Abstract：
:Application of antibody phage display to the identification of cell surface antigens with restricted expression patterns is often complicated by the inability to demonstrate specific binding to a certain cell type. The specificity of an antibody can only be properly assessed when the antibody is of sufficient high affinity to detect low-density antigens on cell surfaces. Therefore, a robust and simple assay for the prediction of relative antibody affinities was developed and compared to data obtained using surface plasmon resonance (SPR) technology. A panel of eight anti-CD46 antibody fragments with different affinities was selected from phage display libraries and reformatted into complete human IgG1 molecules. SPR was used to determine K(D) values for these antibodies. The association and dissociation of the antibodies for binding to CD46 expressed on cell surfaces were analysed using FACS-based assays. We show that ranking of the antibodies based on FACS data correlates well with ranking based on K(D) values as measured by SPR and can therefore be used to discriminate between high- and low-affinity antibodies. Finally, we show that a low-affinity antibody may only detect high expression levels of a surface marker while failing to detect lower expression levels of this molecule, which may lead to a false interpretation of antibody specificity.
journal_name
J Immunol Methodsjournal_title
Journal of immunological methodsauthors
Geuijen CA,Clijsters-van der Horst M,Cox F,Rood PM,Throsby M,Jongeneelen MA,Backus HH,van Deventer E,Kruisbeek AM,Goudsmit J,de Kruif Jdoi
10.1016/j.jim.2005.04.016keywords:
subject
Has Abstractpub_date
2005-07-01 00:00:00pages
68-77issue
1-2eissn
0022-1759issn
1872-7905pii
S0022-1759(05)00126-2journal_volume
302pub_type
杂志文章abstract：:Chinese hamster lymphocyte cultures were supplemented with foetal calf serum (FCS) or hose serum (HS). Addition of 2-mercaptoethanol (2-ME) to cultures supplemented with these sera resulted in a marked increase lymphocyte proliferation. The PHA concentrations necessary for optimal lymphocyte stimulation were much lowe...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(82)90299-x
更新日期：1982-01-01 00:00:00
abstract：:Serologically and biochemically pure HLA antigens and other beta 2-microglobulin-associated proteins were isolated using a rapid immunoprecipitation technique with a monoclonal anti-beta 2-microglobulin reagent A88. Elution of the immunoprecipitate with beta 2-microglobulin isolated from kidney patients proved to be e...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(81)90237-4
更新日期：1981-01-01 00:00:00
abstract：:Fungi represent emerging infectious threats to human populations worldwide. Mice and other laboratory animals have proved invaluable in modeling clinical syndromes associated with superficial and life-threatening invasive mycoses. This review outlines salient features of common vertebrate animal model systems to study...
journal_title：Journal of immunological methods
pub_type： 杂志文章,评审
doi：10.1016/j.jim.2014.03.022
更新日期：2014-08-01 00:00:00
abstract：:A limiting dilution method for the efficient transformation by Epstein-Barr virus (EBV) of human B lymphocytes has been applied to the production of human monoclonal antibodies to ovarian cancer-associated antigens. Limited numbers (e.g., 2 X 10(5)) of EBV-infected B lymphocytes from ovarian cancer patient spleen, lym...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(87)90274-2
更新日期：1987-12-24 00:00:00
abstract：:The U937 cell line has been shown to generate tumour necrosis factor alpha (TNF-alpha) in response to soluble stimuli such as PMA and LPS, but only after treatment with GM-CSF. We report here the generation of TNF-alpha from U937 cells following phagocytosis of opsonised zymozan particles without the need for pre-trea...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(92)90141-f
更新日期：1992-08-10 00:00:00
abstract：:In this study, we developed a one-step, washing-free and amplification-free assay for protein analysis using gold nanoparticle probes (GNPs) and dynamic light scattering (DLS) technique. The target protein concentration was determined by analyzing the level of GNP aggregation caused by antibody-antigen interactions us...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2009.07.015
更新日期：2009-09-30 00:00:00
