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C Charpin

Publications and source records attributed to C Charpin.

At least 73 records · Page 4Linked to original sources
PubMed1991

Fel d I allergen distribution in cat fur and skin.

Immunohistochemical procedures were performed to ascertain Fel d I antigen (Ag) distribution in cat fur and skin biopsy specimens and to analyze Fel d I allergen concentrations in fur. One hundred strands of fur and 24 skin biopsy specimens (6 by 4 by 3 mm) from shaved areas were collected from 11 different cats. Freshly depilated hairs were immunostained by free-floating monoclonal anti-Fel d I, avidin-biotin-peroxidase complex, and either processed for scanning electron microscopic examination or mounted on glass slides for computer-assisted densitometric analysis (SAMBA system). Skin biopsy specimens were promptly frozen and sectioned just before the immunohistochemical processing. Densitometric analysis of fur demonstrated that immunoprecipitate concentrations were tenfold higher at the root than at the tip. However, this finding may be explained by decrease of the thickness of the hair cortex that varied in similar proportions. The Ag accumulated on the strand surface but may focally penetrate into the medulla through the scale-like cortical interstices. In skin biopsy specimens, Fel d I Ag was found in epithelial squamous cells, within the epidermis and hair follicles, on the surface of the epidermis and hair follicles, and in sebaceous gland cells. These data suggest that Fel d I Ag is produced by sebaceous cells and, to a lesser extent, by basal squamous epithelial cells and that it is stored mainly on the surface of the epidermis and fur.

Allergens↗
PubMed1991

Purification and characterisation of a breast-cancer-associated glycoprotein not expressed in normal breast and identified by monoclonal antibody 83D4.

Monoclonal antibody (mAb) 83D4 was generated using formol-fixed paraffin-embedded human breast carcinoma tissue as the immunogen. Previous studies demonstrated that it was reactive with breast carcinoma tissues, but not with normal breast. The antigen identified by mAb 83D4 was detected, using ELISA, in MCF7 breast carcinoma cell line membrane extracts, in primary breast and colon carcinoma tissue extracts and in pleural effusion fluid from patients with metastatic breast cancer. No reactivity with 83D4 was found in either human milk fat globule membranes or skimmed milk. 83D4 reactive antigen was found to be a heterogeneous high molecular weight (MW) protein (apparent Mr:300-400 to over 1000 kDa) by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. The antigen was purified from MCF7 cells, breast and colon carcinomas and effusion fluid, by perchloric acid solubilisation followed by immunoaffinity chromatography with 83D4. The immunopurified antigen from MCF7 cells and pleural effusion fluid was further analysed by gel filtration and ion-exchange chromatography, which confirmed the high MW and indicated the charge heterogeneity of the reactive molecules. The 83D4 reactive antigen strongly bound to wheat-germ agglutinin and weakly to peanut lectin. No binding was found with lentil lectin or concanavalin A. Antigenic activity was strongly reduced by trypsin and subtilysin digestion and by treatment with sodium periodate, but it was not affected by neuraminidase. These results imply the glycoprotein nature of the 83D4-defined antigen and the involvement of carbohydrate, but probably not sialic acid, in the epitope. Purified 83D4 antigen did not display reactivity for mAb HMFG-1, directed against a polymorphic epithelial mucin, PEM, using ELISA, but bound mAb CC49 and weakly mAb B72.3, antibodies which define a tumour associated glycoprotein, TAG-72. Moreover CC49 and 83D4 showed similar reactivity pattern in immunoblotting assays. A double determinant radioimmunoassay confirmed that 83D4 antigen carries epitopes for mAb B72.3 and CC49. Competition radioimmunoassays clearly distinguished the 83D4 defined epitope from those recognised by B72.3 and CC49, demonstrating that antibody 83D4 identifies a unique epitope. It is suggested that the antigens identified by mAb 83D4 and by mAb B72.3 and CC49 may form part of the same family of carcinoma associated glycoproteins.

Antibodies, Monoclonal↗
PubMed1991

Two monoclonal antibodies identify antigens preferentially expressed on normal human breast cells versus breast cancer cells.

