Journal of Immunological Methods (J IMMUNOL METHODS)
The Journal of Immunological Methods is devoted to covering techniques for: (1) Quantitating and detecting antibodies and/or antigens and haptens based on antigen-antibody interactions. (2) Fractionating and purifying immunoglobulins, lymphokines and other molecules of the immune system. (3) Isolating antigens and other substances important in immunological processes. (4) Labelling antigens and antibodies with radioactive and other markers. (5) Localizing antigens and/or antibodies in tissues and cells, in vivo or in vitro. (6) Detecting, enumerating and fractionating immunocompetent cells. (7) Assaying for cellular immunity. (8) Detecting cell-surface antigens by cell-cell interactions. (9) Initiating immunity and unresponsiveness. (10) Transplanting tissues. (11) Studying items closely related to immunity such as complement, reticuloendothelial system and others. In addition the journal will publish articles on novel methods for analysing the organization, structure and expression of genes for immunologically important molecules such as immunoglobulins, T cell receptors and accessory molecules involved in antigen recognition, processing and presentation. Submitted full length manuscripts should describe new methods of broad applicability to immunology and not simply the application of an established method to a particular substance - although papers describing such applications may be considered for publication as a short Technical Note The Recombinant Technology section will contain articles relating to modification by recombinant techniques of molecules of immunological interest; isolation of novel binding proteins by phage display; gene therapy; transfection; and expression. Immunology Protocols is a section providing detailed, step-by-step descriptions of new and established techniques in immunology. Articles on the molecular biological analysis of immunologically relevant receptor binding sites are also invited.
- Impact factor2.2Show impact factor historyHide impact factor history
- WebsiteJournal of Immunological Methods website
Other titlesJournal of immunological methods, Immunological methods, JIM
Material typePeriodical, Internet resource
Document typeJournal / Magazine / Newspaper, Internet Resource
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- Pre-print can not be deposited for The Lancet
Publications in this journal
Article: Usage of standardized antigen-presenting cells improves ELISpot performance for complex protein antigens. Journal of Immunological MethodsJournal of Immunological Methods 03/2013;
Article: Human monocyte-derived dendritic cells from leukoreduction system chambers after plateletpheresis are functional in an in vitro co-culture assay with intestinal epithelial cells.: Tiscornia I, Sánchez-Martins V, Hernández A, Bollati-Fogolín M.Journal of Immunological Methods 07/2012;
Article: WISH cell line: from the antiviral system to a novel reporter gene assay to test the potency of human IFN-α and IFN-β.: Bürgi Mde L, Prieto C, Etcheverrigaray M, Kratje R, Oggero M, Bollati-Fogolín M.Journal of Immunological Methods 04/2012;
Article: Novel approach to recognition of predicted HIV-1 Gag B3501-restricted CD8 T-cell epitopes by HLA-B3501(+) patients: confirmation by quantitative ELISpot analyses and characterisation using multimers.[show abstract] [hide abstract]
ABSTRACT: Exploring the intricacies of CD8(+) T-cell epitope recognition using emerging technologies to combine assessment of affinity, phenotype and resulting polyfunctional efficacy advances our understanding of HIV-1 immunopathogenesis and disease progression. Complexities within T-cell antigen recognition, such as epitope:MHC binding, stability and affinity, appear to influence the distinction between protective and ineffective anti-HIV-1 immune responses, which are thought to govern rate of disease progression. This study utilises the novel ProImmune REVEAL and ProVE(R) technology of rapid peptide synthesis, binding and affinity assays, and pentamer synthesis in conjunction with flow cytometry and simultaneous assessment of multiple CD8(+) T-cell effector functions in response to HLA-B3501-restricted HIV-1 Gag peptides, to discover new T-cell epitopes. The predicted HLA-B3501-restricted peptides, HPVHAGPIA and YPLTSLRSL, and relevant pentamers were used in parallel to validate T-cell epitopes on clinical HIV-1(+) samples, confirming correlation between the expected superior immunogenicity of newly discovered epitopes and the ex vivo T-cell response. Such a platform should be employed in prophylactic and therapeutic vaccine settings.Journal of Immunological Methods 01/2009; 341(1-2):76-85.
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ABSTRACT: The masking effects of antigens by chemical fixation, processing, embedding media interactions, represent a serious problem for immunohistochemical purposes. Fortunately, different approaches in antigen retrieval exist. These techniques are relatively recent and continuously expanding. This review focuses on the present state of the art in antigen retrieval methods for immunohistochemistry in light and electron microscopy. Moreover, a brief discussion on the chemical aspects of fixation, mechanism of retrieval, as well as its efficacy, is given.Journal of Immunological Methods 01/2009; 341(1-2):1-18.
