Monoclonal antibody-defined antigens of human prostate cancer cell line PC3 (original) (raw)
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International Urology and Nephrology, 1998
Prostatic epithelium basically consists of secretory-luminal, basal and endocrine-paracrine cells. Immunohistochemical procedures are frequently used for showing the cells reflecting different differentiations. In this study, 40 prostatic tissue specimens submitted to the Department of Pathology of In0niJ University, Research Hospital, between 1991 and 1996 were examined. Half of the cases were diagnosed as cancer and the other half had various benign lesions. Of the cases 22.5% (n = 9) were needle biopsy material whereas the remainder, 47.5% (n = 19), were from prostatectomy and 30% (n = 12) were transurethral resection of the prostate (TURP) specimens. High molecular weight anti-cytokeratin antibodies (HMW anti-cytokeratin) stained basal cells both in all normal prostatic tissue and benign prostatic lesions, but in the majority of cancers (70%, n = 14) negative immunoreactivity was seen. Nevertheless, in some of the cancer cases (30%, n = 6) basal cell anti-cytokeratin staining was shown. Negative immunoreactivity with HMW anti-cytokeratin is important in distinguishing between malignant and benign lesions, whereas positive staining is not every time in favour of benign lesions. With the usage of prostate specific antigen (PSA) it was seen that all of the malignant and benign prostatic lesions stained positively. Basal cells in hyperplastic glands were not stained with this stain. Irregular, and in some areas, intense (PSA) immunoreactivity is present in precancerous and malignant lesions. Endocrine cells, which are represented with Chromogranin-A (Chr-A) immunoreactivity and reflecting neuroendocrine differentiation, are present in 75% (n = 15) of benign lesions and in 50% (n = 10) of cancer cases. It was thought that the lesser number of these cells in neoplastic lesions in comparison to the non-tumoral lesions is correlated with the disorder of mechanism that regulates the cell growth. Both in neoplastic and nontumoral tissues the prostatic epithelial cells showed the three markers, namely HMW anti-cytokeratin, PSA, and Chr-A, which may reflect the multidirectional differentiation of these cells from a pluripotent origin.
Production and Characterization of Monoclonal Antibodies against Human Prostate Specific Antigen
Avicenna Journal of Medical Biotechnology, 2015
Background Prostate Specific Antigen (PSA) is an important laboratory marker for diagnosis of prostatic cancer. Thus, development of diagnostic tools specific for PSA plays an important role in screening, monitoring and early diagnosis of prostate cancer. In this paper, the production and characterization of a panel of murine monoclonal antibodies (mAbs) against PSA have been presented. Methods Balb/c mice were immunized with PSA, which was purified from seminal plasma. Splenocytes of hyperimmunized mice were extracted and fused with Sp2/0 cells. By adding selective HAT medium, hybridoma cells were established and positive clones were selected by ELISA after four times of cloning. The isotypes of produced mAbs were determined by ELISA and then purified from ascitic fluids using Hi-Trap protein G column. The reactivities of the mAbs were examined with the purified PSA and seminal plasma by ELISA and western blot techniques. Furthermore, the reactivities of the mAbs were assessed in P...
A Prostate Antigen in Sera of Prostatic Cancer Patients1
A prostate antigen has been detected by a rocket immunoelectrophoresis technique in 17 of 219 sera obtained from patients with advanced prostatic cancer. Sera from 175 pa tients with nonprostatic cancers, including those with late-stage disease of the breast, lung, colon, rectum, stomach, and pan creas, were antigen negative as were 20 samples each from normal adults and age-matched males. Antigen in sera showed immunological identity with antigen in prostate tissue as deter mined by immunoprecipitation peak enhancement experi ments. Using antibody affinity chromatography and radioimmunoprecipitation techniques, the antigen in sera was purified and subjected to sodium dodecyl sulfate electrophoresis; it exhibited a molecular weight of approximately 36,000, similar to that of antigen isolated from prostatic tissue.
Journal of Immunological Methods, 2004
Prostate specific antigen (PSA) is the most important marker for prostate cancer. Antibodies against minor variants of PSA may be useful in the development of novel diagnostic tests for prostate cancer, but it has been difficult to produce such antibodies by protein immunization. In this study, we have compared the characteristics of monoclonal antibodies (MAbs) obtained by genetic immunization with those obtained by protein immunization. The whole coding region of PSA-cDNA was cloned in a mammalian expression vector pCDNA-3. Six mice were immunized four times by intra-muscular (i.m.) injection of the PSA-pCDNA3 plasmid. The MAbs produced were characterized with respect to subclass, epitope specificity, binding to various molecular forms of PSA and affinity. After intra-muscular injection of DNA, anti-PSA antibodies were detected in the serum of all mice, but the antibody titers were markedly lower than after protein immunization. After fusion of the spleen cells from the mice, five hybridomas producing MAbs to PSA were obtained. The MAbs were of IgG1 and IgG2a isotype and they all recognized equally different forms of free PSA, namely enzymatically active, nicked and proPSA. Epitope mapping showed that these MAbs reacted with the same antigenic regions as those obtained by protein immunization. Thus, genetic immunization leads to production of anti PSA MAbs with similar characteristics to those obtained by immunizing with PSA protein. As applied in the present study, it is less efficient than protein immunization, but it is a useful technique when the antigen is not available in the quantities needed for immunization.