In order to obtain antibodies with specificity toward normal mammary epithelial antigenic determinants, we immunized BALB/c mice with normal milk cells and screened the hybridomas against an undifferentiated breast cancer cell line H466B, peripheral blood lymphocytes and normal fibroblasts. Two hybridomas were generated, which produced BA6 (IgG1) and CA4 (IgM) monoclonal antibodies (MAbs). These MAbs did not react with 5 breast cancer cell lines. In cryostat sections of normal human breast tissue, BA6 was reactive with 6/6 and CA4 was reactive with 12/13 specimens both showing an apical staining of epithelial cells. Conversely staining of malignant cells in breast cancer biopsies was observed in 4/33 specimens with BA6 and in 4/19 specimens with CA4. Computerized image analysis (SAMBA) of immunostained sections showed homogeneous distribution of staining, with a high percentage of stained cell surfaces in normal breast (mean percentages of positive surfaces : BA6 : 75% and CA4 : 82%) while, in malignant samples, staining was heterogeneous, with a mean percentage of positive surface of 25% for BA6 and 12% for CA4. Both MAbs reacted strongly with human milk fat globule membranes (HMFGM) and skimmed milk. FPLC size exclusion chromatography of skimmed milk showed that CA4 and BA6 reactive materials eluted in distinct peaks in high molecular weight ranges. Electrophoretic separation of HMFGM followed by CA4 staining detected a high molecular weight reactive band (Mr 380-600 kDa). CA4 and BA6 reactivity was reduced by protease treatment of the antigen but was not affected by neuraminidase digestion, by methanol extraction or by Na-metaperiodate oxidation. After perchloric acid treatment of HMFGM, BA6 activity was lost while the CA4 activity was found in the soluble fraction. The results reported suggest that the two MAbs identify two distinct novel epitopes of normal breast cells.

Animals↗
PubMed1991

Monoclonal antibody 1BE12 immunoreactivity with human endometrium. Correlations with hormone receptors and proliferation cell markers.

The monoclonal antibody (MAb) 1BE12 has recently been reported to react with several human normal and abnormal tissues. In human endometrium, it reacts more strongly with carcinomas than with normal tissue. To investigate the effectiveness of MAb 1BE12 in identifying cell proliferation in human endometrial cancers, 1BE12 immunocytochemical assays (ICAs) were performed on frozen (n = 47) and paraffin (n = 100) sections with subsequent computer-assisted microcytophotometric (SAMBA) evaluation of immunoprecipitate distribution. MAb 1BE12 immunoreactivity was not impaired by tissue fixation and paraffin embedding. It reacted with normal proliferative endometrium but not with normal secretory endometrium, and immunoreactivity increased with the degree of cell proliferation and malignancy, the amount of immunostaining being greater in invasive carcinomas than in normal proliferative endometrium and endometrial hyperplasia. ICAs showed no correlation between MAb 1BE12 immunoreactivity and estrogen and progesterone receptor antigenic sites. On the other hand, MAb 1BE12 staining in frozen sections increased with Ki67, EGFR, pHER-2/neu, and cathepsin immunostaining. These findings suggest that ICAs on frozen and paraffin-embedded biopsy specimens using MAb 1BE12 along with other markers can be useful for early detection and grading of endometrial carcinoma. The relevance of MAb 1BE12 to the selection of patients for laser ablation of the endometrium rather than hysterectomy is also discussed.

Antibodies, Monoclonal↗
PubMed1991

[Production of a monoclonal antibody (B1N) recognizing human nuclear antigen associated with cell proliferation].

This report describes the preliminary characterization of a novel antigen reactive with a murine monoclonal antibody designated B1N produced in our laboratory. This antibody (IgM) reacts in IFI with mammals and also insect cells, by staining in a speckled fashion the nucleus of these cells. Immunoblotting analysis of Hela and murine D55 nuclear extracts revealed a polypeptide with an apparent molecular weight of 120kD (p120). In this work we demonstrated that: 1. this polypeptide appeared in human peripheral blood lymphocytes only when they were induced to proliferate in vitro after phytohemagglutinin stimulation; 2. this polypeptide was no longer detected in D55 resting cells, following serum deprivation; 3. the MAb B1N specifically revealed the nucleus of proliferating cells on frozen sections of uterine tissue. These data strongly suggest that the p120 nuclear antigen expression is associated with the proliferation state of cells.

Animals↗
PubMed1990

Characterization and distribution in human tissues of a glycoproteic antigen defined by monoclonal antibody 1BE12 raised against the human breast cancer cell line T47D.

