Teisserenc H, Ogg GS, Romero P, Weynants P, Cerundolo V
JournalJ Immunol
PubMed ID10358133
Adoptive immunotherapy using CTL has provided some clinical benefit to patients with metastatic melanoma. Use of cloned CTL of known specificity might improve clinical effect, but technical difficulties have limited exploration of this possibility. We have used fluorescence-driven cell sorting to clone tumor-specific CTL after staining with tetrameric MHC class ... More
Construction and application of a yeast surface-displayed human cDNA library to identify post-translational modification-dependent protein-protein interactions.
AuthorsBidlingmaier S, Liu B,
JournalMol Cell Proteomics
PubMed ID16321969
Although post-translational modifications such as phosphorylation mediate fundamental biological processes within the cell, relatively few methods exist that allow proteome-wide identification of proteins that interact with these modifications. We have constructed a yeast surface-displayed human cDNA library and utilized it to identify protein fragments with affinity for phosphorylated peptides derived ... More
Oral nickel tolerance: Fas ligand-expressing invariant NK T cells promote tolerance induction by eliciting apoptotic death of antigen-carrying, effete B cells.
AuthorsNowak M, Kopp F, Roelofs-Haarhuis K, Wu X, Gleichmann E,
JournalJ Immunol
PubMed ID16585548
Whereas oral nickel administration to C57BL/6 mice (Ni(high) mice) renders the animals tolerant to immunization with NiCl2 combined with H2O2 as adjuvant, as determined by ear-swelling assay, it fails to tolerize Jalpha18-/- mice, which lack invariant NKT (iNKT) cells. Our previous work also showed that Ni(high) splenic B cells can ... More
Co-localization of amyloid beta and tau pathology in Alzheimer's disease synaptosomes.
AuthorsFein JA, Sokolow S, Miller CA, Vinters HV, Yang F, Cole GM, Gylys KH,
JournalAm J Pathol
PubMed ID18467692
The amyloid cascade hypothesis proposes that amyloid beta (Abeta) pathology precedes and induces tau pathology, but the neuropathological connection between these two lesions has not been demonstrated. We examined the regional distribution and co-localization of Abeta and phosphorylated tau (p-tau) in synaptic terminals of Alzheimer's disease brains. To quantitatively examine ... More
Kinetics and efficacy of positive selection in the thymus of normal and T cell receptor transgenic mice.
AuthorsHuesmann M, Scott B, Kisielow P, von Boehmer H
JournalCell
PubMed ID1868548
DNA-labeling studies in alpha beta T cell receptor (TCR) transgenic mice show that the lifespan of immature CD4+8+ thymocytes is 3.5 days irrespective of whether they are selected for maturation or not. While nonselected cells die, the binding of the TCR to thymic major histocompatibility complex molecules rescues CD4+8+ cells ... More
Early onset of CD8 transgene expression inhibits the transition from DN3 to DP thymocytes.
AuthorsWack A, Coles M, Norton T, Hostert A, Kioussis D
JournalJ Immunol
PubMed ID10903721
In this paper we show that the effects of transgenic coreceptor expression on thymocyte development depend on the onset of transgene expression. Thus, a CD8 transgene expressed on CD44+CD25+ (DN2) and CD44-CD25+ (DN3) cells causes a partial block at the stage when TCRbeta selection takes place and diminishes expansion at ... More
Use of major histocompatibility complex class I tetramers to monitor tumor-specific cytotoxic T lymphocyte response in melanoma patients.
AuthorsCerundolo V
JournalCancer Chemother Pharmacol
PubMed ID10950154
There is now considerable evidence that human tumors often express antigens that render them susceptible to lysis by cytotoxic T lymphocytes (CTLs). These findings have raised hope for the development of cancer vaccines to trigger a tumor-specific immune response in cancer patients. To optimize the immunogenicity of cancer vaccines, it ... More
A role for c-myc in the regulation of thymocyte differentiation and possibly positive selection.