abstract：:In this paper, we describe a fluorimeter-based, closed-tube, polymerase chain reaction (PCR) assay for the detection and quantification of the mRNA of porcine interleukin 1alpha (IL1alpha) and interleukin 2 (IL2) cytokines in peripheral blood leukocytes (PBLs) using melting curve analysis and compare it to a standard ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(99)00114-3
更新日期：1999-11-19 00:00:00
abstract：:The EuroFlow Consortium developed a fully standardized flow cytometric approach from instrument settings, through antibody panel, reagents and sample preparation protocols, to data acquisition and analysis. The Swiss Cytometry Society (SCS) promoted a study to evaluate the feasibility of using such standardized measur...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2017.07.013
更新日期：2019-12-01 00:00:00
abstract：:The production and characterisation of 17 monoclonal antibodies to human parathyroid hormone-related protein (PTH-rP) 1-34 is described. Five of the antibodies were shown to be of high avidity (Ka 4 X 10(10)-1.9 X 10(11) L/M) and able to detect 15-100 pg PTH-rP 1-34 per tube by RIA. None cross-reacted with PTH 1-34, a...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(90)90346-w
更新日期：1990-02-20 00:00:00
abstract：:Monoclonal antibodies (mAbs) have emerged as a promising tool for cancer therapy. Differing approaches utilize mAbs to either deliver a drug to the tumor cells or to modulate the host's immune system to mediate tumor kill. The rate by which a therapeutic antibody is being internalized by tumor cells is a decisive feat...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2015.04.024
更新日期：2015-08-01 00:00:00
abstract：:The sandwich solid phase radioimmunoassay procedure has been adapted to the serological diagnosis of tuberculosis by the use of disposable plastic haemagglutination trays in which the wells were sensitized with soluble mycobacterial antigen and the subsequent uptake of globulin from test sera detected with radioiodina...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(75)90069-1
更新日期：1975-01-01 00:00:00
abstract：:Humans exposed to rabies virus must be promptly treated by passive immunization with anti-rabies antibody and active immunization with rabies vaccine. Currently, antibody prepared from pooled human serum or from immunized horses is utilized. However, neither of these reagents are readily available, entirely safe, or c...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(01)00353-2
更新日期：2001-06-01 00:00:00
abstract：:Human peripheral blood lymphocytes were stained simultaneously with the lipophilic membrane label PKH-2 and surface markers and analysed by multiparameter flow cytometry. A biphasic staining distribution was observed for T (CD3+) as well as B (CD19+) cells. Within the T subset, CD4+ cells stained almost exclusively wi...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(92)90211-b
更新日期：1992-09-18 00:00:00
abstract：:Human vascular endocan is a proteoglycan exhibiting tumorigenic activity through both its glycan and protein cores. Endocan mRNA is identified as being one of the most significant molecular signatures defining a poor prognosis in lung, breast, kidney, prostate, and thyroid malignancies. The survival inversely correlat...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2012.02.009
更新日期：2012-04-30 00:00:00
abstract：:Within EuroFlow, we recently developed screening tubes for hematological malignancies and immune deficiencies. Pipetting of antibodies for such 8-color 12-marker tubes however is time-consuming and prone to operational mistakes. We therefore evaluated dried formats of the lymphocytosis screening tube (LST) and of the ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2017.03.011
更新日期：2019-12-01 00:00:00
abstract：:HIV-1 negative regulatory factor (Nef) can inhibit CTL recognition by downregulation of HLA-A and HLA-B on the cell surface. In contrast, HLA-C is not affected by Nef and a growing number of studies demonstrate an important role of HLA-C for the control of HIV-1. So far, only a limited number of HLA-C restricted CTL e...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2015.01.005
更新日期：2015-03-01 00:00:00
abstract：:Historically, tumour markers are substances selectively released by tumour cells in the blood stream so that they can be detected in the serum or other body fluids for clinical monitoring of various malignancies. The term is now extended to cell or tissue characteristics, such as cytogenetic markers, oncogenes or abno...
