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Heddy Zola

Publications and source records attributed to Heddy Zola.

At least 19 recordsLinked to original sources
PubMed2006

CD molecules 2006--human cell differentiation molecules.

The Human Leucocyte Differentiation Antigens Workshops (HLDA) have since 1984 provided a forum for the characterization and study of leucocyte surface molecules and antibodies against them. HLDA devised the CD nomenclature, which is sanctioned by IUIS. The HLDA Council reviewed and modified the objectives of HLDA in 2004, and changed the name of the organization to Human Cell Differentiation Molecules (HCDM) to reflect the broader objectives. Workshop studies under the HCDM banner proceeded during 2005 and early 2006, culminating in a meeting in May 2006. At that meeting the Council, acting as Nomenclature Committee, approved a number of new CD designations and changes to some pre-existing CD designations, which are summarized in this report.

Antigens, CD↗
PubMed2005

Expression of toll-like receptors on B lymphocytes.

Toll-like receptors (TLRs) are a family of trans-membrane receptors that play an important role in the innate immune system. Most studies examining the cellular expression of TLRs on immune cells have focussed on neutrophils, monocytes and dendritic cells, but there is little evidence of TLRs being expressed on lymphocytes. Using 3-colour flow cytometry, expression of TLR-1, TLR-2, TLR-3, TLR-4, and TLR-9 on peripheral blood lymphocyte populations was determined. Further examination of TLRs on CD5- and CD5+ CD19+ B cell subsets was performed. The binding of TLR1 and TLR9 antibodies was detected on 15-90% of resting B cells, but not on resting T-cells. The higher expression of TLR1 and TLR9 on CD5+ B cells compared to CD5- B cells may reflect the role of B1 cells in more primitive, less specific antibody responses.

Antibodies, Monoclonal↗
PubMed2005

Cross-reactivity of anti-human chemokine receptor and anti-TNF family antibodies with common marmoset (Callithrix jacchus) leukocytes.

The common marmoset (Callithrix jacchus) is a New World primate species frequently employed for immunological models of human disease. We used flow cytometry to screen a panel of new anti-human antibodies from the HLDA8 workshop to establish cross-reactivity with marmoset peripheral blood mononuclear cells. Seventy-seven antibodies were screened of which nine antibodies showed binding. Cross-reactivity of anti-human monoclonal antibodies with CC and CXC chemokine receptors CCR3, CCR6, CCR7, and CCR8 was demonstrated on untreated marmoset mononuclear cells. Stimulation of marmoset mononuclear cells with ConA and/or PMA-ionomycin resulted in an up-regulated expression of CXCR1, CXCR3, and CXCR4. The expression of TNF-family related molecules TACI and APRIL on marmoset mononuclear cells was also identified. These studies extend the range of cross-reactive antibodies to now include anti-chemokine and anti-TNF family antibodies for this important pre-clinical model species and should provide useful tools for investigation of immunological processes in marmoset monkey models.

Animals↗
PubMed2005

Development of a cluster of differentiation antibody-based protein microarray.

Protein microarrays combine aspects of DNA microarrays and ELISA for the parallel interrogation of a biological sample using a multiplex of protein biomarkers. Here we report the development of a protein microarray consisting of a subset of CD antibodies and CRP. Several preparations (culture supernatant, ascites fluid and purified Ig) of each antibody were used in a forward phase protein microarray. Microarrays were fabricated using a non-contact printer delivering 300 pL (+/-30 pL) to specific locations on polyacrylamide gel-based substrates. Following production, microarrays were blocked for non-specific binding and incubated with sera conjugated directly with Cy3. Using CRP as a control biomarker, 12 clinical samples (inflammatory conditions and controls) were interrogated using the protein microarray format and results compared to CRP measured by conventional immunoassay. The data obtained from the microarray correlated with CRP assessed by immunoassay. Subsequently CRP 'positive' samples were interrogated for CD antigen expression; which revealed CD25 and CD45RO expression in all samples. Whilst this study focussed on a subset of CD antibodies, it is anticipated that this array could be expanded to include a larger number of CD antibodies and allow screening of sera from multiple conditions in order to identify disease markers.

Antibodies↗
PubMed2005

Characterisation of the protein composition of peripheral blood mononuclear cell microsomes by SDS-PAGE and mass spectrometry.

Approximately 340 leucocyte plasma membrane proteins have been characterised by the eight Human Leucocyte Differentiation Antigen workshops held between 1982 and 2004, based primarily on their reactivity with monoclonal antibodies. The human genome is predicted to encode approximately 34,000 cDNA transcripts, of which between 15% and 20% are predicted to contain one or more transmembrane helices. We have used SDS-PAGE separation coupled with mass spectrometry-based peptide mass tag identification to identify novel plasma membrane proteins in microsome preparations prepared from mononuclear cells obtained from human peripheral blood. A total of 361 distinct proteins were identified in a single preparation, including 37 known leucocyte plasma membrane proteins, 27 potential novel plasma membrane proteins whose expression on PBMC is poorly characterised, and 51 other proteins for which the subcellular location could not be determined. Expression analysis using cDNA panels indicates that several of these novel plasma membrane proteins are differentially expressed in lymphocyte subsets. These results show that previously unidentified lymphocyte plasma membrane proteins can be identified using this approach.