The purpose of thymocyte differentiation is to establish the T cell repertoire and eliminate nonfunctional and autoreactive T cells. In an analysis of potential regulators of this process, we have found that c-myc expression is down-regulated dramatically during early thymocyte differentiation and subsequently up-regulated along with TCR/CD3 in CD4+8+ cells. ... More
Ex vivo staining of metastatic lymph nodes by class I major histocompatibility complex tetramers reveals high numbers of antigen-experienced tumor-specific cytolytic T lymphocytes.
AuthorsRomero P, Dunbar PR, Valmori D, Pittet M, Ogg GS, Rimoldi D, Chen JL, Liénard D, Cerottini JC, Cerundolo V
JournalJ Exp Med
PubMed ID9802976
Characterization of cytolytic T lymphocyte (CTL) responses to tumor antigens has been impeded by a lack of direct assays of CTL activity. We have synthesized reagents ("tetramers") that specifically stain CTLs recognizing melanoma antigens. Tetramer staining of tumor-infiltrated lymph nodes ex vivo revealed high frequencies of tumor-specific CTLs which were ... More
Early establishment of diverse T cell receptor profiles for influenza-specific CD8(+)CD62L(hi) memory T cells.
AuthorsKedzierska K, Venturi V, Field K, Davenport MP, Turner SJ, Doherty PC
JournalProc Natl Acad Sci U S A
PubMed ID16754852
Single-cell analysis of endogenous, primary CD8(+) T cell responses to the influenza D(b)NP(366) and D(b)PA(224) epitopes indicates that prominent clonotypes bearing "public" or "shared" T cell receptors (TCRs) subset early into CD62L(hi) and CD62L(lo) populations. The CD62L(lo) effectors divide more and are rapidly eliminated during the contraction phase, whereas stable ... More
A PDZ domain-based detection system for enzymatic assays.
AuthorsFerrer M, Hamilton AC, Inglese J
JournalAnal Biochem
PubMed ID11814291
A time-resolved fluorescence resonance energy transfer (TR-FRET) detection method based on the formation of a PDZ domain.peptide ligand complex has been developed for enzymatic assays as an alternative to immuno-based detection strategies. The enzyme substrate is a "masked" biotinylated PDZ domain peptide ligand containing the consensus sequence Ser-X-Val-COOH. The critical ... More
Comparison of helium-neon and dye lasers for the excitation of allophycocyanin.
AuthorsLoken MR, Keij JF, Kelley KA
JournalCytometry
PubMed ID3542436
Allophycocyanin (APC) has a broad absorption spectrum permitting several different lasers to be used to excite this dye in a flow cytometer. A comparison was made between a dye laser and a helium-neon (HeNe) laser for the excitation of APC as an immunofluorescent chromophore. The ratio of fluorescence of stained ... More
Human CD1d-glycolipid tetramers generated by in vitro oxidative refolding chromatography.
AuthorsKaradimitris A, Gadola S, Altamirano M, Brown D, Woolfson A, Klenerman P, Chen JL, Koezuka Y, Roberts IA, Price DA, Dusheiko G, Milstein C, Fersht A, Luzzatto L, Cerundolo V
JournalProc Natl Acad Sci U S A
PubMed ID11248072
CD1 molecules are specialized in presenting lipids to T lymphocytes, but identification and isolation of CD1-restricted lipid specific T cells has been hampered by the lack of reliable and sensitive techniques. We here report the construction of CD1d-glycolipid tetramers from fully denatured human CD1d molecules by using the technique of ... More
Receptor proximity, not intermolecular orientation, is critical for triggering T-cell activation.
AuthorsCochran JR, Cameron TO, Stone JD, Lubetsky JB, Stern LJ
JournalJ Biol Chem
PubMed ID11384988
Engagement of antigen receptors on the surface of T-cells with peptides bound to major histocompatibility complex (MHC) proteins triggers T-cell activation in a mechanism involving receptor oligomerization. Receptor dimerization by soluble MHC oligomers is sufficient to induce several characteristic activation processes in T-cells including internalization of engaged receptors and up-regulation ... More
Rapid generation of broad T-cell immunity in humans after a single injection of mature dendritic cells.
Dendritic cells (DCs) are potent antigen-presenting cells that initiate protective T-cell immunity in mice. To study the immunogenicity of DCs in humans, we injected 9 healthy subjects subcutaneously with a control injection of autologous monocyte-derived, mature DCs, followed 4-6 weeks later by DCs pulsed with keyhole limpet hemocyanin (KLH), HLA-A*0201-positive ... More
High frequency of skin-homing melanocyte-specific cytotoxic T lymphocytes in autoimmune vitiligo.
