抗人淋巴细胞E受体单克隆抗体的研究

用杂交瘤技术制备了一识别E受体单克隆抗体Tlla(JN 20-5),间接免疫荧光分析表明Tlla(JN20-5)与多种T细胞反应,并识别人外周血全部T细胞。此抗体对E玫瑰花生成有明显阻断作用,其相应抗原又与E受体一样对胰酶消化敏感。经PHA刺激不同时间后Tlla(JN20-5)与另一CP_2抗体一样,同外周血单个核细胞反应百分率无明显增加,但反应强度增加。标准CO_2抗体Leu 5b、Anti-CCT3都有阻断FITC标记Tlla(JN20—5)的作用,说明Tlla(JN20—5)为一识别E玫瑰花受体的单克隆抗体。     

抗人淋巴细胞E受体单克隆抗体的研究

用杂交瘤技术制备了一识别E受体单克隆抗体T11a(JN20-5),间接免疫荧光分析表明T11a(JN20-5)与多种T细胞反应,并识别人外周血全部T细胞。比抗体对E玫瑰花生成有明显阻断作用,其相应抗原又与E受体一样对胰酶消化敏感。经PHA刺激不同时间后T11a(JN20-5)与另一CP_2抗体一样,同外周血单个核细胞反应百分率无明显增加,但反应强度增加。标准CO_2抗体Leu5b、Anti-CCT_3都有阻断F1TC标记T11a(JN20-5)的作用,说明T11a(JN20—5)为一识别E玫瑰花受体的单克隆抗体。     

用OKT_(11)证明人淋巴细胞45℃孵育上清中存在50KD的抗原

人类T淋巴细胞能与绵羊红细胞(SRBC)形成自然玫瑰花,因此提出在T淋巴细胞表面存在SRBC的受体(即E受体)。Mendes等将人外周血淋巴细胞于45℃保温1小时,经保温后的淋巴细胞形成E玫瑰花的能力大大降低;若将其再置于上清液中温育,其形成E玫瑰花的能力又可得到部分恢复。因此推测,淋巴细胞在45℃孵育后的上清液中可能存在E受体。近年来,运用杂交瘤技术产生抗E;受体的单克隆抗体已经获得成功。OKT_(11)、9.6等抗E受体的单克隆抗体都与T淋巴细胞上50KD的抗原反应。我们利用免疫沉淀技术,将人外周血T淋巴细     

抗人淋巴细胞T8抗原样McAb的研制和初步鉴定

用杂交瘤技术,将经人外周血E花环阳性细胞免疫小鼠脾细胞与NS-1骨髓瘤细胞融合后,产生一分泌IgG_1亚型McAb杂交瘤株。其所分泌抗体经微量放射免疫测定及间接免疫荧光法分析,表明它只能与T细胞系、76％的胸腺细胞及22％的外周血T淋巴细胞反应,不与其它各种不同细胞反应。将此抗体所识别入外周血T淋巴细胞亚群与抗Leu-2a识别T8~+细胞、抗Leu-3a识别T4~+细胞比较,发现此抗体与抗Leu-2a识别同一群细胞。此抗体能从T细胞表面沉淀出30KD(还原条件)或78KD(非还原条件)分子,并完全阻断FITC标记抗Leu-2a与T细胞的结合,说明此抗体是识别T8抗原样的McAb。     

人淋巴细胞表面抗原的研究——Ⅴ.单克隆抗体anti-CCT1和anti-CCT3所识別的决定簇及其功能

近几年来,已发现抗人T细胞表面TP50蛋白(E受体或与E受体有关)的单克隆抗体(单抗),包括T11A,9.6,35.1,Leu5,D66,LF2,多数能抑制E玫瑰花形成,有的能抑制T细胞多种功能,有的能诱     

人对白细胞间介素-2的反应及其产生的细胞基础 用单克降抗体鉴定的自身玫瑰花形成细胞的作用

作者用自身玫瑰花形成T细胞(Tar)和花生凝集素(PNA)将T细胞亚群(T_4~+与T_8~+)进一步分成不同的组份,分析了这些不同组份产生白细胞间介素-2(IL-2)和对其反应的情况,同时提及了对IL-1的反应。用梯度离心及E玫瑰花法从健康人外周血中分离获取T细胞,以OKT_4和OKT_8单克隆抗体加补体的阴性选择法纯化分离T_4~+和     

