Summary Report of the TD-3 Workshop: Characterization of 83 Antibodies against Prostate-Specific Antigen

Abstract

Twelve research groups participated in the ISOBM TD-3 Workshop in which the reactivity and specificity of 83 antibodies against prostate-specific antigen (PSA) were investigated. Using a variety of techniques including cross-inhibition assays, Western blotting, BIAcore, immunoradiometric assays and immunohistochemistry, the antibodies were categorized into six major groups which formed the basis for mapping onto two- and three-dimensional (2-D and 3-D) models of PSA. The overall findings of the TD-3 Workshop are summarized in this report. In agreement with all participating groups, three main antigenic domains were identified: free PSA-specific epitopes located in or close to amino acids 86–91; discontinuous epitopes specific for PSA without human kallikrein (hK2) cross-reactivity located at or close to amino acids 158–163; and continuous or linear epitopes shared between PSA and hK2 located close to amino acids 3–11. In addition, several minor and partly overlapping domains were also identified. Clearly, the characterization of antibodies from this workshop and the location of their epitopes on the 3-D model of PSA illustrate the importance of selecting appropriate antibody pairs for use in immunoassays. It is hoped that these findings and the epitope nomenclature described in this TD-3 Workshop are used as a standard for future evaluation of anti-PSA antibodies.

Identification of novel prostate‐specific antigen‐binding peptides modulating its enzyme activity

“…PSA was immobilized to the solid phase by mAb H117 which binds to the same epitope as mAb 5E4 [29]. The capture antibody was covalently coupled onto the surface of the CM5 sensor chip according to the manufacturer's instructions, using coupling levels of 5000 resonance units (RU).…”

“…For characterization of the binding site of the peptides on PSA, monoclonal anti PSA antibodies recognizing various epitopes on PSA were used [29]. Each mAb (33 nm) was incubated in wells containing PSA bound by mAb 5E4.…”

Targeting Free Prostate-Specific Antigen forIn VivoImaging of Prostate Cancer Using a Monoclonal Antibody Specific for Unique Epitopes Accessible on Free Prostate-Specific Antigen Alone

“…15,40,41 In the present study, the specific targeting of fPSA was investigated by using PSA30, an IgG1 that is specific for an epitope region uniquely available on fPSA alone. 30,32 We successfully employed labeling with 125 I and evaluated targeting by DAR and biodistribution studies in both LNCaP-tumor-bearing mice and nontumor-bearing animals. The uptake of 125 I-PSA30 on DAR was compared with the uptake of co-injected 18 F-labeled metabolic probes (FDG and choline).…”

“…31 Comprehensive epitope mapping studies have demonstrated that PSA30 has very similar binding characteristics to some of the originally developed mouse mAbs (e.g., 5A10) specific for fPSA alone, 17,19,30,32 and PSA30 recognizes an epitope covered by ACT that is accessible only on fPSA.…”

Standardization as a Private Enterprise

“…For proteins, reference methods typically will require well-characterized antibodies. The establishment of an epitope map for PSA has facilitated development of assays that measure free and complexed PSA equally (8 ). The epitopes recognized by many hK2 antibodies have already been characterized (9 ), and it is to be hoped that the authors extend these studies by mapping all available antibodies.…”