AuthorsOgg GS, Rod Dunbar P, Romero P, Chen JL, Cerundolo V
JournalJ Exp Med
PubMed ID9743539
Vitiligo is an autoimmune condition characterized by loss of epidermal melanocytes. Using tetrameric complexes of human histocompatibility leukocyte antigen (HLA) class I to identify antigen-specific T cells ex vivo, we observed high frequencies of circulating MelanA-specific, A*0201-restricted cytotoxic T lymphocytes (A2-MelanA tetramer+ CTLs) in seven of nine HLA-A*0201-positive individuals with ... More
Novel roles of CpG oligodeoxynucleotides as a leader for the sampling and presentation of CpG-tagged antigen by dendritic cells.
AuthorsShirota H, Sano K, Hirasawa N, Terui T, Ohuchi K, Hattori T, Shirato K, Tamura G
JournalJ Immunol
PubMed ID11418633
Oligodeoxynucleotides containing CpG motifs have been highlighted as potent Th1 activators. We previously reported that Ag and CpG, when conjugated together, synergistically promoted the Ag-specific Th1 development and inhibited the Th2-mediated airway eosinophilia. In this study, we examined the mechanisms underlying the synergism of the covalent conjugation. The CpG-OVA conjugate ... More
Orphan nuclear receptors constitutive androstane receptor and pregnane X receptor share xenobiotic and steroid ligands.
AuthorsMoore LB, Parks DJ, Jones SA, Bledsoe RK, Consler TG, Stimmel JB, Goodwin B, Liddle C, Blanchard SG, Willson TM, Collins JL, Kliewer SA
JournalJ Biol Chem
PubMed ID10748001
Xenobiotics induce the transcription of cytochromes P450 (CYPs) 2B and 3A through the constitutive androstane receptor (CAR; NR1I3) and pregnane X receptor (PXR; NR1I2), respectively. In this report, we have systematically compared a series of xenobiotics and natural steroids for their effects on mouse and human CAR and PXR. Our ... More
Wnt/ß-catenin signalling induces MLL to create epigenetic changes in salivary gland tumours.
AuthorsWend P, Fang L, Zhu Q, Schipper JH, Loddenkemper C, Kosel F, Brinkmann V, Eckert K, Hindersin S, Holland JD, Lehr S, Kahn M, Ziebold U, Birchmeier W
Journal
PubMed ID23736260
We show that activation of Wnt/ß-catenin and attenuation of Bmp signals, by combined gain- and loss-of-function mutations of ß-catenin and Bmpr1a, respectively, results in rapidly growing, aggressive squamous cell carcinomas (SCC) in the salivary glands of mice. Tumours contain transplantable and hyperproliferative tumour propagating cells, which can be enriched by ... More
Use of a phosphotyrosine-antibody pair as a general detection method in homogeneous time-resolved fluorescence: application to human immunodeficiency viral protease.
AuthorsCummings RT, McGovern HM, Zheng S, Park YW, Hermes JD
JournalAnal Biochem
PubMed ID10094778
A homogeneous time-resolved fluorescence (HTRF) assay has been developed for human immunodeficiency viral (HIV) protease. The assay utilizes a peptide substrate, differentially labeled on either side of the scissile bond, to bring two detection components, streptavidin-cross-linked XL665 (SA/XL665) and a europium cryptate (Eu(K))-labeled antiphosphotyrosine antibody, into proximity allowing fluorescence resonance ... More
Tetramer-guided analysis of TCR beta-chain usage reveals a large repertoire of melan-A-specific CD8+ T cells in melanoma patients.
AuthorsValmori D, Dutoit V, Liénard D, Lejeune F, Speiser D, Rimoldi D, Cerundolo V, Dietrich PY, Cerottini JC, Romero P
JournalJ Immunol
PubMed ID10861093
The assessment of the TCR repertoire expressed by tumor-specific CD8+ T lymphocytes has been hampered to date by the difficulty of targeting the analysis to lymphocytes directed against a single epitope. In the present study we have used fluorescent A2/Melan-A tetramers in conjunction with anti-CD8 and anti-TCR beta-chain variable (BV) ... More
JAB1 is essential for B cell development and germinal center formation and inversely regulates Fas ligand and Bcl6 expression.