B细胞表面蛋白CD_(72)/Lyb-2是CD_5的配体

糖蛋白CD_5表达于所有成熟T 细胞和小部分B 淋巴细胞膜表面上。其在免疫上的确切作用尚不清楚。有报道指出CD_5在人和小鼠都可作为一个受体,以和CD_2,CD_(28)相类似的方式介导一个对T 细胞的共同刺激信号。抗CD_5抗体能刺激由CD_3结合TCR 介导的T 细胞增殖,并且也刺激IL-2分泌和其受体的表达,同时也诱导细胞内Ca~(++)浓度的增加。     

Wu T_(11)(CD2)-小鼠抗人绵羊红细胞受体杂交瘤细胞系的建立

本文报道用T细胞系JMN1免疫BALB/c小鼠,然后取免疫脾细胞与小鼠骨髓瘤细胞系X 63Ag8.653融合。用扁桃腺冰冻切片免疫酶染色法,筛选出一株稳定分泌IgG1亚类的抗绵羊红细胞受体(ER)单扰(McAb)的杂交瘤—WuT11。通过E玫瑰花结形成抑制试验,膜抗原抽提物对WuT11的间接免疫荧光阻断试验,对外周血单个核细胞(PBMC)及其E~ 、E~-层细胞,T、B和非T非B细胞系的反应特征以及FACS的反应曲线等,证明WuT11与OKT11和anti-CCT3相类似,竞争抑制试验证明WuT11与anti-CCT3识别同一表位。用WuT11进行亲和层析,提取的相应抗原经SDS-PAGE,测得分子量为50KDa的糖蛋白,证明WuT11属CD_2,为识别人ER的单抗。     

HI_(47),一种抗人成熟B细胞单克隆抗体制备、鉴定及其生物学特性研究

用人扁桃腺细胞免疫BALB/c 小鼠的脾细胞与小鼠NS_1骨髓癌细胞融合,得到一种单克隆抗体HI_(47)(IgG_3),能够结合家兔补体,主要与分化后期的人B细胞反应,还可与巨噬细胞、树突状细胞、内皮细胞及部分激活T细胞结合。HI_(47)识别抗原的分子量为37/35KD,其分布与CD_(20)相似。第四届国际人类白细胞分化抗原会议将HI_(47)命名为CD_(20)+X。本文还初步探讨了HI_(47)抗体抑制SAC诱导B细胞活化增殖的作用。     

慢性乙型肝炎患者T细胞功能的研究

本文以31例慢性乙型肝炎患者为研究对象,用CD系列单克隆抗体检测了患者外周T细胞亚群分型,观察ConA诱导的白细胞介素-2分泌功能,以及非特异性T细胞抑制功能(Ts)对B细胞抗体分泌的影响。结果表明,慢性乙肝患者体内存在T细胞免疫功能的异常,其外周血CD_4~+细胞比例低下,CD~+细胞比例增高,导致CD_4/CD_3比值下降,这种变化与HBV复制活跃的指标HBsAg~+密切相关。因此血清HBeAg~+是反映患者外周血T细胞亚群变化较敏感的指标。此外慢性乙肝患者T细胞产生IL-2的功能明显障碍,同时伴有ConA诱导的Ts功能低下,这种功能的异常与外周血CD_4~+与CD_8~+细胞亚群的变化无明显的统计相关性。     

人CD137单抗诱导不同状态T细胞增殖和凋亡的研究

为了研究一种新的T细胞共刺激分子—CD137对不同状态T细胞增殖和凋亡的双重调节作用。采用3 H TdR掺入法测定T细胞增殖 ,用流式细胞仪测定细胞凋亡。结果显示 :(1)CD137单抗可与T细胞表面的CD137抗原结合 ,明显增强PHA刺激T细胞增殖 ,使T细胞增殖指数较PHA单独作用高 2～ 3倍 ,但CD137单抗单独不能刺激T细胞增殖 ;(2 )对于慢性活化的T细胞 ,CD137单抗可协同PHA诱导T细胞凋亡 ,使T细胞凋亡率从PHA单独作用的 19 2 0 %增加到 36 31% ,CD137单抗单独并不能诱导慢性活化T细胞凋亡。CD137单抗一方面可协同PHA刺激静止状态T细胞的增殖 ,另一方面可协同PHA诱导慢性活化T细胞凋亡 ,对T细胞起双重调节作用。     

