Abstract：
:We describe a simple, rapid and reproducible in vitro culture system in which human peripheral blood lymphocytes (PBLs), donated 24 h prior to initiation of culture can be stimulated to produce antigen-specific antibodies. Peripheral blood lymphocytes purified by Ficoll-Hypaque centrifugation were passed over a G10 Sephadex column and then activated in vitro in the presence of 0.003% staphylococcus Cowan A, 2.8 x 10(-6) M indomethacin and appropriate concentrations of tetanus toxoid antigen. After the first 24 h in culture, a five-fold concentrated supernatant from an allogeneic mixed lymphocyte culture was added. The cell surface phenotypes of the PBLs were analyzed by flow cytometry at the initiation and termination of culture, in order to provide a comprehensive characterization of the cellular composition of a successful in vitro stimulation system. Our results clearly show that the majority of peripheral blood B cells can be induced to an activated stage (blast transformation) and interleukin 2 (IL-2) receptor expression, following very simple manipulations of the lymphoid population. Tetanus toxoid-specific antibody production can be readily generated in this cell population. In contrast, T cells were not activated to express IL-2 receptors and reach blast transformation, and did not show appreciable proliferation. Our system provides a population of B cells producing antibodies of desired specificity which could be utilized for the generation of human hybridomas or could serve as a donor population for antibody engineering via the combinatorial library approach. Careful light scattering and cell surface phenotypic analyses of the cells entering, proliferating and differentiating in these cultures enabled several novel observations to be made.
journal_name
J Immunol Methodsjournal_title
Journal of immunological methodsauthors
Owen JA,Muirhead K,Jensen C,Jonak ZLdoi
10.1016/0022-1759(95)00256-1subject
Has Abstractpub_date
1996-03-28 00:00:00pages
39-49issue
1eissn
0022-1759issn
1872-7905pii
0022175995002561journal_volume
190pub_type
杂志文章abstract：:A one-step immunospecific affinity chromatographic method for purification of mouse alpha-fetoprotein (AFP) directly from amniotic fluid is described. The procedure employed a rabbit anti-AFP immunoglobulin-G entrapped in a polyacrylamide gel matrix prepared as a gel slurry. The presence of impurities could not be dem...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(79)90146-7
更新日期：1979-01-01 00:00:00
abstract：:Apoptosis and primary necrosis are the two modes of cell death induced by a lethal injury. The majority of structural and biochemical events occurring during cell death can be analysed by flow cytometry. The 7-aminoactinomycin D (7-AAD) assay can be used to detect the loss of membrane integrity during apoptosis of mur...
journal_title：Journal of immunological methods
pub_type： 杂志文章,评审
doi：10.1016/s0022-1759(02)00072-8
更新日期：2002-07-01 00:00:00
abstract：:A simple and specific enzyme-linked immunosorbent assay (ELISA) has been developed to detect circulating IgG and IgM anti-idiotypic antibodies directed against anti-HBs molecules using 96-well polyvinyl microtitre plates as the solid phase and HRPO-labelled goat anti-HBs as conjugate. Anti-idiotype reactions were obse...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(87)90316-4
更新日期：1987-02-11 00:00:00
abstract：:A simple and reproducible technique for murine platelet isolation is described. Such preparations then become useful in determining in vitro platelet secretory responses and platelet aggregation. ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(80)90226-4
更新日期：1980-01-01 00:00:00
abstract：:Small haptens such as methylamphetamine (MW 149) cannot, on their own, induce an immune response. It is also unlikely that they fill the binding site of any antibody that recognises them. Under such circumstances any attached label might be expected to enter the area of the binding site and exert an influence on overa...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(91)90375-p
更新日期：1991-07-05 00:00:00
abstract：:Macrophages can be separated from other cell types by their ability to readily attach and spread on glass or on plastic surfaces which are treated for optimal growth of cultured cells (tissue culture-treated plastic). To detach macrophages from these surfaces, techniques must be used which require prior preparation of...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(84)90142-x
更新日期：1984-03-30 00:00:00
abstract：:Goat anti-mouse antibodies conjugated with colloidal 40 nm gold particles (G40) were used as secondary layers to stain human T lymphocytes in an attempt to extend the polarized light epi-illumination microscopic technique to flow cytometric multiparameter analysis. G40-labelled T cells were further stained with phycoe...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(87)90211-0