Cell Membrane↗
PubMed2005

The HLDA8 blind panel: findings and conclusions.

There were over 600 antibodies submitted to HLDA8, with many of unknown specificity. Of these, 101 antibodies were selected for a blind panel study that also included 5 negative controls and 27 positive controls of known CD specificity making a total of 133 antibodies in the final panel. Of the 101 unknowns, 31 antibodies were identified during the course of this blind panel study as being specific for known molecules and included some specific for MHC class II antigens, CD45 isoforms and the Dombrock antigen. Several antibody pairs among those in the blind panel were found to have very similar staining patterns and were therefore compared by immunohistochemical and/or Western blot analyses for identity.

Antibodies↗
PubMed2005

CD molecules 2005: human cell differentiation molecules.

The immune system works through leukocytes interacting with each other, with other cells, with tissue matrices, with infectious agents, and with other antigens. These interactions are mediated by cell-surface glycoproteins and glycolipids. Antibodies against these leukocyte molecules have provided powerful tools for analysis of their structure, function, and distribution. Antibodies have been used widely in hematology, immunology, and pathology, and in research, diagnosis, and therapy. The associated CD nomenclature is commonly used when referring to leukocyte surface molecules and antibodies against them. It provides an essential classification for diagnostic and therapeutic purposes. The most recent (8th) Workshop and Conference on Human Leukocyte Differentiation Antigens (HLDA), held in Adelaide, Australia, in December 2004, allocated 95 new CD designations and made radical changes to its aims and future operational strategy in order to maintain its relevance to modern human biology and clinical practice.

Antigens, CD↗
PubMed2005

The human leucocyte differentiation antigens (HLDA) workshops: the evolving role of antibodies in research, diagnosis and therapy.

The 8th International Workshop on Human Leucocyte Differentiation Antigens (chaired by Zola H and managed by Swart B) was run over a 4-year period and culminated in a conference in December 2004. Here we review the achievements of the HLDA Workshops and provide links to information on CD molecules and antibodies against them, including the 93 new CDs assigned in the 8th Workshop. We consider what remains to be achieved (including an estimate of the number of leucocyte surface molecules still to be discovered), and how the field can best move forward.

Allergy and Immunology↗
PubMed2004

Detection of low-abundance membrane markers by immunofluorescence--a comparison of alternative high-sensitivity methods and reagents.

The analysis of membrane molecules using antibodies detected by immunofluorescence staining and flow cytometry is used widely in research and diagnostic immunology. Conventional staining techniques readily detect molecules present at concentrations of around 2000 molecules per cell, but some molecules are expressed and function at much lower abundance. We described previously a method for the detection of molecules present at 100 molecules per cell or less based on the use of phycoerythrin as the fluorophore, a three-layer amplification process, and careful selection of available reagents. In recent years, a number of new reagents, fluorophores and kits, have become available, some of them intended for high-sensitivity applications. In this paper, a number of these reagents have been compared with the published method. While some of the reagents gave variable results or high nonspecific staining in our hands, several reagents were comparable with the published method. Furthermore, the new fluorophores allow improved simultaneous detection of two low-abundance markers.

Antibodies↗
PubMed2004

Generation of murine monoclonal antibodies to Haemophilus influenzae type b capsular polysaccharide by in vivo immunization.

Haemophilus influenzae type b (Hib) is a pathogenic gram-negative bacterium associated with human disease, especially in young children. Protective immune response to Hib results from antibodies developed against the polyribosylribitol phosphate (PRP) capsular polysaccharide of the bacterium. Several investigators in the study of immune response to Hib have produced human monoclonal antibodies to PRP. Only two previous groups have reported the generation of murine anti-PRP monoclonal antibodies using either immunizing mice with inactivated Hib bacteria or a combination of in vivo and in vitro immunization of mice with PRP conjugate antigen (PRP-D). In this present study, we generated murine anti-PRP monoclonal antibody secreting hybridomas for the first time by simple in vivo immunization with PRP conjugate antigen (PRP-T). The anti-PRP antibodies from one hybridoma clone (B10) are further characterized and potential applications are discussed.

Agglutination Tests↗
PubMed2004

Characterization of colonic and mesenteric lymph node dendritic cell subpopulations in a murine adoptive transfer model of inflammatory bowel disease.