WUT6-抗人胸腺细胞共同性分化抗原来交瘤细胞系的建立

本文报导了用人胎儿胸腺细胞免疫BALB/c小鼠,取其脾细胞与小鼠骨髓瘤细胞系NS-1融合,获得一株能稳定分泌抗人共同性胸腺细胞单克隆抗体的杂交瘤细胞株,W_(?)T_(?)。它与72％胸腺细胞、皮肤Langerhan's细胞及部分T细胞系呈阳性反应;外周血T、D细胞、粒细胞、红细胞、血小板及B细胞系、Null细胞系均阴性。FACS检测结果及W_(?)T_(?)阳性细胞的组织分布特点均同HIT_6相似。在胸腺、皮质细胞及髓质中少数胞体较大呈树突状的细胞呈阳性反应。在扁桃体实质中少数散在细胞阳性。上皮基底层中的Langerhan's细胞则呈强阳性反应。用W_(?)T_(?)亲和层析柱提取胸腺细胞膜上相应的靶抗原分子,证明W_(?)T_(?)识别分子量为51KDa的抗原。免疫竞争抑制试验表明,W_(?)T_(?)同抗原的结合可被HIT_6完全阻断。说明W_(?)T_(?)与HIT_6识别相同的抗原决定簇。     

利用基因重组抗原研制人CD20单克隆抗体及其功能的研究

目的　利用基因重组抗原免疫小鼠 ,制备人CD2 0单克隆抗体 ,研究其特异性和功能。方法　用表达重组人CD2 0基因的细胞NIH 3T3免疫BALB c小鼠 ,取其脾细胞与Sp2 0细胞融合 ,以间接免疫荧光法筛选杂交瘤上清。免疫沉淀法及流式细胞术鉴定其识别抗原的相对分子质量 (Mr)和特异性 ;以MTT法、流式细胞术及形态学方法检测诱导细胞凋亡和抑制细胞增殖的功能。结果　利用杂交瘤技术获得了 1株抗人CD2 0的单克隆抗体 1 2 8,它具有CD2 0抗体特异的细胞反应谱 ,其识别的抗原Mr 为 33× 10 3 。MTT实验证实 1 2 8能显著抑制Daudi和Raji细胞生长。结论　利用基因重组抗原制备了 1株能够稳定分泌抗人CD2 0的单克隆抗体的杂交瘤细胞株 1 2 8,此单抗具有抑制CD2 0阳性细胞增殖和诱导其凋亡的功能。     

Pneumonitis in bone marrow transplant recipients results from a local immune response.

Eighteen recipients of allogeneic T cell-depleted bone marrow who developed 22 episodes of interstitial pneumonitis were investigated by bronchoalveolar lavage for the cause of pneumonitis. The cells obtained were examined using a panel of monoclonal antibodies with immunocytochemical techniques to identify lymphocyte subsets and the presence of surface molecules indicative of lymphocyte activation. The majority of patients had an excess of lymphocytes in lavage and most of these cells were positively stained with the McAb recognizing the CD8 antigen (suppressor/cytotoxic type T cells). Although the proportions of CD4+ (helper type) T cells were below normal, the absolute numbers were within normal limits, thus the CD4:CD8 ratio was consistently 1:1 or less. A large proportion of the CD8+ cells displayed HLA-DR molecules (RFDR1+), interleukin-2 (IL-2) receptors (CD25+) and high concentration of CD7 antigen (RFT2+). Further analysis revealed that most CD8+ cells were CD5+ (RFT1+) yet a large proportion (20-40%) were CD5-. A majority of CD8+ cells was also CD38+ (RFT10+) and Leu7+. No clear correlation between the emergence of a raised proportion of activated CD8+ cells and diagnosed cytomegalovirus infection was found. These results demonstrate, however, that cells with the phenotype of the resident T cells of the bronchial epithelium (CD8+CD5-) emerge to the air spaces and express activation markers. This raises the intriguing paradox of an aggressive local immune response occurring in an otherwise immunosuppressed group of patients.     

识别人白细胞免疫标志的单克隆抗体

A monoclonal antibody (48)against human Pan-Leucocytes was prepared. This antibody reacted with all haemopoietic cells tested, but not with red cells and platelets by indirect immunofluorescent staining. It was also disclosed that this antibody is only bound to lymphoid tissues or leucocytes scattered in other tissues with immunoperoxidase staining. The reactivities of McAb 48 with large a mount of various target cells were identical with anti-HLE, McAb of CD45 group. Therefore McAb 48 also recognizes T200 antigen and belongs to CD45 group. The value of McAb 48 in differential diagnosis of malignant lymphoma is discussed.     