Spleen cells from inbred Biozzi mice, immunized against the human breast cancer cell line T47D, were fused with murine myeloma SP2O cells to generate monoclonal antibodies. One of these, 1BE12, of IgM isotype, reacted with five of six human breast tumor cell lines, while no binding was detectable with normal lymphocytes, RBC, or fibroblasts. The antigen recognized by monoclonal antibody 1BE12 was localized on the surface of T47D and MCF7 cells and was detected in cell-free supernatants of cultures. The antigen was found also on the surface of milk secretory cells. Immunohistochemical staining of frozen and paraffin-embedded sections of human tissues showed apical polarized reactivity in normal breast glands, while in all breast cancers staining was either cytoplasmic or membranous and heterogeneously distributed. Immunostaining was also observed in some other normal epithelia, including salivary gland, gastroduodenal mucosa, exocrine pancreas, and cervix. The antigen was not detectable in secretory endometrium, whereas proliferative endometrium was strongly stained. Colon carcinoma, and cancers of the bladder and endometrium were strongly reactive. No staining was detected in melanoma, lymphoma, mesothelioma, non-small cell lung carcinoma, and thyroid, renal, and ovarian carcinomas. Lectin absorption of MCF7 membrane extracts reduced 1BE12 binding. A large reduction in 1BE12 reactivity was observed after digestion of T47D and MCF7 membrane extracts with proteases. Treatment with sodium periodate resulted in complete loss of antigenicity, while neuraminidase treatment did not affect 1BE12 binding. These findings suggest that the 1BE12 epitope is expressed on the carbohydrate moiety of a glycoprotein and does not contain sialic acid. Immunoblotting of the perchloric acid-soluble fraction of MCF7 membrane extracts after electrophoresis in 1% agarose detected the antigen as a high molecular weight species (Mr greater than 900,000). The antigen was purified by perchloric acid extraction of MCF7 membrane preparations followed by affinity chromatography on 1BE12 antibody coupled to Sepharose-4B and gel exclusion fast protein liquid chromatography. No reactivity of the purified material was found with monoclonal antibodies directed against human milk fat globule membrane-associated mucins HMFG1 and DF3.

Antibodies, Monoclonal↗
PubMed1990

Correlation between laminin and type IV collagen distribution in breast carcinomas, and estrogen receptors expression, lymph node and vascular involvement.

The laminin (Lam) and type IV collagen (Coll IV) and estrogen receptor (ER) immunodetections were assessed in a large series of 400 human breast carcinomas. In all the cases the patient's age, the tumor size, the histological type and grade, the presence or the absence of axillary lymph node metastasis and of vascular invasion in tumor borders, and ER tumor content were recorded. Monoclonal anti-Lam, anti-Coll IV were applied with the avidin-biotin-peroxidase complex and monoclonal anti ER with peroxidase anti-peroxidase complex, on frozen sections. A computerized system of image analysis referred to as SAMBA (TITN) with specific software for tissue sections analysis permitted a multiparametric quantitative analysis of immunostained surfaces. With this system, in each tumor, the cellularity, the percentage of Lam, Coll IV and receptor positive surfaces versus the total cell surface and versus the epithelial (keratin positive) surface, the integrated optical density IOD histograms were obtained and correlated to morphometrical and standard histological data. From this study, it was shown that: (1) Lam and Coll IV immunostained epithelial basement membranes in carcinomas were correlated to the presence of estrogen and progesterone receptor antigenic sites within the tumors, with a significant decrease of the positive staining in ER-ICA negative tumors in comparison to ER-ICA/PR-ICA positive tumors. (2) The combined densitometric and morphometric evaluation demonstrated a decrease of Lam and Coll IV immunostaining in malignant tumors, correlated to (i) the presence of peritumorous vascular invasion and (ii) keratin positive cells in bone marrow (iii) axillary lymph node involvement. It is concluded that the variations in Lam and Coll IV antigens distribution may be relevant indicators of tumor metastatic potential in breast carcinomas and that computerized image analysis enables the standardization of the evaluation antigens distribution.

Adult↗
PubMed1990

Production of a monoclonal antibody as immunohistochemical marker on paraffin embedded tissues using a new immunization method.

This report describes a method for the production of murine monoclonal antibodies (MAbs) against cellular antigens preserved during formol fixation and paraffin embedding of human tissues in an attempt to select markers that would be useful in immunopathology. Hybridomas were prepared using spleen cells from mice immunized with cell suspensions obtained from formalin-fixed paraffin block sections of a human breast carcinoma. A monoclonal antibody 83 D4 was selected, which was reactive with paraffin embedded breast carcinoma tissues, but not with normal breast. The reactive antigen has a high molecular weight (400-1000 kD) and was detected on the cell surface of live human breast cancer cell lines and on frozen tissues sections. These results demonstrate that the MAb 83 D4 identifies a native breast tumor associated epitope conserved during tissue fixation and embedding and could be used as an immunohistochemical marker.

