Cytotoxicity assay antibody

H. Gazzano-Santoro, P. Ralph, T. Ryskamp, A. Chen, and V. Mukku, A non-radioactive complement-depen-dent cytotoxicity assay for anti-CD20 monoclonal antibody, J. Imunol. Methods, 202, 163 (1997). 

Cell proliferation can be measured by different techniques in various in vitro models and is used as one of the most specific endpoints for DNT evaluation. A commonly used assay is the bromo-deoxyuridine (BrdU) incorporation where analogues nucleotides are integrated into newly synthesized DNA permitting indirect detection of rapidly proliferating cells by fluorescent labelled anti-BrdU antibodies [49], Cytotoxicity assays, such as MTT and ala-marBlue, are also sometimes used to measure proliferation [49]. However, these assays do not distinguish between cell death and inhibited cell proliferation which is of particular importance in DNT studies. 

There are also chemical methods such as HPLC, the most widely used method after bioassays LC-MS the enzyme-inhibition assays such as protein phosphatase inhibition assays, an inexpensive technique [75,76] and the immunoassays (monoclonal antibodies to OA and DTX-1, and enzyme-linked immunoabsorbent assay). There are several commercial kits available, and the cytotoxicity assays obviate the morphological changes caused by the DSP toxin activity in various cell lines for example, human KB cells, salmon and rat hepatocytes, and cultured neurons. 

Diaz, T. M. Pertega, S. Ortega, D. Lopez, E. Centeno, A. Manez, R. Domenech, N. FDA/PI flow cytometry assay of complement-mediated cytotoxicity of antibodies generated during xenotransplantation. Cytometry, Part A 2004, 62A, 54-60. 

Types II and III Hypersensitivity. No simple animal models are currently available to assess Type II (antibody-mediated cytotoxicity) hypersensitivity reactions. IgE antibodies and immune complexes in the sera of exposed animals can be assayed using ELISA or RIA techniques that require the use of specific antibodies to the drug. 

Spectrophotometric plate readers Perkin-Elmer s lambda reader, an automated microprocessor-controlled, microplate reader, offers the flexibility of configuring a reliable, user-friendly, versatile system, capable of accommodating a wide variety of assays requiring calorimetric measurement on microscale (<300pl) samples. These assays include ELISA, protein determination, cytotoxicity, cytoproliferation and antibody sensitivity testing. 

All the antibody—toxin conjugates bind in their entirety to the appropriate immunoaffinity HPLC column and are completely separated from free antibody. There is no adverse affect on the integrity of the conjugates as judged by gel electrophoresis, size exclusion HPLC, and assays of cytotoxic potency (5)... 

Recently, Sen et al. (2001) have described methods for generating highly effective HER-2-specific cytotoxic T cells by arming activated T cells with anti-CD3 X anti-HER-2 bispecific antibody. In this method OKT3 and 9184 anti-HER-2 monoclonal antibodies were conjugated and used to arm T cells that were subsequently tested for binding, cytotoxicity, and cytokine secretion assays. Armed T cells aggregate and selectively kill HER-2-positive breast cancer cells (MCF-7). 

L1210 cell assay, macrophage inhibition assay, inhibition of antibody-dependent cellular cytotoxicity, rosette inhibition assay... 

Pro-inflammatory cytokines are important mediators of inflammation and tissue destruction. This section describes two cell-based assays that were used to screen for inhibitors of cytokine production and some of the compounds discovered using these screens. The two screens were important elements of a collaboration between Xenova Ltd and the Suntory Institute of Biomedical Research to find microbial metabolites with potential utility for the treatment of rheumatoid arthritis. Both screens were cell stimulatory assays with similar formats, the principle of which is illustrated in Figure 3. Treatment of cells with a particular stimulus activates a signal transduction pathway, one of the end results of which is production of a cytokine, which is secreted into the assay medium. After a separation step, the cytokine of interest is measured quantitatively in the supernatant by dissociation enhanced lanthanide fluorescence immunoassay (DELFIA) using a europium-labeled tertiary antibody. At the same time, cytotoxic properties of test substances are determined by assessing their effect on proliferation of the separated cells. 

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