Historical evolution,overview, and therapeutic manipulation of co-stimulatory molecules

Co-stimulatory molecules are key mediators in the regulation of immune responses and knowledge of its different families, structure, and functions has improved in recent decades. Understanding the role of co-stimulatory molecules in pathological processes has allowed the development of strategies to modulate cellular functions. Currently, modulation of co-stimulatory and co-inhibitory molecules has been applied in clinical applications as therapeutic targets in diseases and promising results have been achieved.     

Stereospecific monoclonal antibodies to nicotine and cotinine and their use in enzyme-linked immunosorbent assays

Stereospecific monoclonal antibodies (McAb) have been prepared against the tobacco alkaloid (S)-(-)-nicotine and its major metabolite (S)-(-)-cotinine. Nine anti-nicotine and 4 anti-cotinine hybridomas, selected by a screening procedure that utilized immunoprecipitation of the ³H-labeled natural isomers of nicotine or cotinine, were grown in the ascites fluid of pristane-primed syngeneic BALB/c mice. Antibodies in concentrations up to 7.5 mg/ml ascites and with binding affinities that generally exceeded 10⁸ M⁻¹ were obtained. Enzyme-linked immunosorbent assays (ELISAs) were developed in which nicotine or cotinine derivatives bound covalently to poly- -lysine werecoated onto wells of polyvinyl chloride microtiter plates. Coated wells were incubated sequentially with McAb in the presence or absence of inhibitor, rabbit anti-mouse immunoglobulin, then horseradish peroxidase-labeled protein A (HRP-SpA) before addition of substrate. The antibodies are highly specific and show minimal cross-reactivity with several nicotine metabolites and other structurally related compounds. In the respective assays, only 0.25 ng (S)-(-)-nicotine and 0.12 ng (S)-(-)-cotinine are required to give 50% inhibition of antibody binding, and as little as 0.05 ng nicotine and 0.02 ng cotinine give 15% inhibition. These assays are 5–10 times more sensitive than analogous ELISAs developed with rabbit antisera and HRP-SpA or conventional radioimmunoassays (RIAs) that utilize the rabbit antisera and ³H-labeled ligands. There was good correlation between the levels of nicotine (r = 0.967) found in saliva samples from smokers and non-smokers assayed by McAb-based ELISAs and conventional RIAs.     

抗早孕因子单克隆抗体的制备与鉴定

目的建立分泌抗EPF(Early pregnancy factor,早孕因子)单克隆抗体的杂交瘤细胞株,纯化单抗并鉴定.方法用本实验室已纯化的早孕和肿瘤源性EPF作为抗原刺激Balb/c小鼠,用免疫后的小鼠脾细胞与同系小鼠骨髓瘤细胞(NS-1)融合,经4次克隆化,获得可稳定分泌抗EPF单克隆抗体的细胞株,注入Balb/c小鼠腹腔制备腹水型单抗,Protein-A亲和层析纯化,SDS电泳和Western-blot等方法分析纯化结果.结果融合后获得一株稳定分泌抗EPF抗体的细胞株(C3D11),克隆化后,获得稳定分泌抗EPF单克隆抗体的细胞株,将增殖后的细胞注射Balb/c小鼠腹腔获得腹水型单抗,以亲和层析法纯化,SDS-PAGE分析显示纯化后去掉了大部分杂蛋白,免疫印迹分析抗体纯度较高,与抗原匹配性良好.结论本研究制备的EPF单克隆抗体为特异性抗EPF抗体.     

抗hER合成肽单克隆抗体的制备与鉴定

用合成的人雌激素受体（ｈＥＲ）抗原决定簇多肽（氨基酸序列从１５１～１６５）与ＫＬＨ的偶联物为抗原，免疫ＢＡＬＢ／Ｃ小鼠，经鼠－鼠杂交，ＥＬＩＳＡ和免疫细胞化学法筛选，有限稀释克隆化，得到一株分泌抗ｈＥＲ合成肽的杂交瘤细胞株（２Ｂ１２）。用ＥＬＩＳＡ法测定。该纯化抗体１０ｎｇ／ｍｌ仍与抗原反应，其免疫球蛋白为ＩｇＧ２ａ．ｋ。与ＭＣＦ－７乳腺癌细胞溶解蛋白的Ｗｅｓｔｅｒｎｂｌｏｔ分析，在分子量６７ｋＤ     

Immunohistochemical examination of lung cancers using monoclonal antibodies reacting with sialosylated Lewisx and sialosylated Lewisa

Summary In order to explore the relationship between the expression of cancer-associated glycolipids such as sialylated Le^x (SLEX) and sialylated Le^a (SLEA) and the histological subtypes of lung cancers, 30 cases of small cell carcinoma (SCC) and 47 cases of non-small cell carcinoma (non-SCC) were examined immunohistochemically using monoclonal antibodies reacting with SLEX and SLEA. The forty-seven cases of non-SCC included 20 cases of adenocarcinoma, 20 of squamous cell carcinoma and 7 of large cell carcinoma. Tumour cells of most non-SCCs expressed SLEX and SLEA. In adenocarcinomas, the number of tumour cells having SLEX and SLEA was more than that of squamous cell carcinomas, large cell carcinomas and SCC. In SCC, 14 of the 30 cases were found to be positive for both antigens. Although the cancer cells of 11 cases of 17 intermediate cell type SCC had both antigens, the cells of only 3 of 13 oat cell tumours expressed SLEX and SLEA. The present study shows that SLEX and SLEA are useful markers for lung adenocarcinomas, that most cases of intermediate cell type of SCCs have characteristics similar to non-SCC but that many oat cell tumours lack them.     