AuthorsSitte S, Gläsner J, Jellusova J, Weisel F, Panattoni M, Pardi R, Gessner A
JournalJ Immunol
PubMed ID22327073
Jun activation domain-binding protein 1 (JAB1) regulates ubiquitin-dependent protein degradation by deneddylation of cullin-based ubiquitin ligases and, therefore, plays a central role in regulating proliferation and apoptosis. Because these processes are decisive for B cell development, we investigated JAB1 functions in B cells by establishing a mouse strain with a ... More
Epithelial cell-specific laminin 5 is required for survival of early thymocytes.
AuthorsKim MG, Lee G, Lee SK, Lolkema M, Yim J, Hong SH, Schwartz RH
JournalJ Immunol
PubMed ID10861052
The gene LamC2 encoding the gamma2 chain of laminin 5, an epithelial cell-specific extracellular matrix protein, was identified in a PCR-based subtracted cDNA library from mouse thymic stromal cells. The mRNA existed in two alternative forms (5.1 and 2.4 kb). The full-length message was highly expressed in SCID thymus and ... More
Amplification of fluorescent in situ hybridisation signals in formalin fixed paraffin wax embedded sections of colon tumour using biotinylated tyramide.
AuthorsMcKay JA, Murray GI, Keith WN, McLeod HL
JournalMol Pathol
PubMed ID9536283
Fluorescent in situ hybridisation (FISH) is a powerful tool for the evaluation of chromosomal alterations in formalin fixed paraffin wax embedded sections of colorectal cancer. However, initial experiments using a two-step detection system for digoxigenin labelled chromosome specific centromeric probes resulted in a complete lack of hybridisation signal from a ... More
Single Molecule Detection Technologies in Miniaturized High Throughput Screening: Fluorescence Correlation Spectroscopy.
AuthorsMoore KJ, Turconi S, Ashman S, Ruediger M, Haupts U, Emerick V, Pope AJ
JournalJ Biomol Screen
PubMed ID10838431
Fluorescence assay technologies used for miniaturized high throughput screening are broadly divided into two classes. Macroscopic fluorescence techniques (encompassing conventional fluorescence intensity, anisotropy [also often referred to as fluorescence polarization] and energy transfer) monitor the assay volume- and time-averaged fluorescence output from the ensemble of emitting fluorophores. In contrast, single-molecule ... More
Dok-related protein negatively regulates T cell development via its RasGTPase-activating protein and Nck docking sites.
AuthorsGugasyan R, Quilici C, I ST, Grail D, Verhagen AM, Roberts A, Kitamura T, Dunn AR, Lock P
JournalJ Cell Biol
PubMed ID12093790
Downstream of kinase (Dok)-related protein (DokR, also known as p56(dok)/FRIP/Dok-R) is implicated in cytokine and immunoreceptor signaling in myeloid and T cells. Tyrosine phosphorylation induces DokR to bind the signal relay molecules, RasGTPase-activating protein (RasGAP) and Nck. Here, we have examined the function of DokR during hematopoietic development and the ... More
Recombinant adenovirus coexpressing covalent peptide/MHC class II complex and B7-1: in vitro and in vivo activation of myelin basic protein-specific T cells.
AuthorsChen J, Huber BT, Grand RJ, Li W
JournalJ Immunol
PubMed ID11466346
Previous studies have demonstrated that an MHC class II molecule with an antigenic peptide genetically fused to its beta-chain is capable of presenting this peptide to CD4(+) T cells. We hypothesized that covalent peptide/class II complex may direct the accessory molecules to exert their function specifically onto T cells in ... More
Influence of EDTA and heparin on lipopolysaccharide binding and cell activation, evaluated at single-cell level in whole blood.
