High reactivity of monoclonal antibody (TO73) with human malignant tumor cell line

The biological characteristics d a new monoclonal antibody (TO73) reacting with a vincristine-resistant human leukemic cell line (KY-VCR) were, evaluated. Immunological and electron-immunological studies showed that TO73 reacted with the surface glycoprotein of KY-VCR. TO73 was found to have no effect on cell growth and intracellular uptake of vincristine. In human neoplastic cell ilnes TO73 was found to react with 11 of 27 (41%) cell lines. With regard to de novo primary tumor with one exception, TO73 did not react with any of the examined primary tumor cells. The patient with TO73-positive leukemia died of induction failure due to drug resistance. Complete remission was achieved in the other leukemic patients. These results indicate that TO73 antigen may be associated with immortalization of tumor cells and poor prognosis in some cases.     

Immunohistochemical reactivity of a monoclonal antibody prepared against human breast carcinoma

Summary The reactivity profile of an IgM monoclonal antibody, MBR1, raised against the human breast cancer cell line MCF7, was studied in a variety of human tumours and non-neoplastic tissues by light microscopic immunohistochemistry. The range of reactivity included specific types of non-neoplastic epithelial cells and a number of epithelial tumours. Most mammary carcinomas reacted with MBR1, but adenocarcinomas and squamous carcinomas from different sites were also strongly positive. Different patterns of immunoreactivity were apparent in microscopically normal tissues, in tissues with inflammatory changes and in carcinomas. Heterogeneous staining, despite morphological similarities, was documented in neoplastic and non-neoplastic epithelial cells. The reactivity of MBR1 was different from that reported for other monoclonal antibodies, but revealed similarities to that of monoclonal antibodies and polyclonal sera against human milk fat globule membrane.     

Differential diagnosis of undifferentiated malignant tumors with monoclonal antibody T29/33

An immunoperoxidase method for distinguishing lymphomas from nonlymphoid neoplasms with monoclonal antibody T29/33 is described. This antibody recognizes a 200,000-dalton pan-hematopoietic glycoprotein antigen. Staining in nearly 200 hematopoietic tumors was positive for T29/33, although three of six plasmacytomas were negative for this antibody. Five undifferentiated tumors that were proved to be lymphomas by subsequent electron microscopic and immunohistologic studies were positive for T29/33. Conversely, 11 of 12 undifferentiated tumors with ultrastructural and clinical features of carcinoma or sarcoma were T29/33-negative. The only exception was one sarcoma that was T29/33-positive. Thus, monoclonal antibody T29/33 is a valuable tool for characterizing neoplasms that cannot be diagnosed by histopathologic examination alone.     

Immunoreactivity with monoclonal antibody A-80 and nuclear DNA content in benign and malignant human breast disease.

Immunohistochemical analysis with the monoclonal antibody (MoAb) A-80, recognizing a tumor-associated cytoplasmic mucin-type glycoprotein, and cytometric nuclear DNA assessment were performed on 314 surgical specimens of the human mammary gland. The series included 36 benign conditions, 34 epithelial hyperplasias, 40 carcinomas in situ, and 204 primary invasive carcinomas. Normal breast parenchyma, benign tumors, and other nonmalignant lesions were all of DNA diploid (euploid) type and rarely expressed the A-80 glycoprotein. Differences in MoAb A-80 immunoreactivity and nuclear DNA content were noted among subtypes of epithelial hyperplasias. Fifteen of 34 epithelial hyperplasias were of DNA aneuploid type and the majority were A-80 immunoreactive. Of these 15 immunoreactive aneuploid epithelial hyperplasias, atypical intraductal hyperplasia was the most common subgroup. None of the 19 epithelial hyperplasias of DNA euploid type immunoreacted. Most of the intraductal (33 of 40) and invasive (180 of 204) carcinomas immunostained with MoAb A-80. The majority of the A-80 immunoreactive malignant tumors were of DNA aneuploid type (26 of 33 carcinomas in situ and 108 of 180 invasive mammary carcinomas). The results suggest that expression of the A-80 glycoprotein occurs at an early stage of malignant transformation. Genetically stable (euploid) mammary tumors seem to immunoreact with MoAb A-80 less frequently than genetically unstable (aneuploid) tumor variants. Combined analysis with MoAb A-80 and of nuclear DNA content in premalignant and malignant mammary lesions could be a useful tool of differential diagnostic and prognostic value.     