Article: Multiplexed Elispot assay.[show abstract] [hide abstract]
ABSTRACT: Micron scale latex beads are well established as highly biocompatible reagents. Imbibing two fluorescent dyes into the interior of the beads enables the creation of a family of combinatorially colored labels. Previous use of such beads, in flow cytometry for example, has focused on beads of approximately 5 microm diameter. We show here that 280 nm combinatorially labeled particles can be used to create ELISA-style assays in 200 microm scale virtual wells, using digital microscopy as the readout. The utility of this technique is illustrated by profiling the secreted cytokine footprints of peripheral blood mononuclear cells in a multiparametric version of the popular Elispot assay. Doing so reveals noncanonical classes of T lymphocytes. We further show that the secreting cell type can be concurrently identified by surface staining with a cell type specific antibody conjugated to the same multiplexed beads.Journal of Immunological Methods 01/2009; 341(1-2):127-34.
Article: Gene gun-supported DNA immunisation of chicken for straightforward production of poxvirus-specific IgY antibodies.[show abstract] [hide abstract]
ABSTRACT: Orthopoxviruses code for numerous immunomodulatory proteins, the structure and function of which are clarified inadequately. Antibodies constitute a potent tool to study such proteins, enabling conclusions on protein location and time course of expression. However, common antibody production in mice or rabbits requires tedious protein expression and injection, as well as blood collection at regular intervals. To simplify this procedure, IgY antibodies specific for poxviral proteins (F1L and p28) were generated by immunisation of chickens, because antibody retrieval from eggs allows the non-invasive generation of huge amounts of antibodies. The main intentions were (i) to decrease invasiveness, (ii) to immunise with native forms of proteins and (iii) to circumvent previous protein expression and purification. Therefore, chicken were immunised with DNA expression vectors coding for conserved domains of the selected proteins delivered for the first time by a gene gun. Four weeks after initial immunisation specific antibodies were found in the egg yolk as proven by immunofluorescence staining of poxvirus-infected cells. The specific IgY titre rose to 1:80,000 and was stable for more than 120 days. With this investigation we present an universal procedure for IgY design and production that can be applied for various issues in the future.Journal of Immunological Methods 01/2009; 341(1-2):146-53.
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ABSTRACT: Natural killer (NK) cells represent the first line of defense against transformed or virally infected cells. Upon triggering of activating receptors NK cells can respond by secreting cytokines such as interferon-gamma or tumor necrosis factor-alpha and by the release of cytotoxic granules, resulting in the lysis of susceptible target cells. The importance of NK cells becomes clear in patients with impaired NK cell function or development. These patients suffer from recurrent illness and have particular problems in controlling viral infections despite their functional adaptive immune response. A detailed analysis of NK cell function is therefore of great importance. Here we describe a fast and comprehensive NK cell assay. The assay is performed in whole blood samples, eliminating the need for the isolation of PBMC or pure NK cells, while still allowing for the stimulation of the samples with cytokines. In each sample the absolute NK cell number is determined. The cytolytic activity is assayed by the lysis of (51)Cr labeled target cells and by determining the externalization of CD107a in the NK cells. Furthermore, cytokine production is detected by intracellular FACS analysis. Due to the strong reduction of required material this approach utilizes less than 3.5 ml of heparinized whole blood and is particularly applicable for frequent monitoring the immune function of adult and especially of pediatric patients.Journal of Immunological Methods 01/2009; 341(1-2):154-64.
