Immunocytochemical demonstration of S-phase cells by anti-bromodeoxyuridine monoclonal antibody in human brain tumor tissues

Nagashima, Tadashi; DeArmond, Stephen J.; Murovic, Judith A.; Hoshino, Takao

doi:10.1007/bf00688136

Cited by 164 publications

(59 citation statements)

References 15 publications

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“…One hundred twenty-one specimens obtained by endoscopic biopsy from patients with primary gastric cancers at the Department of Surgery 11, Kanazawa University, Kanazawa, Japan, between 1980 and1985, were analyzed for PCNA immunoreactivity. Biopsy specimens were immediately fixed in 10% formalde-hyde solution overnight and embedded in paraffin.…”

Section: Methodsmentioning

confidence: 99%

Analysis of proliferative activity using anti-proliferating cell nuclear antigen antibody in gastric cancer tissue specimens obtained by endoscopic biopsy

Yonemura

Kimura

Fushida

et al. 1993

Cancer

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Section: Methodsmentioning

confidence: 99%

Analysis of proliferative activity using anti-proliferating cell nuclear antigen antibody in gastric cancer tissue specimens obtained by endoscopic biopsy

Yonemura

Kimura

Fushida

et al. 1993

Cancer

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“…The technique has been described in detail elsewhere (18). Briefly, tissue sections were deparafhized for 15 min in xylene, rinsed three times (5 min each time) in 100% ETOH, and incubated for 30 min in 2 N HC1.…”

Section: Peroxidase Immunohistochemical Analysismentioning

confidence: 99%

“…Indirect peroxidase staining of tissue sections is a far more reliable method than counting the frequency of mitoses to determine the proliferative potential of glial tumors (18). The LIs obtained in the present study were comparable to those obtained by 3H-thymidine labeling studies in our institution (11,14), which were very high in medulloblastomas (12.0 f 1.3%, & SE) and glioblastomas (9.3 & 1.0%), low in welldifferentiated gliomas (< 1.0%), and intermediate in anaplastic astrocytomas (4.0 0.8%).…”

Section: Reutiue Fluorescplce Inte)(sitymentioning

confidence: 99%

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Cell kinetic studies of in situ human brain tumors with bromodeoxyuridine

et al. 1985

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At the time of surgery, 18 patients with various brain tumors were given a 1-h i.v. infusion of bromodeoxyuridine (BrdUrd), 150-200 mg/m2. At an infusion rate of 200 mg/m2/h, serum BrdUrd levels of 8 pM were achieved. After the infusion, tumor tissue was obtained and divided into two portions. One portion was fixed in 70% ethanol, embedded in paraffin, and sectioned; the sections were deparaffinized, denatured with 2 N HCl, and reacted with monoclonal antibodies against BrdUrd (anti-BrdUrd MAb). BrdUrd-labeled nuclei were demonstrated satisfactorily by an indirect peroxidase method. The other portion was dissociated into single cells with a DNase enzyme cocktail and reacted with FITC-conjugated anti-BrdUrd MAb to determine the percentage of BrdUrd-labeled cells or with chromomycin A3 for DNA analysis. The single-cell suspensions were analyzed by flow cytometry.The fraction of S-phase cells in the tissue sections was similar to both the percentage of BrdUrd-labeled nuclei and the S-phase fraction determined by flow cytometric analysis. The results obtained with BrdUrd-labeled nuclei were similar to those obtained from previous autoradiographic studies of various brain tumors exposed to a pulse of 3H-thymidine. Since BrdUrd is not radioactive and is nontoxic at the dosage used, these techniques, together with the histopathological diagnosis, may help to predict the biological malignancy of individual tumors.Key terms: Cell kinetics, human brain tumor, glioma, bromodeoxyuridine, immunocytochemistry, flow cytometry, monoclonal antibody, S-phase fraction, serum BrdUrd level The labeling index obtained by autoradiographic analysis of tissue exposed to a pulse of 3H-thymidine has been used to estimate the proliferative potential of in situ human tumors (25). Application of this technique has been severely limited because of the potential radiation hazard to normal tissue and to the environment and because the studies are tedious to perform and require several months to complete. Thus, the results are not available soon enough to be considered in the design of chemotherapy regimens for individual patients. With the development of flow cytometry (FCM), it became possible to obtain a measurement similar to the labeling index by quantitating the intensity of DNA fluorescence in individual tumor cells (27). Unfortunately, the difficulty of obtaining clean single-cell suspensions and the lack of sophisticated computer programs to analyze the DNA histograms has prohibited routine use of FCM to measure the proliferative capacity of most human tumors.Recently, Gratzner (7) developed a monoclonal antibody W b ) that can identify nuclei containing bromodeoxyuridine (BrdUrd). Anti-BrdUrd MAb can be detected by direct conjugation of FITC to the antibody, by indirect conjugation of FITC using a n FITC-tagged secondary antibody, or by immunoperoxidase methods. This is an important breakthrough for studies of cell kinetics. BrdUrd, like 3H-thymidine, is incorporated into nuclear DNA during DNA synthesis (6, 26), but is neither radioac...

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“…Recently, Nagashima et al 11,12) successfully em ployed anti-bromodeoxyuridine (BrdUrd) monoclo nal antibody for rapid detection of S phase cells in rat and human brain tumors. However, this proce dure requires that BrdUrd be incorporated into the nucleus of every S phase cell.…”

Section: Introductionmentioning

confidence: 99%

Demonstration of proliferating cell nuclear antigen (PCNA/Cyclin) in glioma cells.

Honda

Nakane

1987

Neurol. Med. Chir.(Tokyo)

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Proliferating cell nuclear antigen (PCNA/cyclin), a non-histone, acidic nuclear protein commonly believed to be strictly confined to proliferating cells, was successfully demonstrated exclusively in the nuclei of human glioma cells both in vitro and in situ by an indirect immunoperoxidase method. Speckled nucleoplasmic staining for PCNA, with variable nucleolar staining, was observed in approx imately 37% of cultured human glioma (KY) cells in exponential growth, whereas only 14% of sub cutaneous KY tumor cells from inoculated nude mice stained unequivocally positive for nuclear PCNA. A direct comparison of this method and autoradiography would be valuable. However, im munocytochemical demonstration of PCNA appears to be a simple method of estimating the pro liferative activity of glioma cells, since the number of PCNA-positive KY cells closely corresponds to the number of S phase cells that exhibit nuclear uptake of bromodeoxyuridine.

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Immunocytochemical demonstration of S-phase cells by anti-bromodeoxyuridine monoclonal antibody in human brain tumor tissues

Cited by 164 publications

References 15 publications

Analysis of proliferative activity using anti-proliferating cell nuclear antigen antibody in gastric cancer tissue specimens obtained by endoscopic biopsy

Analysis of proliferative activity using anti-proliferating cell nuclear antigen antibody in gastric cancer tissue specimens obtained by endoscopic biopsy

Cell kinetic studies of in situ human brain tumors with bromodeoxyuridine

Demonstration of proliferating cell nuclear antigen (PCNA/Cyclin) in glioma cells.

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