Ulcerative colitis and Crohn's disease, collectively termed inflammatory bowel diseases (IBD), are chronic inflammatory diseases of the intestine that afflict more than 4 million people worldwide. Intestinal inflammation is characterized by an abnormal mucosal immune response to normally harmless antigens in the gut flora. In Crohn's disease, the pathogenic mucosal immune response is a typical T helper (TH1) type cell response, whereas ulcerative colitis is predominantly associated with a TH2 response. We are interested in the role of dendritic cells in early immunologic events leading to T cell activation and chronic intestinal inflammation. Using a murine adoptive transfer model of IBD, we found an accumulation of dendritic cells in colon and mesenteric lymph nodes during the early stage of IBD before the appearance of epithelial lesions and tissue degradation. In situ immunostaining and flow-cytometric analysis revealed that approximately 50% of colonic dendritic cells were CD11b B220 myeloid dendritic cells and 50% expressed the CD11b B220 plasmacytoid phenotype. In corresponding mesenteric lymph nodes, approximately 16% were plasmacytoid dendritic cells. Colonic myeloid dendritic cells were shown to express the co-stimulatory molecule CD40. Both, colonic myeloid and plasmacytoid dendritic cells released interferon-alpha in situ and stimulated T cell proliferation ex vivo. Our results show that dendritic cells can mature in the intestine without migrating to mesenteric lymph nodes. Mature intestinal dendritic cells may form a nucleation site for a local T cell response and play an important role in the pathogenesis of IBD.

Animals↗
PubMed2003

Human Leucocyte Differentiation Antigen nomenclature: update on CD nomenclature. Report of IUIS/WHO Subcommittee.

The 7th International Workshop on Human Leucocyte Differentiation Antigens (HLDA7) studied a number of newly characterised molecules relevant to human leucocyte differentiation and function. The HLDA organisation, which devised and continues to maintain the CD nomenclature, is responsible, under the auspices of IUIS and WHO, for the nomenclature of all leucocyte differentiation markers. The 7th Workshop redefined a number of (principally carbohydrate) molecules, and assigned CD names to approximately 80 new molecules. This update lists, in tabular form, the redefined and newly assigned names, together with antibodies, which have been confirmed under Workshop conditions as specific for the new and redefined molecules. The major features of the cellular expression patterns are summarised, and a LocusLink accession number provided to enable the reader to access more detailed information through http://www.ncbi.nlm.nih.gov/LocusLink.

Antigens, CD↗
PubMed2003

Isolation of antigen-specific B cells.

Cell separation techniques are important in immunology. Major cell populations can be separated successfully with high purity. However, isolation of cells which are specific for particular antigens is more challenging because of the relatively small numbers of antigen-specific cells, and the lack of independent markers available to determine the purity of the isolated population. In this review, the literature describing three principal techniques used to separate antigen-specific cells has been reviewed. Particular emphasis has been placed on yield and purity; the two most important parameters of any purification method. The most promising isolation methods have used immunomagnetic sorting and multiparametric flow cytometric analysis.

Animals↗
PubMed2002

CD antigens 2002.

Explore the source record for details and available documents.

Antibodies, Monoclonal↗
PubMed2002

FcgammaRIIb expression on human germinal center B lymphocytes.

IgG antibody can specifically suppress the antibody response to antigen. This has been explained by the hypothesis that signaling through the B cell antigen receptor is negatively modulated by the co-ligation of immunoglobulin with the receptor for IgG, FcgammaRIIb. We hypothesized that inhibitory signaling through FcgammaRIIb would be counter-productive in germinal center cells undergoing selection by affinity maturation, since these cells are thought to receive a survival/proliferative signal by interacting with antigen displayed on follicular dendritic cells. We have identified and characterized a population of B lymphocytes with low/negative FcgammaRIIb expression that are present in human tonsil. Phenotypically these cells correspond to germinal center B cells and comprise both centroblast and centrocyte populations. In examining expression at the molecular level we determined that these B cells do not express detectable mRNA for FcgammaRIIb. We examined several culture conditions to induce expression of FcgammaRIIb on germinal center cells but could not determine conditions that altered expression. We then examined the functional consequence of cross-linking membrane immunoglobulin and the receptor for IgG on human B lymphocytes. Our results cast some doubt on the value of anti-IgG as a model for antigen-antibody complexes in studying human B cell regulation.

B-Lymphocyte Subsets↗
PubMed2002

An economical 20 litre bench-top fermenter.

We describe an economical 20 litre bench-top fermenter suitable for production of recombinant antibody fragments in bacterial expression systems. The bacterial culture contained within a polycarbonate carboy is mixed (400-600 rpm) and aerated (1 vessel vol./min) by a high-shear radial flow impeller mounted on a hollow stainless steel shaft, through which pressurised air is pumped. Air is dispersed as fine bubbles into the culture medium by the turbine impeller, without the need for a porous sparger. A stainless steel baffle stabilised by a gliding counterweight increases mixing. The components can easily be disassembled for cleaning and sterilisation. Temperature (range 20-37 degrees C) and pH (range 7.0-7.5) are controlled manually. Using the apparatus, it proved possible to achieve Escherichia coli cell culture densities equivalent to an optical density at 600 nm (OD(600)) of 30-32, compared with OD(600) 4-6 in shake flasks. A yield of 40 mg/litre/day of a recombinant antibody fragment was obtained with the fermenter, which was 15-fold more than the yield of 2.5mg/litre/day achieved in shake flasks. The fermenter may be particularly suited for research purposes.

Antibodies↗