journal_title：Journal of immunological methods
pub_type： 杂志文章,评审
doi：10.1016/0022-1759(92)90072-2
更新日期：1992-06-24 00:00:00
abstract：:Labeling with [35S]methionine at a high specific activity is essential to the facile preparation of 2-dimensional gel electrophoretograms with the analytical 2-dimensional charge-size separation procedure (Anderson's ISODALT system). Mitogen-activated T and B lymphocytes subjected to low methionine concentrations woul...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(86)90051-7
更新日期：1986-04-03 00:00:00
abstract：:Antigen receptor-mediated activation of T and B lymphocytes results in activation of phospholipase C-gamma isozymes with subsequent hydrolysis of membrane inositol phospholipids. As a method of screening autoimmune or immunodeficient patients for early receptor signaling defects, we have developed a rapid technique fo...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(95)00135-w
更新日期：1995-10-12 00:00:00
abstract：:In this study the exposure period of the lymphocyte-myeloma cell mixture to the fusogen was evaluated for its influence upon the yield of total hybridoma colonies and those which secreted monoclonal antibodies. Sp2/0 and FOX-NY myeloma cells were fused for varying periods with murine splenic lymphocytes immunized with...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(85)90207-8
更新日期：1985-08-02 00:00:00
abstract：:Receptor-mediated antibody internalization is a key mechanism underlying several anti-cancer antibody therapeutics. Delivering highly toxic drugs to cancer cells, as in the case of antibody drug conjugates (ADCs), efficient removal of surface receptors from cancer cells and changing the pharmacokinetics profile of the...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2016.02.001
更新日期：2016-04-01 00:00:00
abstract：:We developed a reliable method for reconstituting thymectomized, lethally irradiated guinea pigs. Injection of 2.5-10 X 10(7) syngeneic bone marrow cells into adult thymectomized, lethally irradiated guinea pigs produced survival of 46-100% of treated animals. Gentamycin sulfate (5 mg/kg of body weight) for 10 days wa...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(84)90187-x
更新日期：1984-05-25 00:00:00
abstract：:High-titered antisera against human melanoma- and leukemia-associated antigens have been produced in non-human primates. The data from this and from other studies suggest that this model may prove a valuable source of highly specific antisera against a variety of human tumor-associated antigens or alloantigens. Our me...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(76)90048-x
更新日期：1976-01-01 00:00:00
abstract：:Peritoneal eosinophilia frequently occurs in patients undergoing peritoneal dialysis. We have devised a method for isolating large numbers of these peritoneal eosinophils from pediatric patients on continuous peritoneal dialysis. Patients were selected on the basis of previous high peritoneal eosinophil counts and had...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(90)90152-l
更新日期：1990-02-09 00:00:00
abstract：:Ferromagnetic iron dextran particles were prepared by reacting a mixture of ferrous chloride and ferric chloride with dextran polymers under alkaline conditions. Particles purified by gel filtration chromatography were in the size range of 30-40 nm, had an electron dense core of about 15 nm, were stable against aggreg...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(82)90007-2
更新日期：1982-08-13 00:00:00
abstract：:A microELISA for the estimation of human serum thymosin alpha 1 is described. In this assay, antibody to thymosin alpha 1 is pre-incubated with the standard or serum at 4 degrees C. Unbound antibody in the liquid-phase then binds with solid-phase thymosin alpha 1. The method is sufficiently sensitive for measuring ser...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(85)90163-2
更新日期：1985-06-12 00:00:00
abstract：:While a great deal has been learned about the genetic control of B lymphocyte behavior through the observation of primary B cells from mice bearing targeted gene mutations, such studies can be restricted by the limited number and longevity of the cells ex vivo, by their heterogeneity, and by the inability to apply fur...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(99)00094-0
更新日期：1999-08-31 00:00:00
abstract：:The oxidative burst generation capacity of granulocytes can be reliably detected by a flow cytometric procedure using lysed whole blood, dihydrorhodamine 123 (DHR), and phorbol myristate acetate (PMA). This assay is used to detect chronic granulomatous disease (CGD) and the CGD carrier state. To assess the feasibility...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(95)00210-3
更新日期：1995-12-15 00:00:00
abstract：:To date, in vivo apoptosis within the thymus has been assessed using morphological criteria and/or detection of a DNA ladder indicative of oligonucleosomal fragmentation of the DNA. Here, we have used a fluorometric method to investigate activation of the caspase protease family in the thymus following in vivo inducti...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(99)00054-x
更新日期：1999-06-24 00:00:00
abstract：:Many labs wish to measure cytokines in an accurate, reproducible, and rapid manner. An antibody-based membrane array for measuring cytokines has been developed based on the same technology as the traditional ELISA. The aim of this study was to compare results obtained with the traditional ELISA method with those from ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2003.10.011
更新日期：2004-01-01 00:00:00