AuthorsBrunialti MK, Kallás EG, Freudenberg M, Galanos C, Salomao R
JournalCytometry
PubMed ID11857593
BACKGROUND: The use of whole blood (WB) in studying lipopolysaccharide (LPS)-induced cellular activation preserves the milieu in which LPS-cell interaction occurs in vivo. However, little information is available on using such a system at a single-cell level. We evaluated LPS binding and cell activation in WB by using flow cytometry. ... More
An antigen-targeted approach to adoptive transfer therapy of cancer.
Previous attempts to treat human malignancies by adoptive transfer of tumor-specific CTLs have been limited by the difficulty of isolating T cells of defined antigen specificity. The recent development of MHC class I/antigenic peptide tetrameric complexes that allow direct identification of antigen-specific T cells has opened new possibilities for the ... More
A second uniquely human mutation affecting sialic acid biology.
AuthorsAngata T, Varki NM, Varki A
JournalJ Biol Chem
PubMed ID11546777
Siglecs are immunoglobulin superfamily member lectins that selectively recognize different types and linkages of sialic acids, which are major components of cell surface and secreted glycoconjugates. We report here a human Siglec-like molecule (Siglec-L1) that lacks a conserved arginine residue known to be essential for optimal sialic acid recognition by ... More
Distinct BMI-1 and EZH2 expression patterns in thymocytes and mature T cells suggest a role for Polycomb genes in human T cell differentiation.
AuthorsRaaphorst FM, Otte AP, van Kemenade FJ, Blokzijl T, Fieret E, Hamer KM, Satijn DP, Meijer CJ
JournalJ Immunol
PubMed ID11342607
BMI-1 and EZH2 Polycomb-group (PcG) proteins belong to two distinct protein complexes involved in the regulation of hematopoiesis. Using unique PcG-specific antisera and triple immunofluorescence, we found that mature resting peripheral T cells expressed BMI-1, whereas dividing blasts were EZH2(+). By contrast, subcapsular immature double-negative (DN) (CD4(-)/CD8(-)) T cells in ... More
Polyethylene glycol-modified GM-CSF expands CD11b(high)CD11c(high) but notCD11b(low)CD11c(high) murine dendritic cells in vivo: a comparative analysis with Flt3 ligand.
AuthorsDaro E, Pulendran B, Brasel K, Teepe M, Pettit D, Lynch DH, Vremec D, Robb L, Shortman K, McKenna HJ, Maliszewski CR, Maraskovsky E
JournalJ Immunol
PubMed ID10861034
Dendritic cells (DC) are potent APCs that can be characterized in the murine spleen as CD11b(high)CD11c(high) or CD11b(low)CD11c(high). Daily injection of mice of Flt3 ligand (FL) into mice transiently expands both subsets of DC in vivo, but the effect of administration of GM-CSF on the expansion of DC in vivo ... More
High frequencies of naive Melan-A/MART-1-specific CD8(+) T cells in a large proportion of human histocompatibility leukocyte antigen (HLA)-A2 individuals.
AuthorsPittet MJ, Valmori D, Dunbar PR, Speiser DE, Liénard D, Lejeune F, Fleischhauer K, Cerundolo V, Cerottini JC, Romero P
JournalJ Exp Med
PubMed ID10477554
Using fluorescent HLA-A*0201 tetramers containing the immunodominant Melan-A/MART-1 (Melan-A) tumor-associated antigen (Ag), we previously observed that metastatic lymph nodes of melanoma patients contain high numbers of Ag-experienced Melan-A-specific cytolytic T lymphocytes (CTLs). In this paper, we enumerated and characterized ex vivo Melan-A-specific cells in peripheral blood samples from both melanoma ... More
Characterization of a novel proapoptotic caspase-2- and caspase-9-binding protein.
AuthorsBonfoco E, Li E, Kolbinger F, Cooper NR
JournalJ Biol Chem
PubMed ID11350957
Caspases play important roles in regulating apoptotic signaling pathways. Here we report the cloning, by the yeast two hybrid system with dominant negative caspase-2 as "bait," of a proapoptotic molecule named proapoptotic caspase adaptor protein (PACAP), encoded by a 372-base pair open reading frame. Binding of this novel protein to ... More
The rearranged V(H) domain of a physiologically selected anti-single-stranded DNA antibody as a precursor for formation of IgM and IgG antibodies to diverse antigens.
