Abstract：
:Proper evaluation of immunogenicity during clinical development of biotherapeutics is a major challenge to bioanalytical scientists, in part due to matrix interference in anti-drug antibody (ADA) and neutralizing antibody (NAB) assays. If not addressed, matrix interference could confound the immunogenicity assessment of a given biotherapeutic in clinical development. To support clinical development of a B cell maturation antigen (BCMA)-CD3 bispecific antibody, a cell-based NAB assay was developed as part of a tiered approach to evaluating the immunogenicity of the drug. The assay endpoint (T cell activation) was chosen based on its strong association with the mechanism of action of the drug. The BCMA-CD3 bispecific antibody activates T cells through simultaneous binding of CD3 on T cells and BCMA on target cells. In this system, T cell activation was assessed through the measurement of luciferase activity in an engineered Jurkat cell line. In the presence of NAB, the degree of T cell activation measured by the amount of luciferase activity can be reduced. During method development, soluble BCMA (sBCMA) interference in the NAB assay was apparent. The binding of sBCMA to the anti-BCMA domain of the bispecific drug led to reduced T cell activation, which caused false positive results in NAB testing. To mitigate this interference, several strategies to eliminate sBCMA were investigated. Among the procedures tested, a bead-based approach proved most effective in depleting sBCMA, while maintaining robust assay performance and achieving fit-for-purpose sensitivity. Using this sample pretreatment procedure, the NAB assay tolerated sBCMA up to 2 μg/mL, or approximately four times the estimated median sBCMA concentration in serum samples from patients with active multiple myeloma.
journal_name
J Immunol Methodsjournal_title
Journal of immunological methodsauthors
Wang Y,Luong M,Guadiz C,Zhang M,Gorovits Bdoi
10.1016/j.jim.2019.112642subject
Has Abstractpub_date
2019-11-01 00:00:00pages
112642eissn
0022-1759issn
1872-7905pii
S0022-1759(19)30207-8journal_volume
474pub_type
杂志文章abstract：:In this report the use of surface-linkage to expand the potential experimental and therapeutic applications of single chain antibody (scFv) constructs is reviewed. A strategy for the generation and functional characterization of surface-linked scFvs that bind selectively to the T-cell proteins CD3epsilon, CD28, and CD...
journal_title：Journal of immunological methods
pub_type： 杂志文章,评审
doi：10.1016/s0022-1759(00)00344-6
更新日期：2001-02-01 00:00:00
abstract：:This communication describes an automated micro-adherence inhibition assay. Tumor-specific immunity was demonstrated with B16 melanoma and MCA-38 colon adenocarcinoma, both of which are syngeneic to the same strain of mouse (C57B16/J). Abrogation of the leukocyte adherence inhibition (LAI) response of sensitized leuko...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(77)90086-2
更新日期：1977-01-01 00:00:00
abstract：:Using in vitro established tumour cell lines attempts were made to assess the suitability of tetrazolium salt reduction (MTT) assay to replace the conventional radioactive base techniques for measuring cell proliferation and cell killing. The optimum conditions for MTT loading time, concentration of MTT and the time f...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(93)90012-v
更新日期：1993-03-15 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：:Two sensitive monoclonal antibody (MAb)-based enzyme-linked immunosorbent assays (ELISAs), one for activin A (homodimer of beta A subunits) and one for activin B (homodimer of beta B subunits) in plasma have been developed. The activin A ELISA had an effective range of 0.2-50 ng/ml while the activin B ELISA's range wa...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(93)90100-l
更新日期：1993-09-27 00:00:00
abstract：:A simple technique for raising specific antiserum to the native molecule of Epstein-Barr virus (EBV)-encoded nuclear antigen-2 (EBNA-2) from Raji cells is described. The procedure involves the use of immunoblotting to identify the EBNA-2 polypeptide followed by subsequent excision from an SDS-gel and immunization of e...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(86)90543-0
更新日期：1986-03-13 00:00:00
abstract：:A simple method for the simultaneous isolation of IgG2, IgG1, IgA and IgM from porcine colostral whey is described. It makes use of DEAE-cellulose chromatography followed by repeated Sepharose 6B exclusion chromatography after the removal of fat, casein and lipoproteins, Fused rocket immunoelectrophoresis is used to s...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(77)90058-8