更新日期：1987-07-16 00:00:00
abstract：:A new quantitative assay for migration inhibitory factor (MIF) employs 3H-labelled cultured mouse or human lymphoid cells migrating from capillary tubes. Capillaries filled with labelled cells are placed in liquid scintillation counting vials, along with the MIF-containing sample and are removed at the end of a five-h...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(78)90258-2
更新日期：1978-01-01 00:00:00
abstract：:Dispersed fluorocarbon oil droplets were used as the antigen-carrier particle in an immunological agglutination assay. The extent of agglutination was found to vary with the percentage of antigen to that of inert protein used in the solution to coat and stabilize the oil droplets. Maximum sensitivity was observed when...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(86)90533-8
更新日期：1986-03-13 00:00:00
abstract：:The presence of KIR4.1 antibodies has been proposed to be a characteristic of Multiple Sclerosis (MS). This could have a significant impact on disease management. However, the validation of the initial findings has failed till date. Conflicting results have been attributed to difficulties in isolating the lower-glycos...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2017.03.008
更新日期：2017-06-01 00:00:00
abstract：:A microscale ELISA system, designated as the 'C1q-micro ELISA,' has been developed for the measurement of circulating immune complexes. The sensitivity of the method ranged between 0.1 and 3 microgram equivalents of aggregated human gamma-globulins. The procedure requires small quantities of serum and reagents, and it...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(82)90308-8
更新日期：1982-01-01 00:00:00
abstract：:In developing countries the acquisition and use of radioisotopes with their short shelf lives poses enormous problems. The objective of this study was to develop a simple, sensitive, non-isotopic method for the measurement of TSH in human serum. A conventional sandwich enzyme-linked immunosorbent assay (ELISA) techniq...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(88)90315-8
更新日期：1988-11-25 00:00:00
abstract：:It is known that membranoproliferative glomerulonephritis (MPGN), hypocomplementaemia and C3 nephritic factor (C3NeF) are closely related to each other, and the presence or absence of C3NeF in the serum is important for evaluating the nature of MPGN. However, some difficulties have been encountered in detecting this f...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(90)90234-m
更新日期：1990-07-20 00:00:00
abstract：INTRODUCTION:Peripheral blood mononuclear cells (PBMCs) are frequently used for genomic analyses, but several factors can affect the yield and integrity of nucleic acids, including the methods of cell collection and isolation. The goal of this work was to analyze the utility of systematic isolation of different immune ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2016.12.005
更新日期：2017-03-01 00:00:00
abstract：:This paper describes a novel method for following the changes in mouse tumour-infiltrating immune cell populations by repeated sampling of tumours by fine needle aspiration (FNA), followed by flow cytometry. Using this technique we were able to collect samples from P815 mouse mastocytomas, and identify and enumerate s...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2015.06.015
更新日期：2015-10-01 00:00:00
abstract：:MicroRNAs are small regulatory molecules that demonstrate useful biomarker potential. They have been recognised in biofluids, where they are protected from degradation by encapsulation into extracellular vesicles (EVs). A number of commercial products are available for the isolation of EVs and their RNA content; howev...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2015.12.011
更新日期：2016-02-01 00:00:00
abstract：:The epitope specificities of monoclonal antibodies against carcinoembryonic antigen (CEA) were determined in a solid-phase, biotin-avidin enzyme immunoassay. Constant amounts of biotin-labeled antibodies were incubated with the immobilized antigen in the presence of decreasing amounts of unlabeled antibodies. The biot...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(84)90157-1
更新日期：1984-03-30 00:00:00
abstract：:Helper and suppressor activity of T cell lines were investigated using an assay system which is based on measurement by the enzyme-linked immunosorbent assay (ELISA) of the antibody produced in vitro. T cell lines were established from spleens of BALB/c mice immunized with ovalbumin and human serum albumin. The T cell...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(83)90063-7
更新日期：1983-02-25 00:00:00