Article: A novel method for long term bone marrow culture and genetic modification of murine neutrophils via retroviral transduction.[show abstract] [hide abstract]
ABSTRACT: Neutrophils are a critical component of the innate immune response to invading microbial pathogens. However, an excessive and/or prolonged neutrophil response can result in tissue injury that is thought to underlie the pathogenesis of various inflammatory diseases. The development of novel therapeutic strategies for inflammatory diseases depends on an improved understanding of regulation of neutrophil function. However, investigations into neutrophil function have been constrained in part by the difficulty of genetically modifying neutrophils using current techniques. To overcome this, we have developed a novel method for the genetic modification of murine bone marrow derived progenitor cells using retroviral transduction followed by long term bone marrow culture to generate mature neutrophils. These neutrophils are functionally mature as determined by morphology, surface marker (Gr1, CD11b, CD62L and CXCR2) expression, and functional attributes including the ability to generate superoxide, exocytose granule contents, chemotax, and phagocytose and kill bacteria. Further, the in vitro matured neutrophils are capable of migrating to an inflammatory site in vivo. We utilized this system to express the Bcl-2 transgene in mature neutrophils using the retroviral vectors pMIG and pMIT. Bcl-2 overexpression conferred a substantial delay in spontaneous apoptosis of neutrophils as assessed by annexin V and 7-amino-actinomycin D (7AAD) staining. Moreover, Bcl-2 overexpression did not alter granulopoiesis, as assessed by morphology and surface marker expression. This system enables the genetic manipulation of progenitor cells that can be differentiated in vitro to mature neutrophils that are functional in vitro and in vivo.Journal of Immunological Methods 12/2008; 340(2):102-15.
Article: An 11-color flow cytometric assay for identifying, phenotyping, and assessing endocytic ability of peripheral blood dendritic cell subsets in a single platform.[show abstract] [hide abstract]
ABSTRACT: Human peripheral blood dendritic cells (PBDC) are a rare population comprised of several distinctive subsets. Analysis of these cells has been hindered by their low frequency. In this study, we report a novel direct ex vivo 11-color flow cytometric assay that combines subset identification with analysis of activation status and endocytic ability of three major PBDC subsets (CD1c(+)CD11c(+) "MDC1," CD141(+)CD11c(+) "MDC2," and CD303(+)CD11c(-) "PDC") within a single platform. This method eliminates the need for DC enrichment, isolation, or prolonged culture. Human peripheral blood mononuclear cells (PBMC) from healthy donors are incubated with FITC-dextran directly ex vivo, prior to cell surface staining with various markers. As expected, PBDC identified by this assay express low levels of CD40 and CD86 directly ex vivo, and significantly upregulate expression of these molecules upon stimulation with toll-like receptor ligands LPS and CpG oligonucleotides. In addition, PDC internalize FITC-labeled dextran poorly in comparison to MDC1 and MDC2 subsets. Specificity of FITC-dextran endocytosis is further verified by imaging flow cytometry. Furthermore, the combination of surface markers used in this assay reveals a previously unreported CD4(+)CD11c(+)CD303(-)CD1c(-)CD141(-) cell population. Taken together, this assay is a rapid and cost-effective method that avoids manipulation of PBDC while providing direct ex vivo high-dimensional flow cytometry data for PBDC studies.Journal of Immunological Methods 12/2008; 341(1-2):106-16.
Article: Delayed processing of blood increases the frequency of activated CD11b+ CD15+ granulocytes which inhibit T cell function.[show abstract] [hide abstract]
ABSTRACT: We tested whether granulocytes, which contaminate PBMC isolates after prolonged blood storage at room temperature, are responsible for inhibited T cell function in aged blood. We extend previous observations by characterizing these contaminating granulocytes as CD11b+ CD15+ cells comparable to activated CD11b+ CD15+ granulocytes induced by incubation of blood with FMLP. Granulocyte contamination of PBMC was observed within 6-8 h after venipuncture and room temperature storage (2.3 fold increase), and increased 11.3-fold by 24-26 h in comparison to PBMC from fresh blood. Refrigerated 22-26 hour storage of blood exacerbated granulocyte contamination (84-fold increase). In contrast, granulocyte contamination was markedly reduced if blood was diluted in RPMI-1640 medium (3.9-fold increase) or PBS (1.8-fold increase) prior to 22-26 hour room temperature storage. Granulocyte contamination significantly correlated with reduced CD3zeta chain expression, a marker of T cell dysfunction. Correspondingly, T cell proliferation following PHA stimulation was significantly decreased in PBMC with contaminating granulocytes from aged blood (77% of control) or FMLP treated blood (44% of control). Minimizing granulocyte contamination in PBMC of aged blood by cell sorting, or by reducing granulocyte activation by diluting blood in PBS prior to storage, increased CD3zeta chain expression and increased T cell proliferation following stimulation. These data indicate that granulocytes inhibit T cell function in aged blood. Therefore, preventing granulocyte activation in blood specimens is critical to maintain optimal T cell function. This may be accomplished by limiting the time from venipuncture to PBMC isolation to <8 h and may be extended to 26 h by simply diluting blood in PBS prior to room temperature storage.Journal of Immunological Methods 12/2008; 341(1-2):68-75.