Antibodies, Monoclonal↗
PubMed1990

Image cytometry of aneuploidy, growth fraction (MoAb Ki-67) and hormone receptors (ER, PR) immunocytochemical assays in breast carcinomas.

DNA nuclear content was assessed in human breast carcinomas (n = 132) using image cytometry. Optical density histograms of Feulgen stained cell imprints from fresh tissue samples, subsequently frozen for immunocytochemical assays, were determined by the SAMBA system and used for the DNA index, the ploidy balance (PB) and the proliferation index (PI) computation. The three parameters were correlated to (i) histological data (tumour grade, vascular and/or lymph node invasion) and to (ii) growth fraction (Ki67), hormone receptor antigenic sites (ER, PR) and intramedullar (bone marrow) biopsies and anti-KL1-positive epithelial cells. It was shown that 57% of breast carcinomas were aneuploid. Aneuploidy PI significantly correlated to the criteria of poor prognosis such as high tumour grade, vascular and lymphatic invasion and to increased Ki67-positive cells, and the absence of or low ER and PR. Since image cytometry is easy to handle and perfectly suitable for current diagnostic practice in pathology departments, particularly for tumour cell ploidy assessment and standardized analysis of immunostaining procedures with morphological control of the preparation, we conclude that image cytometry, as performed with the SAMBA, must be regarded as a relevant tool for prognosis evaluation and therapy guidance in individual patients.

Aneuploidy↗
PubMed1989

Immunodetection in fine-needle aspirates and multiparametric (SAMBA) image analysis. Receptors (monoclonal antiestrogen and antiprogesterone) and growth fraction (monoclonal Ki67) evaluation in breast carcinomas.

Immunocytochemical assays (ICA) using monoclonal antiestrogen receptors (ER ICA), antiprogesterone receptors (PR ICA), and monoclonal antibody Ki67 (Ki67 ICA) were performed in 127 breast carcinomas. The immunostaining procedures were applied on frozen tissue sections, tumour imprints, and fine-needle aspirates in order to compare the variations in the distribution of the antigens detected in the three different types of preparations. Positive reactions detected with peroxidase-antiperoxidase and avidinbiotin-peroxidase, and alkaline phosphatase-antialkaline phosphatase complexes were evaluated through a computerized system of image analysis referred to as SAMBA 200 (SAMBA TITN, Grenoble, France). Application programs specifically developed for the analysis of tissue sections and of cytologic preparations were applied. This system allowed a multiparametric, accurate, reliable, reproducible and automatized evaluation of the heterogeneity of the antigenic sites in tumors. For each markers positive cell surface (PS), and integrated and mean optical densities (IOD, MOD) and IOD histograms were compared. It was shown that (1) there was no significant variation in optical densities in cell imprints and aspirates whereas PS significantly (P less than 0.01) differed in both preparations; (2) there were significant differences of the optical densities between tissue sections and cytological preparations, either imprints or aspirates, likely due to randomly cut nuclei in tissue sections; and (3) there was a significant difference between the PS of tissue sections and aspirates but no significant difference between tissue sections and imprints. It is concluded that fine-needle aspiration constitutes a convenient method for cell sampling, reliable for the diagnosis of malignancies. However, it may not reflect the heterogeneity of cell subpopulations in tissue.

Antibodies, Monoclonal↗
PubMed1989

Myopathy associated with desmin type intermediate filaments. An immunoelectron microscopic study.

The muscle biopsy performed in a 58-year-old woman with a myopathy involving pelvic girdle and lower limbs displayed unusual intermediate filament aggregates by light and electron microscopy. No cardiac involvement was detected. The filamentous aggregates selective for type 1 fibers were found in subsarcolemmal and intermyofibrillar areas closely related to Z bands. Immunohistochemical study by light and electron microscopy using polyclonal and monoclonal antibodies and avidin-biotin peroxidase method revealed that aggregates consisted of desmin filaments. Desmin positive material was unstained with vimentine antibodies.

Cytoskeleton↗
PubMed1989

Immunocytochemical assays in human endometrial carcinomas: a multiparametric computerized analysis and comparison with nonmalignant changes.