abstract：:An immunoblot (Western) assay was developed employing a species-specific monoclonal antibody to a 43 kDa Mycoplasma pneumoniae membrane polypeptide and a species-specific monoclonal antibody to 29 kDa Legionella pneumophila outer membrane protein. This assay could simultaneously detect these two different antigens dir...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(87)90291-2
更新日期：1987-11-05 00:00:00
abstract：:An alternate method was designed to couple a limited quantity of protein to an affinity support when a conventional technique was unsuccessful. This was achieved through the introduction of a small number of sulfhydryl groups to the ligand by reaction with 2-iminothiolane which resulted in a limited number of reactive...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(88)90034-8
更新日期：1988-08-09 00:00:00
abstract：:An immunomagnetic technique to detect and identify Salmonella serogroup C1 has been developed. Monoclonal antibodies (MAbs) specific for the O antigen 6,7 of Salmonella lipopolysaccharide (LPS) coupled to magnetic beads were used to isolate the salmonellae. Captured bacteria were easily identified by acridine orange s...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(91)90387-u
更新日期：1991-03-01 00:00:00
abstract：:In vitro differentiation of human monocytes (Mo) provides large amounts of mature and functionally competent macrophages (M phi) which may be used as potentially powerful anticancer agents for adoptive immunotherapy. Granulocyte macrophage-colony stimulating factor (GM-CSF) was evaluated for its ability to influence l...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(91)90046-i
更新日期：1991-10-25 00:00:00
abstract：:A method of tanned erythrocyte agglutination of wide applicability is described for the study of DNA antibodies. The practical aspects of the method have been explored in estimation of antibodies to DNA in human cases of systemic lupus erythematosus, discoid lupus erythematosus, rheumatoid arthritis, other collagenose...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(78)90067-4
更新日期：1978-01-01 00:00:00
abstract：:The use of cells as a cryopreserved, readily available reagent has facilitated high-throughput screening of new drug candidates by bioassay. This practice is considerably less labor intensive and allows more flexibility in laboratory testing than traditional continuous cell culture. We have shown that this practice ca...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2011.05.009
更新日期：2011-07-29 00:00:00
abstract：:To develop a stable immuno-assay system for quantification of human hemoglobin (Hb), the interaction between various antibodies and Hb was studied using a surface plasmon resonance (SPR) biosensor in the BIAcore equipment (Amersham Pharmacia Biotech) with an immobilized anti-Hb antibody sensor chip. When polyclonal an...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(00)00144-7
更新日期：2000-04-21 00:00:00
abstract：:A simple and sensitive liposome immunoassay for gentamicin was developed using the cytolytic agent, phospholipase C, instead of complement. Liposomes entrapping a fluorescent marker, calcein, were prepared by the reverse-phase evaporation method from a mixture of dimyristoylphosphatidylcholine and cholesterol (molar r...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(94)90397-2
更新日期：1994-04-15 00:00:00
abstract：:We have previously shown a polymorphism of human Fc receptors for mouse IgG1 using an EA rosette technique in which human erythrocytes sensitized with a murine IgG1 monoclonal antibody against glycophorin A acted as indicator cells. We now describe a method to quantitate this EA rosetting using the pseudoperoxidase ac...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(87)90223-7
更新日期：1987-07-16 00:00:00
abstract：:Antibody-dependent cell-mediated cytotoxicity (ADCC) is an important mechanism of action implicated in the clinical efficacy of several therapeutic antibodies. In vitro ADCC assays employing effector cells capable of inducing lysis of target cells bound by antibodies are routinely performed to support the research and...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2014.03.021
更新日期：2014-05-01 00:00:00
abstract：:The detection of site-directed anti-idiotypic antibodies is usually based on their ability to inhibit the binding of antigen to idiotype, in either solid- or fluid-phase radioimmunoassays. Passage of serum over antigen-coupled immunosorbents for the purpose of removing the idiotypes from complexes with putative anti-i...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(87)90172-4
更新日期：1987-06-26 00:00:00
abstract：:A method for quantitative RT-PCR using ELISA detection was developed and applied to the quantitation of IL-1 beta mRNA in clinical samples. To compensate for the 'tube effect' of RT-PCR, a synthetic RNA, pRSET RNA, was added to sample solutions as an internal standard and co-amplified with IL-1 beta mRNA. Sense primer...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(94)00321-m
更新日期：1995-03-27 00:00:00