International Journal of Cancer, 1984
Two monoclonal antibodies (KH-I and KH-2) against a transplanted fibrosarcoma (KMT-17) in WKA rab were produced by fusing a mouse myeloma (Pl-X63-Ag8.653) with spleen cells from syngeneic rats hyperimmunized with KMT-17. Both antibodies showed complement-dependent cytotoxicity against KMT-17. By absorption of cytotoxicity. KH-I reacted with homologous tumor, other syngeneic fibrosarcomas (KMT-80 and KMT-75). and lung and kidney from normal rats. However, KH-2 reacted with many kinds of tumors and various normal tissues. Antigen specificity was tested by complement fixation and/or solidphase radioimmunoassay using glycolipids isolated from KMT-17 cells and authentic glycolipids. KH-I reacted with globotriglycosyl ceramide which war not detected on
Andrologia, 2009
Somatic cell hybrids were made from mouse myeloma cells and spleen cells derived from BALB/c mice immunized with homogenized epithelial fractions of BPH. The screening by immunoperoxidase staining on human prostate and non-prostate tissue resulted in one monoclonal antibody identifying a prostate specific antigen. Upon SDS-PAGE and Western blot this antigen exhibited a single band at the position of 34 kDa molecular weight. The immunoreactivity of the prostate antigen was found to be localized excluxsively in the epithelial lining of ducts and secretions of normal prostate, BPH and prostate cancer. Anti-p34 antibody reacted with an antigenic determinant on the PSA molecule and inhibited the binding of Anti-PSA antibody to PSA by about SO to 90% in the RIA. Zusammenfassung. Die Lymphozyten-Hybridisierung erfolgte zwischen Myelomzellen und Milzzellen imrnunisierter BALB/c-Mause. Zur Immunisierung wurde eine homogenisierte Epithelfraktion aus BPH-Geweben verwandt. Das Screening-Verfahren erfolgte immunhistologisch von Prostata und nicht-prostatischen Organen. Mit dieser Technik konnte ein monoklonaler Antikorper identifiziert werden, der ein Prostata-spezifisches Antigen erkennt. Das Antigen hat ein Molekulargewicht von 34 kDa und ist in den Driisenepithelien der normalen Prostata, der BPH und des Prostatakarzinoms lokalisiert. Der Antikorper reagiert mit antigenen Determinanten des PSA-Molekiils und inhibiert zu SO bis 90 % die Bindung des Anti-PSA-Antikorpers an PSA.
Immunochemical characterization of surface antigens of TerC, a teratocarcinoma-derived cell line
Proceedings of the National Academy of Sciences of the United States of America, 1979
Rabbit and mouse antisera prepared against teratocarcinoma cells precipitate both glycoproteins and glycolipids from detergent extracts of radiolabeled cells. Extracts of immunoprecipitates with chloroform/methanol, 2:1 (vol/vol) have been resolved on thin-layer gels into multiple peaks. There are more species seen in extracts of teratocarcinoma cells than in extracts of the crossreacting cultured cell line, cl Id. The teratocarcinoma antigens may be extracted out of chloroform/methanol into buffered saline. Incubation in these secondary extracts converts unreactive cells (lymphocytes) to cells reactive with antisera against teratocarcinoma. Furthermore, the coated cells absorb at least 80% of the activity of antisera against teratocarcinoma targets.
Immunopeptidometric Assay for Enzymatically Active Prostate-Specific Antigen
Clinical Chemistry, 2004
Background: Determinations of certain forms of prostate-specific antigen (PSA) have been shown to increase the specificity for prostate cancer (PCa). One such variant, proteolytically active PSA, is a potentially useful tumor marker, but it is not specifically recognized by antibodies. Using phage display libraries, we previously identified a "family" of peptides that bind specifically to active PSA. We used these to develop an immunopeptidometric assay (IPMA) that specifically detects this form of PSA. Methods: Microtitration plates coated with a PSA antibody were used to capture PSA, and a PSA-binding glutathione S-transferase (GST) fusion peptide was used as a tracer. Bound tracer was detected with an antibody to GST labeled with a europium chelate. PSA isoenzymes with high and low enzymatic activity were used to study binding specificity. Results: The IPMA detected enzymatically active PSA but not internally cleaved PSA and pro-PSA, which are enzymatically inactive. The assay detected 1-10% of free PSA in serum from PCa patients. Conclusions: Peptides identified by phage display can be used to develop assays with unique specificities for enzymatically active PSA. IPMA represents a new assay principle with wide potential utility.