The immunohistochemical reactivity of a new anti-epithelial monoclonal antibody (MAb b-12) against breast carcinoma and other normal and neoplastic human tissues

Summary A mouse monoclonal antibody, MAb b-12, has been described previously (Stähli et al. 1985) which reacts with a M_r 350 kD glycoprotein with mucin-like characteristics (Stähli et al. 1987) expressed in cytoplasm and on the surface of human breast carcinoma cell lines (MCF-7 and ZR-75-1). In the present report the immunohistochemical reactivity of this MAb with normal and malignant human tissues is analyzed. Pre-experiments showed that the epitope b-12 is resistant to formalin treatment allowing the use of tissue processed by standard paraffin embedding methods. 167 normal and 408 neoplastic tissues were tested by indirect immunofluorescence or the avidin-biotin complex method. MAb b-12 stained the apical cytoplasm of secretory epithelia and their secretions including the acinar and ductular epithelia of the breast. It reacted with all breast carcinomas independent of their histological type or stage, frequently with all but in some cases with a fraction of the tumour cells. Some other carcinomas, primarily those of adenomatous differentiation, were also reactive. In these, however, the fraction of positive tumour cells was usually lower. The b-12 epitope is thus a marker for normal and neoplastic epithelia with secretory functions, particularly for breast carcinomas of all histological types and stages, and perhaps a differentiation marker for abortive adenomatous differentiation in solid carcinomas of the gastro-intestinal, uro-genital or respiratory tract.     

抗人白细胞单克隆抗体1C34-5与正常及肿瘤组织或细胞的反应性

用免疫组化方法及EUSA测检研究抗人白细胞单克隆抗体1C34-5与正常及肿瘤细胞组织的反应性,发现1C34-5仅与正常的白细胞和淋巴造血组织来源的肿瘤细胞反应。在所观察的1例毛细胞白血病、10例恶性淋巴病,2例何杰金氏病的肿瘤组织均与1C34-5呈不同程度的阳性反应,而31例各种不同类型非淋巴造血组织来源的肿瘤均为阴性反应。浸润到肿瘤细胞间或邻近部位的淋巴样细胞也为阳性反应。此抗体在辅助鉴别形态学难以区分的小圆细胞癌和淋巴瘤的病理诊断中,将有一定意义。此外也为观察判断转移癌细胞分布及浸润程度,以及宿主抗肿瘤的免疫反应提供了一种新手段。     

Complement-independent neutralising monoclonal antibody with differential reactivity for strains of human cytomegalovirus

A mouse monoclonal antibody with complement-independent neutralising activity against cytomegalovirus (CMV) and reactive with the 86 kilodalton (kDa) viral glycoprotein H is described. Neutralisation tests against a range of different strains of CMV showed significant crossreactivity, but clear differences were evident between the two prototype viruses AD169 and Davis, and particularly between AD169 and several low-passage recent clinical isolates; CMV present in urine was neutralised weakly if at all.     

一株新型鼠抗人PD-L2单克隆抗体的研制及其生物学特性的鉴定

为了研制特异性识别人PD-L2(B7-DC,CD273)的鼠单克隆抗体,并对其生物学特性和PD-L2分子的表达特性进行初步分析。以高表达人PD-L2分子的基因转染细胞L929/PD-L2作为免疫原,常规免疫BALB/c小鼠,采用B淋巴细胞杂交瘤技术进行细胞融合,以L929/PD-L2作为抗体筛选细胞,L929/mock为对照细胞,经间接免疫荧光标记和流式细胞术分析、反复筛选和多次克隆化培养,筛选出分泌特异性鼠抗人PD-L2单克隆抗体的杂交瘤细胞株;采用Western blot、Ig亚型快速定性试纸法、间接免疫荧光法和竞争结合抑制实验对单抗进行生物学特性的分析,继而利用该单抗进行免疫荧光标记和流式细胞术检测PD-L2在肿瘤细胞株和免疫细胞上的表达特性。结果显示,通过多次融合和反复筛选,成功获得一株特异性鼠抗人PD-L2(B7-DC)的杂交瘤,该杂交瘤分泌的单克隆抗体能特异识别人PD-L2分子。继而利用上述研制获得的单克隆抗体8F2进行免疫荧光标记和流式细胞术检测发现,PD-L2上调性表达在成熟的树突状细胞和调节性T细胞上。提示,成功地获得一株特异性鼠抗人PD-L2单克隆抗体,并对其生物学特性和表达谱进行初步分析,证明其识别抗原表位不同于商品化抗体,是一株新型鼠抗人PD-L2单抗,这为进一步研究PD-1/PD-L2信号通路在免疫应答中的生物学作用提供了有价值的物质基础。     