It has been proposed that autoreactivity of modest affinity contributes to positive selection of a preimmunization B cell repertoire, whereas high-affinity autoreactivity leads to negative selection. This hypothesis predicts that a B cell producing a physiologically selected unmutated ssDNA-binding Ab should be a precursor of cells that respond to diverse ... More
Multiplexed fluorescent bead-based immunoassays for quantitation of human cytokines in serum and culture supernatants.
AuthorsKellar KL, Kalwar RR, Dubois KA, Crouse D, Chafin WD, Kane BE
JournalCytometry
PubMed ID11598944
BACKGROUND: An increasing volume of data suggests a relationship between cytokine levels in human body fluids and disease pathogenesis. Traditionally, many individual assays would be performed to measure the large number of known cytokines and determine their associations with disease. A new technique for the simultaneous measurement of multiple cytokines ... More
Extrafollicular plasmablast B cells play a key role in carrying retroviral infection to peripheral organs.
AuthorsFinke D, Baribaud F, Diggelmann H, Acha-Orbea H
JournalJ Immunol
PubMed ID11342650
B cells can either differentiate in germinal centers or in extrafollicular compartments of secondary lymphoid organs. Here we show the migration properties of B cells after differentiation in murine peripheral lymph node infected with mouse mammary tumor virus. Naive B cells become activated, infected, and carry integrated retroviral DNA sequences. ... More
Flow cytometric quantification of competitive reverse transcription-PCR products.
AuthorsWedemeyer N, Pötter T, Wetzlich S, Göhde W
JournalClin Chem
PubMed ID12194914
BACKGROUND: Competitive PCR of reverse transcribed mRNA sequences is used to quantify transcripts, but the usual approaches are labor-intensive and time-consuming. We describe the non-gel-based quantification of competitive reverse transcription (RT)-PCR products with use of microparticles and flow cytometry. METHODS: PCR products of a target sequence and an internal control ... More
Functional competency of T cell antigen receptors in human thymus.
AuthorsWeiss A, Dazin PF, Shields R, Fu SM, Lanier LL
JournalJ Immunol
PubMed ID3500209
The T cell antigen receptor is likely to play a role in both positive and negative selection in the thymus. Three populations of thymocytes can be distinguished by the level of expression of the CD3-alpha/beta-chain heterodimer of the T cell antigen receptor (CD3/Ti alpha/beta) complex. Cells which fail to express ... More
Distinction between HLA class I-positive and -negative cervical tumor subpopulations by multiparameter DNA flow cytometry.
AuthorsCorver WE, Koopman LA, Mulder A, Cornelisse CJ, Fleuren GJ
JournalCytometry
PubMed ID10942899
BACKGROUND: The study of the molecular-genetic basis of heterogeneity of HLA class I expression in solid tumors is hampered by the lack of reliable rapid cell-by-cell isolation techniques. Hence, we studied the applicability of a flow cytometric approach (Corver et al.: Cytometry 2000;39;96-107). METHODS: Cells were isolated from five fresh ... More
Regulation of NK1.1 expression during lineage commitment of progenitor thymocytes.
AuthorsCarlyle JR, Zúñiga-Pflücker JC
JournalJ Immunol
PubMed ID9862680
We recently identified a stage in fetal ontogeny (NK1.1+/CD117+) that defines committed progenitors for T and NK lymphocytes. These cells are found in the fetal thymus as early as day 13 of gestation, but are absent in the fetal liver. Nonetheless, multipotent precursors derived from both the fetal thymus and ... More
The fate of self-reactive B cells depends primarily on the degree of antigen receptor engagement and availability of T cell help.
AuthorsFulcher DA, Lyons AB, Korn SL, Cook MC, Koleda C, Parish C, Fazekas de St Groth B, Basten A
JournalJ Exp Med
PubMed ID8642340
Self-reactive B cells from tolerant double-transgenic (Dbl-Tg) mice coexpressing hen egg lysozyme (HEL) and rearranged anti-HEL immunoglobulin genes have a relatively short life span when compared to normal B cells, irrespective of whether they are exposed to antigen in multivalent membrane-bound form (mHEL-Dbl-Tg mice) or soluble form (sHEL-Dbl-Tg mice). The ... More
Analysis of the cytolytic T lymphocyte response of melanoma patients to the naturally HLA-A*0201-associated tyrosinase peptide 368-376.