Phenotypic differences within the AB blood type of the feline AB blood group system

Flow cytometry immunolabeling, tube agglutination tests, and thin-layer chromatography immunostaining with two different anti-A monoclonal antibodies (anti-A mAb1 and anti-A mAb2) and one anti-B mAb were used to demonstrate differences in expression of the A and B antigens among erythrocytes from type A and four different type AB cats. Although the flow cytometric patterns of reactivity and agglutination scores for erythrocytes from types A and B cats detected with the anti-A and anti-B mAbs were consistent, reactivity among erythrocytes of different type AB cats was variable. By flow cytometric analysis, 99.9% of type A erythrocytes, no type B erythrocytes, 2.5–4.0% of erythrocytes from type AB cats 1, 3, and 4, and 60.7% of erythrocytes from type AB cat 2 had detectable A antigen when anti-A mAb1 was used. In contrast, 86.4% of type A erythrocytes, no type B erythrocytes, 20.2–38.0% of erythrocytes from type AB cats 1, 3, and 4, and 68.5% of erythrocytes from type AB cat 2 had detectable A antigen when anti-A mAb2 was used. In addition, 86.9% of type B erythrocytes, no type A erythrocytes, 83.1–96.8% of erythrocytes from type AB cats 1, 3, and 4, and 73.0% of erythrocytes from type AB cat 2 had detectable B antigen when the anti-B mAb was used. Agglutination scores of type AB cats were comparable to the percent binding on flow cytometry. Thin-layer chromatography immunostains confirmed differences in the amount of A antigen between erythrocyte glycolipids of type A and AB cats and those of type AB cats 1 and 2. These results suggest that at least two different phenotypes exist within the feline AB blood type, which differ in the amount of A antigen expressed on the erythrocyte surface.     

冰冻切片免疫过氧化物酶染色法在抗细胞性单克隆抗体筛选和鉴定中的应用

本文介绍一种快速准确地筛选和鉴定抗细胞性单克隆抗体(单抗)的冰冻切片免疫过氧化物酶染色技术。它既能筛选融合后的大量单抗(一般在3小时以内能筛选200～400孔杂交瘤细胞上清液);又可根据组织固有结构及细胞分区分布的特性,作为全面鉴定单抗的重要方法。我们运用此法从12次融合中,筛选并经克隆化获得291个单抗,进一步全面鉴定后得到抗T细胞等101个单抗。此外还讨论了消除非特异性染色的体会。     

T-cell lymphomas of the stomach: Morphological and immunological studies characterizing two cases of T-cell lymphoma

Summary Using cytochemical, electron microscopic and immunohistochemical techniques in 20 primary malignant lymphomas of the stomach, we found 18 B-cell and 2 T-cell lymphomas. Primary T-cell lymphoma in the stomach has not been previously reported. The T cells in both cases were reminiscent of T immunoblasts with prominent nucleoli and a basophilic cytoplasm. Case 1 showed a cytological relationship to pleomorphic T-cell lymphoma, large cell type. Case 2 contained in addition some cells not previously described in T-cell lymphomas, resembling immature plasma cells with abundant rough endoplasmic reticulum. Focal positivity to acid phosphatase and dipeptidylaminopeptidase IV suggests the T-cell nature of both lymphomas. In both cases the tumour cells were OKT 11 and OKT 4 positive, and negative for OKT 8. Thus, both cases represent high-grade malignant T-cell lymphomas which correspond phenotypically to T-helper cell lymphoma. Case 2 revealed a further immunohistochemical peculiarity: atypical immunoblasts reacted positively with Ki-1 antibody. Thus, it is a Ki-1 lymphoma of T-cell type.     

Serological studies of monoclonal RH1(D) antibodies with RH1(D) variants

In this paper we chose to emphasize three aspects of our work. First we underlined that “low grade and high grade” D weak red blood cells studied at the DNA level could, when monoclonal antibodies were used, give patterns of positive and negative reactions like partial RH1(D) cells. Secondly, we showed the importance of the technical conditions of the study which are essential for establishing a pattern of reactivity defining an epitope. It appears that the use of papain treated cells at room temperature can be misleading for the definition of epitope especially with IgM antibodies. Lastly we pointed out the interest of Rh variant cells, defined at the gene level, to study the expression of RH1(D) epitopes on the external part of the membrane.