Production and Characterization of Monoclonal Antibodies against the Extracellular Domain of CA 125

Shojaeian, Sorour; Allameh, Abdolamir; Zarnani, Amir Hassan; Chamankhah, Mahmood; Ghods, Roya; Bayat, Ali Ahmad; Jeddi-Tehrani, Mahmood

doi:10.3109/08820130903496785

Cited by 14 publications

(15 citation statements)

References 28 publications

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“…(12) Antigens from various sources were used previously for production of antibodies against CA-125 (OVCAR-3 cells, supernatants of OVCAR-3 cells, ascitic fluids). (13,14) In our preliminary studies, we immunized animals with OVCAR-3-derived protein purified by gel filtration, OVCAR-3 crude cell extracts, commercially available antigens of human origin, and recombinant CA-125 fragment.…”

Section: Discussionmentioning

confidence: 99%

Development and Characterization of Three Novel Monoclonal Antibodies Against CA-125

Michurina

Kerzhner²,

Klimovich³

2014

Monoclonal Antibodies in Immunodiagnosis and Immunotherapy

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Cancer antigen 125 (CA-125) is the most widely used tumor marker for ovarian cancer. Thus, monoclonal antibodies (MAbs) against CA-125 are valuable reagents for the development of diagnostic tests and immunotherapy. We describe here the generation and characterization of three novel hybridoma cell lines producing MAbs against CA-125. CA-125 purified from culture supernatant of ovarian carcinoma cell line OVCAR-3 by affinity chromatography, was used for immunization of BALB/c mice. Three stable cell lines (3C8, 2B6, and 5A12) were selected for production of antibodies against CA-125 and were expanded in mass culture. All three antibodies were shown to recognize linear epitopes. Antibodies 2B6 and 5A12 were determined to recognize epitope cluster B (M 11-like); MAb 3C8 was classified as group A-epitope binders (OC 125-like). The antibodies produced may be used for the development and improvement of CA-125 immunoassays.

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Section: Discussionmentioning

confidence: 99%

Development and Characterization of Three Novel Monoclonal Antibodies Against CA-125

Michurina

Kerzhner²,

Klimovich³

2014

Monoclonal Antibodies in Immunodiagnosis and Immunotherapy

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“…mAbs were generated as described elsewhere . Spleen cells of the immunized mouse were harvested and fused with SP2/0 at a 5:1 ratio using polyethylene glycol 1500 (Sigma, Milwaukee, WI, USA).…”

Section: Methodsmentioning

confidence: 99%

Immunohistochemical characterization of novel murine monoclonal antibodies against human placenta‐specific 1

Ghods

Ghahremani

Darzi

et al. 2014

Biotech and App Biochem

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Human PLAC1 (placenta-specific 1) is a new member of cancer-testis antigens with 212 amino acids, and its expression is restricted to placenta and at much lower levels to testis. Recently, ectopic expression of the PLAC1 transcript has been demonstrated in a wide range of human tumors and cancer cell lines with a proposed function in tumor cell growth. No monoclonal anti-PLAC1 antibody applicable to immunohis-tochemical staining is available so far. To better understand the PLAC1 expression and localization, we aimed to produce monoclonal antibodies (mAbs) against the extracellular region of PLAC1. Mice were immunized with a synthetic peptide corresponding to the C-terminal 11 amino acids of PLAC1 conjugated with a carrier protein. Hybridomas were produced by standard protocol and screened for positive reactivity by enzyme-linked immunosorbent assay. Reactivity of final two clones was then assessed by Western blotting (WB), immunohistochemistry (IHC), and immunocytochemistry (ICC). Both clones showed a specific immunostaining pattern in human term placenta as the positive control. Reactivity was mostly localized to the cytoplasm of syncytiotrophoblasts. One of the clones showed an excellent staining signal in breast, ovary, and prostate cancer cell lines. Importantly, no reactivity was observed with human lymph node cells or prostate. None of the mAbs were able to detect PLAC1 in Western blot. Based on the present results, these mAbs can be used for detection of PLAC1 in IHC and ICC techniques.

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“…To explain the phenomenon of low immunoactivity (Shojaejan et al, 2010), one would assume that aberrant glycosylation of cancer cell-expressed immunoglobulins may be the main reason. To further explain the discrepancy, efforts were made to determine the relative Immunoglobulin distributions of different classes or subclasses of cancer cell-expressed immunoglobulins which were compared with those found in normal human serum.…”

Section: Activity (%)mentioning

confidence: 99%

Molecular and Immuno-Characteristics of Immunoglobulin-like Glycoproteins in Cancer Cell-expressed Biomarker, CA215

Lee

Cheung

et al. 2012

Immunological Investigations

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RP215 monoclonal antibody (Mab) was shown to recognize a specific carbohydrate-associated epitope found in cancer cell-expressed glycoproteins, known as CA215. The membrane-bound and soluble forms of CA215 were detected in almost all of the cancer cells in humans, but rarely found in normal tissues. Through MALDI-TOF MS analysis, it has been reported previously that as much as 40% of the detected tryptic peptides of CA215 showed high degrees of sequence homology to those found in immunoglobulin heavy chains. The cancer cell-derived immunoglobulins were further purified from CA215 by affinity column-linked with goat anti-human IgG for molecular characterizations. Semi-quantitative RT-PCR was used to determine the mRNA levels of various immunoglobulin genes expressed by cancer cells of single or multi-cell origins and compared with those found in normal human serum. The stability of CA215 was investigated under different experimental conditions. It was observed that the RP215-specific epitope in CA215 is stable at neutral pH, in human serum or in mice (half life of 5-18 days), but unstable at extreme pH's (pH ≤ 2.0; pH ≥ 12.0) or high temperatures. Enzyme immunoassays were performed with several secondary antibody probes related to human IgG. It was demonstrated that cancer cell-expressed immunoglobulins with RP215-specific epitope have much lower immunoactivity than that of normal human IgG (≤ 5%), despite the fact that both showed almost identical amino acid sequence in the respective Fc region reported previously. This could be the result of aberrant glycosylation of CA215 in cancer cells. Aberrant glycosylation of glycoproteins may have important biological implications on the proliferation of cancer cells in vitro or in vivo.

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Production and Characterization of Monoclonal Antibodies against the Extracellular Domain of CA 125

Cited by 14 publications

References 28 publications

Development and Characterization of Three Novel Monoclonal Antibodies Against CA-125

Development and Characterization of Three Novel Monoclonal Antibodies Against CA-125

Immunohistochemical characterization of novel murine monoclonal antibodies against human placenta‐specific 1

Molecular and Immuno-Characteristics of Immunoglobulin-like Glycoproteins in Cancer Cell-expressed Biomarker, CA215

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