更新日期：1977-01-01 00:00:00
abstract：:A competitive enzyme immunoassay for rat prolactin using an immunogen and a tracer obtained by recombinant DNA technology is described. Polyclonal antibodies were raised in rabbits immunized with a purified chimeric protein consisting of rat prolactin fused with two synthetic immunoglobulin G binding domains derived f...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(94)90264-x
更新日期：1994-03-10 00:00:00
abstract：:NK cells are lymphocytes of the innate immune system that can kill target cells after activation signal-induced directional secretion of lytic granule contents. This process depends upon F-actin polymerization at the NK cell immunological synapse (NKIS), which is the dynamic organization of molecules at the interface ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2010.02.003
更新日期：2010-04-15 00:00:00
abstract：:Frozen stored human mononuclear cells were compared with non-frozen cells as responder cells in the monocyte chemotaxis assay. Frozen stored cells were found to inhibit lower background migration, resulting in greater experimental-to-control ratios. The use of frozen cells also greatly improved the speed, standardizat...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(97)90023-5
更新日期：1977-01-01 00:00:00
abstract：:Immune complexes attached to and ingested by human polymorphonuclear (PMN) cells were quantified by cytofluorometry using a fluorescence quenching assay which permits differentiation between attachment and ingestion. The fluorescence intensity decreased after ingestion as a result of the low pH in the phagolysosomes. ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(83)90340-x
更新日期：1983-05-27 00:00:00
abstract：:Assays for the analysis of antierythropoietin antibodies (anti-EPO Abs) currently suffer from a high degree of nonspecificity or are cumbersome and time consuming to perform. They are therefore not well suited for the analysis of large numbers of human sera samples, a task that has become increasingly important due to...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2004.08.017
更新日期：2004-11-01 00:00:00
abstract：:As a model for the isolation of a labile or trace protein, the purification of uteroglobin (UGL) by immunoaffinity chromatography is described. Antibody was isolated from sheep antiserum by immunoprecipitation, and coupled to divinylsulphone-activated agarose (Mini Leak). For the immunoabsorption stage rabbit uterine ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(89)90400-6
更新日期：1989-05-12 00:00:00
abstract：:In addition to their known implication in allergy studies, IgE antibodies are becoming an increasingly interesting antibody class in cancer research. However, large-scale purification of IgE antibodies still poses substantial challenges, as they cannot be purified using techniques commonly used for other immunoglobuli...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2020.112914
更新日期：2021-02-01 00:00:00
abstract：:Direct application of membrane fractions to a nitrocellulose support without previous solubilization, denaturation or fixation of the antigen permits the quantitation of antibodies binding to denaturation sensitive epitopes. In experiments reported here, this method was used to determine the rate of EGF receptor expre...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(89)90272-x
更新日期：1989-09-01 00:00:00
abstract：:An enzyme-linked immunosorbent assay (ELISA) for determination of antibodies against the zona pellucida was developed and compared with the already available indirect immunofluorescence (IIF) technique. Sera from 100 women with explained and unexplained infertility were screened for the presence of autoantibodies to t...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(85)90399-0
更新日期：1985-05-10 00:00:00
abstract：:We have recently shown that the semi-continuous cultivation of a mouse hybridoma line in spinner flasks, with a basal defined medium (BDM) devoid of serum and protein, increases the secretion of the immunoreactive monoclonal antibody (MAb) by a factor of ca. 2.4, compared to culture in flasks with serum-containing med...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(90)90446-3
更新日期：1990-05-25 00:00:00
abstract：:An inhibitor to von Willebrand factor (vWf) was detected in the plasma from two patients with histories of mild bleeding and one patient with a severe deficiency in the Factor VIII complex using a competitive enzyme-linked immunosorbent assay (ELISA) procedure. IgG antibodies from the patients' plasmas were shown to b...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(96)00209-8
更新日期：1997-01-15 00:00:00