A monoclonal antibody (HML-1) defining a novel membrane molecule present on human intestinal lymphocytes

A monoclonal antibody, HML-1, was produced by fusion of NSI myeloma cells with spleen cells of a mouse immunized with isolated human intestinal intraepithelial lymphocytes (IEL). Immunofluorescence studies of isolated cells, as well as immunoperoxidase staining of tissue sections, indicated that HML-1 labeled all the various subsets of human intestinal IEL, approximately 40% of lamina propria T cells, 30% mesenteric lymphoblasts and some lymphocytes in other mucosae, particularly IEL. Conversely, it revealed only rare cells in all other lymphoid compartments. Analysis by polyacrylamide gel gradient electrophoresis showed that HML-1 precipitated two major noncovalently bound components of approximate mol. masses of 105 and 150 kDa from human IEL. HML-1 thus defines a novel human membrane antigen present on a subpopulation of lymphocytes preferentially associated with epithelia, and particularly with the intestinal epithelium. The characteristics of this human antigen are very similar to those of an antigen we had previously described in the rat. The possible functional role of this novel class of lymphocyte membrane antigens as well as the nature of the mechanism that triggers their expression remain to be elucidated.     

Murine monoclonal anti-idiotype antibody (alpha) as a probe to detect human monoclonal antibody bound to human tumor tissues

A new immunohistochemical assay was developed for the detection of human monoclonal antibody (HuMAb) bound to human biopsied tumor tissues. A murine anti-idiotype monoclonal antibody, alpha type, 18C6 (IgGl), was raised against an IgM HuMAb, L612, defining a tumor-associated ganglioside antigen (GM3) and used as a probe in a three step cell-binding assay (HuMAb + anti-id + biotinylated anti-mouse Ig). Anti-id 18C6 has an exclusive binding specificity for HuMAb L612, but does not interfere with the binding of L612 to antigen positive melanoma cell lines or to a purified antigen, GM3. The applicability of 18C6 in the three step cell-binding assay was tested first using a melanoma cell line, UCLASO-M12. L612 bound to M12 cells was specifically detected by 18C6 without any background reactivity in ELISA. When this assay was compared with the standard two-step cell-binding assay (HuMAb + peroxidase-conjugated anti-human IgM) using various cultured tumor cell lines, parallel reactivity was observed. The three-step cell-binding assay was then applied to various fresh-frozen human tumor sections. Positive reactivity was demonstrated on various histologic types of human tumor tissues: primary melanoma (10/10), metastatic melanoma (4/4), nevus (10/10), lung cancer (3/6), breast cancer (2/6), and colon cancer (1/1). Adjacent normal tissues were unstained. Control experiments included the cell-binding assay with L612 alone, 18C6 alone. L612 + unrelated mouse IgG, and unrelated IgM HuMAb (L72) + 18C6; but biotinylated anti-mouse IgG did not react with these control preparations. The results indicate that anti-id 18C6 is a highly specific probe to assess the expression of the ganglioside antigenic epitope recognized by the L612 HuMAb on biopsied human tumor tissues.     

A monoclonal antibody (HO-No-1) with specificity for a human nucleolar protein

A murine monoclonal antibody (named HO-No-1) which specifically reacts with a target antigen of 72,000 mol. wt (p72) in the nucleoli of human cells, has been isolated. Using an avidin-biotin peroxidase immunoperoxidase method, it was found that this antibody stains exclusively the nucleolus and not other portions of the nucleus and cytoplasm. This reaction pattern was observed consistently, although to different degrees in 14 human normal tissues and human malignant cell lines. However, after pretreatment with actinomycin D (250 ng/ml) for 2 h, the antibody stains the nucleoplasm uniformly. Thus this study demonstrates evidence for an antibody which reacts specifically with human nucleoli to a protein which could play an important role in rRNA synthesis.     