Immunocytochemical assay (ICAs) were performed on frozen sections from human endometrial samples (n = 89) including normal endometrium, decidua, hyperplasia with and without atypia, and carcinomas. Monoclonal antiestrogen receptor (ER), anti-laminin (Lam), anti-type IV collagen (Coll IV), and anti-Ki67 were applied with avidin-biotin-peroxidase complex or peroxidase-antiperoxidase complex. The results of the ICAs were evaluated through a computerized system of image analysis referred to as SAMBA. It was shown that this system provided for an accurate reliable and reproducible analysis of ICAs in tissue sections. It is concluded that this multiparametric and standardized method of analysis of ICAs can further be applied in correlations with clinical and biochemical data.

Antigens, Surface↗
PubMed1989

Type IV collagen immunostaining and computerized image analysis (SAMBA) in breast and endometrial disorders.

Type IV collagen immunostaining was performed on tissue sections from a large series of non-malignant and malignant disorders of the breast and endometrium. The results were analysed by means of a computerized system of image analysis referred to as SAMBA. It was shown that this system provided an accurate, reliable, reproducible, automated and multiparameteric analysis of collagen IV immunoprecipitates. It was concluded that this standardized method of analyses can be routinely used for the measurement of collagen IV, thus enabling correlations to be sought with histopathological and clinical data.

Basement Membrane↗
PubMed1989

[Growth fraction (Ki67), ploidy balance and proliferation index in tumors of the urogenital tract and breast].

The practical applications of computer-assisted image analysis systems are multiple in oncology. The computerized system of image analysis referred to as SAMBA (TITN) is particularly relevant to analyse coloured images resulting from immunostaining or histochemical procedures assessed either on tissue sections of any type or cytological preparations (imprints, smears). The SAMBA analysis of positive Ki67 surfaces in tissue sections from breast, endometrial, ovarian, cervical or urinary bladder samples enables a multiparametric evaluation of the growth fraction (GF) in intraepithelial, borderline or invasive proliferations. Moreover, the SAMBA analysis after Feulgen staining procedures provides a parametric evaluation of the nuclei densitometry and morphological features and of the chromatin texture, which serve to compute the ploidy balance (BP) and the proliferation index (PI). In benign tumors, GF and PI are low and tumor cells are diploid with an overall high positive PB values. In malignant tumors, GF, PI and the percentage of aneuploid cells increase with tumor grade and stage whereas PB decreases. In borderline proliferations, FC, PI and PB intermediate values are recorded. These new criteria of prognosis should be assessed routinely in pathology departments and the results from these new investigations are likely to be soon implicated in the selection of patient therapy.

Antigens, Surface↗
PubMed1989

[Giant leiomyoma of the inferior vena cava].

The authors report an uncommon case of giant leiomyoma of the inferior vena cava with intracardiac extension and arising from the external iliac vena. This case report illustrates the diagnostical problems usually encountered with this type of tumour. Indeed, despite ultrasound and transverse CT scan, the diagnosis was only assessed by histopathological analysis following a successful surgical removal of the leiomyoma. Only 7 cases of leiomyoma of the inferior vena cava are already reported in the literature. Relationship with intravenous leiomyomatosis of the uterus is also discussed.

Aged↗
PubMed1988

Multiparametric evaluation (SAMBA) of growth fraction (monoclonal Ki67) in breast carcinoma tissue sections.

Breast tissue samples, including normal breast, nonmalignant disorders, and breast carcinomas (n = 257), were tested with monoclonal antibody Ki67 to define the growth fraction in each tissue subgroup. Immunocytochemical assays using anti-Ki67 and avidin-biotin-peroxidase complex and/or alkaline phosphatase anti-alkaline phosphatase were applied in frozen sections. The immunoreactions were analyzed with a computerized system of image analysis referred to as SAMBA (Systeme d'Analyse Microphotometrique à Balayage Automatique). This system permitted a multiparametric and automatized analysis of colored images. The results obtained were: (a) the SAMBA analysis of Ki67-positive staining was accurate, reliable, and reproducible; (b) the anti-Ki67 immunostaining was significantly (P less than 0.01) increased in malignancies and was related to the tumors' degree of differentiation, the vascular invasion, and the presence of axillary lymph node metastases; (c) anti-Ki67 immunostaining is increased (P less than 0.01) in tumors in which estrogen receptor and progesterone receptor antigenic sites are not detected. It is concluded that the SAMBA analysis of the anti-Ki67 immunocytochemical assay provides relevant information in selecting subgroups of patients with higher risk for relapse.

Antibodies, Monoclonal↗