The human tyrosinase gene codes for two distinct antigens that are recognized by HLA-A*0201-restricted CTLs. For one of them, tyrosinase peptide 368-376, the sequence identified by mass spectrometry in melanoma cell eluates differs from the gene-encoded sequence as a result of posttranslational modification of amino acid residue 370 (asparagine to ... More
Specificity of CTL interactions with peptide-MHC class I tetrameric complexes is temperature dependent.
AuthorsWhelan JA, Dunbar PR, Price DA, Purbhoo MA, Lechner F, Ogg GS, Griffiths G, Phillips RE, Cerundolo V, Sewell AK
JournalJ Immunol
PubMed ID10510374
Tetrameric peptide-MHC class I complexes ("tetramers") are proving invaluable as reagents for characterizing immune responses involving CTLs. However, because the TCR can exhibit a degree of promiscuity for binding peptide-MHC class I ligands, there is potential for cross-reactivity. Recent reports showing that the TCR/peptide-MHC interaction is dramatically dependent upon temperature ... More
Functional characterization of CD8(+) antigen-specific cytotoxic T lymphocytes after enrichment based on cytokine secretion: comparison with the MHC-tetramer technology.
AuthorsOelke M, Kurokawa T, Hentrich I, Behringer D, Cerundolo V, Lindemann A, Mackensen A
JournalScand J Immunol
PubMed ID11119258
Cell therapy with antigen-specific T cells holds promise for various diseases including cancer and viral infections. The powerful enrichment procedure based on major histocompatibility complex (MHC)-tetramers, however, is of limited applicability so far. Therefore, the recently developed cell surface affinity matrix technology that allows direct identification and enrichment of life ... More
Paralemmin-1 is expressed in lymphatic endothelial cells and modulates cell migration, cell maturation and tumor lymphangiogenesis.
AuthorsAlbrecht I, Bieri R, Leu A, Granacher P, Hagmann J, Kilimann MW, Christofori G
Journal
PubMed ID23709172
The lymphatic system, the network of lymphatic vessels and lymphoid organs, maintains the body fluid balance and ensures the immunological surveillance of the body. In the adult organism, the de novo formation of lymphatic vessels is mainly observed in pathological conditions. In contrast to the molecular mechanisms governing the generation ... More
A strategy for multiple immunophenotyping by image cytometry: model studies using latex microbeads labeled with seven streptavidin-bound fluorochromes.
AuthorsGothot A, Grosdent JC, Paulus JM
JournalCytometry
PubMed ID8800554
Multiple immunophenotyping is aimed at identifying several cell populations in a single labeling procedure by their ability to bind combinations of specific labeled antibodies. The present work demonstrates the simultaneous discrimination by using image cytometry of aminomethylcoumarin acetate (AMCA), Lucifer yellow (LY), fluorescein isothiocyanate (FITC), R-phycoerythrin (PE), PE-Texas red tandem ... More
Direct isolation, phenotyping and cloning of low-frequency antigen-specific cytotoxic T lymphocytes from peripheral blood.
AuthorsDunbar PR, Ogg GS, Chen J, Rust N, van der Bruggen P, Cerundolo V
JournalCurr Biol
PubMed ID9545200
Cytotoxic T lymphocytes (CTLs) play an important role in controlling viral infections and certain tumours, but characterising specific CTL responses has always been technically limited. Fluorogenic 'tetramers' of major histocompatibility complex (MHC) class I complexes have been exploited recently to quantify the massive expansion of specific CTLs in human immunodeficiency ... More
Functional association of CD9 with the Fc gamma receptors in macrophages.
AuthorsKaji K, Takeshita S, Miyake K, Takai T, Kudo A
JournalJ Immunol
PubMed ID11207280
CD9, a member of the tetraspan family of proteins, is highly expressed on macrophages. Although a clear function for the molecule has yet to be described, we have found that the anti-CD9 mAb activates mouse macrophages. The rat anti-CD9 mAb, KMC8.8, but not the F(ab')(2), induced tyrosine phosphorylation of proteins ... More