abstract：:A technique for detecting the nitroblue tetrazolium-reducing activity of human polymorphonuclear leukocyte enzymes separated on polyacrylamide gels is reported. Slices of gel are incubated in buffer containing nitroblue tetrazolium and either NADPH or NADH. Enzymes capable of reducing nitroblue tetrazolium form a dark...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(82)90058-8
更新日期：1982-10-29 00:00:00
abstract：:An immunoperoxidase method has been developed which allows accurate and sensitive quantitation of the binding of monoclonal antibodies to cell surface antigens. Monolayers of fixed cells were prepared in wells of Terasaki micro-test plates and monoclonal antibodies bound to cell surface antigens were identified by the...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(80)90240-9
更新日期：1980-01-01 00:00:00
abstract：:Transplantation of ascitic P-815 mastocytoma cells intradermally in the ears of syngeneic DBA/2J mice allows a simple and precise quantitation of tumor growth by ear swelling. Tumor growth was retarded and then arrested in allogeneic DBA/1J mice, whereas syngeneic hosts died as a result of tumor dissemination in drain...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(82)90086-2
更新日期：1982-11-26 00:00:00
abstract：:Non-specific radioactive counts retained on the glass fiber filters from a multiple automatic sample harvester were reduced markedly by 3 simple changes in the procedure for harvesting thymidine-labeled whole blood cultured from microtiter plates. These modifications increased the sensitivity, precision and accuracy o...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(81)90225-8
更新日期：1981-01-01 00:00:00
abstract：:Phycobilisomes are cyanobacterial photosynthetic energy transfer complexes partly composed of phycobiliproteins, proteins that are widely used as conjugable fluorochromes for flow cytometry. The brightness and photostability of phycobiliproteins suggest that intact phycobilisomes could constitute even brighter probes ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(01)00367-2
更新日期：2001-08-01 00:00:00
abstract：:A study was undertaken to establish conditions of growth to increase C3b receptor synthesis on a human B lymphoblastoid cell line (Raji) by use of cell differentiation effectors. It appears that whereas two polar compounds HMBA (2mM) and Me2SO(2%) have no or little effect, 5 BrdU (30microM) and db cAMP (5 x10(-4) M) a...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(81)90135-6
更新日期：1981-01-01 00:00:00
abstract：:Immunoabsorbents for the removal of antibodies against cell membrane antigens have been prepared by a variety of techniques in common use. Cells were surface-labelled by lactoperoxidase-catalysed radioiodination, and the different immunoabsorbent preparations were compared in terms of efficiency of incorporation and s...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(78)90222-3
更新日期：1978-01-01 00:00:00
abstract：:Primary cell cultures, such as isolated epithelial cells, neuronal cells, or hepatocytes are characterized by a very low mitotic activity. Monitoring of small changes in cell numbers requires staining with a DNA-specific dye with an extremely high sensitivity and a low inter- and intraassay variability. For this purpo...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(97)00202-0
更新日期：1998-02-01 00:00:00
abstract：:A sensitive, simple and reproducible biotin-avidin amplified double antibody immunoassay to quantitate low concentrations of mouse immunoglobulins is described. The assay is a useful technique to measure trace levels of murine monoclonal antibodies in culture supernatants of hybridoma cells metabolically labeled with ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(85)90355-2
更新日期：1985-10-10 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 relative efficiency of depot (Freund's complete adjuvant (FCA) and saponin), and non-depot (Corynebacterium parvum strain P. acnes I, C. parvum strain IMI585, and levamisole) immunoadjuvants were compared in raising agglutinating and precipitating antibodies against the metacestode fluid of Taenia multiceps of she...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(83)90230-2
更新日期：1983-07-29 00:00:00
abstract：:We describe a simplified and improved proliferation assay based on a conventional ELISA system for the in vitro measurement of cellular proliferation (ELISA proliferation assay = EPA). The assay is based on a new monoclonal antibody (Ki-S3) to the proliferation-specific Ki-67-antigen and is carried out in 96-well micr...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(97)00175-0
更新日期：1998-02-01 00:00:00