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ABSTRACT: Natural Killer cells are cells of the innate immune system that are important for the recognition and clearance of virally infected cells or tumors. Examination of the development and signaling of these cells has been severely hampered due to an inability to over-express proteins in these cells. We developed a novel technique to generate NK cells in vivo, all of which express a gene of interest. IL2Rgamma(c)(-/-)/Rag2(-/-) mice do not develop NK cells due to the lack of IL15 signaling. We infected bone marrow from IL2Rgamma(c)(-/-)/Rag2(-/-) mice with a retroviral construct encoding EGFP and IL2Rgamma(c) connected by an IRES. NK cells selectively developed through expression of IL2Rgamma(c) and 100% of these NK cells were found to be EGFP(+). In order to test the utilization of this method to examine the function of biologically relevant proteins, constitutively active PI3K p110gamma and p110delta isoforms were over-expressed in this system. Constitutively active p110gamma revealed profound effects on NK cell development and function in vivo while p110delta had little effect.Journal of Immunological Methods 12/2008; 340(2):158-63.
Article: Trace level analysis of leached Protein A in bioprocess samples without interference from the large excess of rhMAb IgG.[show abstract] [hide abstract]
ABSTRACT: Resins containing immobilized Staphylococcal Protein A (PA) are widely used in the commercial purification of recombinant human monoclonal antibody (rhuMAb IgG) biotherapeutics. Therefore, a sensitive assay for leached PA is needed to ensure that PA is not present at unacceptable levels as an impurity in the final product. PA impurities are measured by an ELISA using chicken anti-PA antibodies. However, PA in the presence of IgG product forms a PA/IgG complex that interferes in the assay. In this report a multi-product PA ELISA is described, wherein the PA/IgG complex is dissociated by heating in the presence of detergents and chelators prior to the ELISA. The dissociation facilitates the accessibility of the anti-PA antibodies to bind to PA in the immunoassay. Heat is provided by a novel microwave technology which allows brief heating time and high sample throughput using a microtiter plate for sample heating. Thus, broadly applicable dissociation conditions, suitable for all 21 rhMab IgGs tested to date were identified. This approach streamlines the measurement of leached PA, allows higher sample testing throughput, facilitates application across multiple products, and facilitates assay automation. Data comparing in-process samples tested with both the former product-specific ELISA and this new multi-product assay are shown.Journal of Immunological Methods 12/2008; 341(1-2):59-67.
Article: Response surface methodology to determine optimal cytokine responses in human peripheral blood mononuclear cells after smallpox vaccination.[show abstract] [hide abstract]
ABSTRACT: Feasibility, amount of sample aliquots, processing time and cost are critical considerations for optimizing and conducting assays for large-population based studies. Well designed statistical approaches that quickly identify optimal conditions for a given assay could assist efficient completion of the laboratory assays for such studies. For example, assessment of the profile of secreted cytokines is important in understanding the immune response after vaccination. To characterize the cytokine immune response following smallpox vaccination, PBMC obtained from recently vaccinated subjects were stimulated with varying doses of live or UV-inactivated vaccinia virus and cultured for up to 8 days. In this paper, we describe a novel statistical method to identify optimal operating conditions for length in culture and virus MOI in order to measure a panel of secreted Th1, Th2, and inflammatory cytokines. This statistical method is comprised of two components. It first identifies a subset of the possible time in culture by virus MOI combinations to be studied. It then utilizes response surface analysis techniques to predict the optimal operating conditions for the measurement of each secreted cytokine. This method was applied, and the predicted optimal combinations of length in culture and virus MOI for maximum vaccinia-specific cytokine secretion were identified. The use of the response surface methodology can be applied to the optimization of other laboratory assays; especially when the number of PBMC available limits the testing of all possible combinations of parameters.Journal of Immunological Methods 12/2008; 341(1-2):97-105.