Immunomorphological research on human breast tumors using monoclonal antibodies to intermediate filament proteins. The proliferative epithelial structures in fibrocystic disease (dysplasia) and benign tumors

Proliferating epithelial structures were studied immunomorphologically in 16 cases of fibrocystic disease and benign tumours. Monoclonal antibodies were used to prekeratin (PK) C12, normally specific for the lining epithelium, to prekeratin (PK) E3 and to the protein of intermediate filaments in mesenchymal cells--vimentin, normally specific for myoepithelium. The immunofluorescent analysis of the most common proliferating structures has shown that there are elements with a variable combination of PK C12, PK E3 and vimentin among the proliferating cells. While there are cells similar to normal lining epithelium (PK C12) or myoepithelium (PK E3) and/or vimentin), many cells have no analogues either among normal cells, or among cells from ductal, lobular and tubular tumour forms. Since in the most common forms of breast cancer only cells containing PK C12 were found, the immunofluorescent study with the application of monoclonal antibodies to PK C12, PK E3 and vimentin can be recommended in difficult and dubious cases for the recognition of carcinogenic or dysplastic nature of morphologically similar proliferating structures.     

Use of NK1 C3 monoclonal antibody in the assessment of benign and malignant melanocytic lesions.

The monoclonal antibody NK1 C3, synthesised by the Netherlands Cancer Institute, has been used to assess its value in the diagnosis of melanocytic lesions. The antigen recognised by this antibody is not denatured by formalin fixation, with the result that the antibody can be used for retrospective studies on conventionally processed material. Positive results were obtained in primary melanoma (18/18), secondary melanoma (21/21), junctional and compound naevi (32/32), intradermal naevi (9/12), congenital naevi (3/3), so called dysplastic naevi (13/13), blue naevi (5/5), and Spitz tumours (3/14). Non-melanocytic tumours were tested for comparison. The results showed relative but not complete specificity of the antibody for melanocytic tumours, with positive results only in breast and prostate tumours (2/6 and 2/5 respectively). Negative results were obtained with basal and squamous cell carcinoma, appendage tumours, neural tumours, and apudomas. The staining pattern of NK1 C3 was compared with that of antibodies to S100 protein and to neurone specific enolase. Compared with S100 protein NK1 C3 gave stronger staining of a higher percentage of cells in the 12 specimens in which a direct comparison was made. Antibody raised against neurone specific enolase in sheep gave very poor results with heavy background staining. We suggest that NK1 C3 is a useful addition to the battery of monoclonal antibodies of value to the diagnostic histopathologist.     

A novel HLA-A determinant recognized by a cytotoxic human hybridoma IgG1 monoclonal antibody (TrJ14)

Abstract: TrJ14 is a cytotoxic human IgG1Λ hybridoma mAb that recognized a novel HLA-A epitope expressed by lymphoblastoid B cells that are homo- or heterozygous for A2, A3, A11, A30, A31, A33, A68 and A69. Based on these results, the HLA type of cell line TEM (10w9057) was retyped as A66. When peripheral blood T cells isolated freshly from 265 HLA-typed normal individuals served as targets, TrJ14 killed cells expressing two TrJ14-positive HLA-A alleles, as well as the majority of cells having one TrJ14-positive and one TrJ14-negative HLA-A antigen. However, TrJ14 failed to recognize or reacted weakly with most HLA-A2 and -A3 heterozygous normal T cells when A2 or A3 was coexpressed together with a TrJ14-negative antigen. The serological reactivity of TrJ14 correlated with the amino acid valine and aspartic acid at positions 76 and 77 of the αl-domain helix. These amino acids were shared exclusively by all the identified TrJ14⁺ alleles.     

Usefulness of a novel monoclonal antibody against human osteocalcin in immunohistochemical diagnosis

Summary A novel monoclonal antibody against human osteocalcin, recently established in our laboratory, was shown by immunoblotting and immunohistochemistry to react specifically with human osteoblasts. In the present study, the antibody was applied to the immunohistochemical diagnosis of human bone tumours, especially osteoblastic tumours. The antibody reacted with all 27 osteosarcomas. No positive reaction was found either in chondrosarcoma, giant cell tumours of bone, soft tissue tumours or epithelial tumours. A positive reaction was found preferentially in the cytoplasm of most of the osteosarcoma cells, but not in the extracellular matrix. Since the antibody reacted with formalin-fixed and paraffin-embedded tissues, it will be a useful tool for routine immunohistochemical diagnosis of osteoblastic lesions.