Article: Monoclonal antibody development for acrylamide-adducted human haemoglobin; a biomarker of dietary acrylamide exposure.[show abstract] [hide abstract]
ABSTRACT: The spontaneous formation of the neurotoxic carcinogen acrylamide in a wide range of cooked foods has recently been discovered, leading to dietary exposure estimates of 30.8 microg of acrylamide day(-1) for an average 77 kg human male. This is considerably higher than the European legal limit of acrylamide in drinking water, which is approximately 0.2 microg of acrylamide person(-1) day(-1). A recent study of 62,573 women over 11.3 years has observed an increased risk of postmenopausal endometrial and ovarian cancer (but not breast cancer) with increasing dietary acrylamide intake, demonstrating significant risk to human health. As individual acrylamide exposure is affected by dietary habits, cooking methods, and cigarette consumption; accurate extrapolation from estimated dietary exposure is extremely difficult. Quantifying biomarkers of acrylamide exposure therefore remains the most effective means of rapidly determining individual exposure to acrylamide, and correlating exposure with lifestyle choices. Current methodologies for the analysis of blood biomarkers of acrylamide are focused on expensive, slower chromatographic techniques such as GC and LC coupled to mass spectrometry. This paper describes the first successful development of two monoclonal antibodies specific to acrylamide-adducted haemoglobin (IC(50) of 94 ng ml(-1) and 198 ng ml(-1)), that are suitable for use in a high-throughput biomarker immunoassay to determine individual acrylamide exposure. Further development of acrylamide-haemoglobin standards with defined levels of acrylamide adduction will enable a fully quantitative assay, and allow sensitivity comparisons with alternative chromatographic methods of analysis.Journal of Immunological Methods 12/2008; 341(1-2):19-29.
Article: Generating antibodies against the native form of the human prion protein (hPrP) in wild-type animals: a comparison between DNA and protein immunizations.[show abstract] [hide abstract]
ABSTRACT: Generation of therapeutic antibodies against human proteins is hampered by the difficulty of obtaining large quantities of correctly folded immunogens when following classic immunization procedures. Here we compared several genetic immunization protocols for their potential ability to generate high levels of antibodies against proteins expressed in their native form. We chose as a model the prion protein (PrP) because it has been demonstrated that the recognition of the native conformation of PrP is an absolute prerequisite for anti-PrP antibodies to be used as therapeutic tools for prion diseases, a group of lethal neurodegenerative disorders. We designed two human PrP-DNA vectors, containing or not a stimulatory T cell epitope, which were injected into mice following four different protocols: in the naked form with or without electroporation, or protected by cationic polymers or block copolymers. For comparison, other animals received conventional injections of recombinant human PrP with Freund's adjuvant or alum. We found that genetic immunization, carried out especially through DNA electroporation and, to a lesser extent, through injection of block copolymer-protected DNA, was able to generate high amounts of antibodies recognizing native PrP as expressed on the cell surface. Conversely, protein immunizations led to very high levels of antibodies against PrP immobilized on microtiter plates, but unable to recognize the native cell membrane-bound PrP. This clearly demonstrates the usefulness of genetic immunization, when performed under well defined conditions, in raising antibodies to native proteins. These results are of interest not only in view of passive immunotherapy of prion diseases, but also, more generally, in view of generating antibodies to human membrane proteins for immunotherapeutic or immunodiagnostic purposes.Journal of Immunological Methods 12/2008; 341(1-2):41-9.
Article: A yeast display immunoprecipitation method for efficient isolation and characterization of antigens.[show abstract] [hide abstract]
ABSTRACT: Yeast antibody display has found a wide variety of applications including antibody affinity maturation, epitope mapping, and library screening. Here we report a yeast display immunoprecipitation (YDIP) technique that employs yeast cells displaying single-chain antibody fragments (scFv) on their surface as affinity capture reagents to isolate and characterize antigens. We show that displayed single-chain antibody fragments are active in a variety of detergent solutions commonly used for immunoprecipitation and that the antigen-antibody interaction can be accurately quantified by YDIP coupled with flow cytometry. The YDIP method has also been optimized so that it is compatible with commonly used protein characterization tools such as Western blotting, silver staining, and mass spectrometry. From complex protein mixtures, we have used YDIP to isolate, analyze and sequence both soluble and plasma membrane antigens using tandem mass spectrometry. In the case of the membrane antigen, YDIP coupled with tandem mass spectrometry was successful in identifying neural cell adhesion molecule (NCAM) as the antigen for an antibody previously selected as binding to the plasma membranes of brain endothelial cells. The presented method therefore has potential to facilitate antibody-antigen characterization.Journal of Immunological Methods 12/2008; 341(1-2):117-26.
Data provided are for informational purposes only. Although carefully collected, accuracy cannot be guaranteed. The impact factor represents a rough estimation of the journal's impact factor and does not reflect the actual current impact factor. Publisher conditions are provided by RoMEO. Differing provisions from the publisher's actual policy or licence agreement may be applicable.
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Ethnikon Hidryma Ereunōn (Greece)
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