Involvement of "accessory" and antigen receptor molecules in human helper T cell clone activation studied by monoclonal antibody inhibition

The requirements for activation of autocrine proliferation in human helper T cell clones (Th-TCC) by allogeneic cells were examined in monoclonal antibody (MoAb) blocking studies. Stimulation was not blocked by CD4, CD5, CD6, CD7, or CD45 MoAbs, despite high levels of expression of these antigens on the TCC. Only CD2 and CD11a (LFA-1) MoAbs blocked activation, the latter only when peripheral blood mononuclear cells (PBMCs) and not B-lymphoblastoid cell line (B-LCL) cells were used at stimulators. Responses to interleukin 2 (IL 2) were only minimally blocked by any of the MoAbs. All TCC were CD3+ and expressed the alpha/beta chain T cell receptor (TCR) as detected by moAb WT31. Accordingly, CD3 and WT31 MoAbs consistently blocked stimulation by B-LCL, and in addition one anti-DR5 TCC and one anti-DQw3 TCC were blocked by MoAb 42/1C1, which is directed to an idiotypic determinant of the HPB-ALL leukemic line TCR. Only these two TCC reacted with moAb 42/1C1 in flow cytometry. These observations suggest that CD2- but not LFA-1-mediated interactions, as well as TCR and stimulating antigen binding, are absolutely necessary to activate Th-TCC.     

小鼠CTL体外非特异性扩增对其杀瘤活性的影响

目的 在体外用抗CD3单抗、抗CD28单抗和白细胞介素2（IL－2）扩增肿瘤特异性CTL,为肿瘤过继治疗提供足够数量的、具有高度杀瘤活性的效应细胞。方法 采用两种方案培养肿瘤细胞免疫的小鼠脾细胞。（1）抗CD3单抗刺激48h后,加入抗CD3单抗和20U／mlrIL－2（抗－CD3＋IL－2组）;（2）抗CD3单抗和抗CD28单抗同时刺激48h后,加入抗CD3单抗、抗CD28单抗和20U／ml rIL－2（抗－CD3＋抗－CD2＋IL－2组）。分别检测2组效应细胞的增殖水平、杀瘤活性及表型。结果 抗CD3＋IL－2组细胞的^3H－TdR掺入量在第6天、12天、20天分别为22126、52426、2072,抗－CD3＋抗－CD28＋IL－2组细胞的^3H-TdR掺入量在第6天、12天、30天分别为32168、12922、3265,后者明显高于前者。在培养12d时,抗－CD3＋IL－2组细胞对FBL03的最大杀伤活性为66．4％,抗－CD3＋抗－CD28＋IL－2组细胞对FBL－3的最大杀伤活性为77．8％。细胞表型FACS分析结果表明,抗－CD3＋抗－CD28＋IL－2组培养12d后的细胞90％以上为Thy1.2^ 细胞,且CD25^ 细胞在抗－CD3＋抗－CD28＋IL－2组、抗－CD3＋IL－2组分别为23．00％、8．15％。结论 抗CD3单抗、抗CD28单抗和低剂量IL－2同时非特异性刺激,可获得大量扩增的、具有高度杀瘤活性的肿瘤特异性CTL。     

A Novel Human Monoclonal Antibody Rapidly Induces Homotypic Cell Aggregation and Promotes Antibody-Secreting Activity by Human B Lymphoblastoid Cell Line IM-9

A human monoclonal antibody (mAb), designated mNKES, was generated by fusing B cells isolated from an enlarged cervical lymph node of a patient with a carotid body tumor (CBT), with human myeloma cell line KR-12 (6TG). The reactivity of mNKES was tested by the indirect immunofluorescence method. The antigen defined by mNKES was expressed on Burkitt's lymphoma cell lines Raji, Daudi, and Ramos and on B lymphoblastoid cell line IM-9. In addition, mNKES reacted with T cells stimulated with recombinant interleukin-2 (rIL-2) obtained from normal healthy donors. However, mNKES did not react with normal resting human T, B, or adherent cells (monocytes/macrophages). When the reactivity of mNKES and mouse mAbs recognizing the human adhesion-associated antigen (CD10, CD11a, CD11b, CD11c, CD14, CD16, CD18, CD23, CD28, CD29, CD31, CD43, CD44, CD45RA, CD50, CD54, CD58, CD80, CD102, CD106, and HLA class I, and HLA class II antigen) with various cell lines was compared, mNKES reactivity was found to be unique, not resembling that of any of these mouse mAbs. Interestingly, mNKES specifically and rapidly (within 2 hr) induced homotypic cell aggregation of IM-9 cells. This mNKES-induced cell aggregation was completely blocked by the addition of EDTA and when incubated at 4°C. The mAbs reactive with CD11a/CD18 (leukocyte function-associated antigen-1; LFA-1) and CD54 (intercellular adhesion molecule-1; ICAM-1) completely blocked the IM-9 cell aggregation induced by mNKES, and induction of IM-9 cell aggregation by mNKES was significantly blocked in the presence of the protein kinase C inhibitors sphingosine and H-7 and completely blocked by cytochalasin B and cytochalasin D, which inhibit microfilament formation. Regarding biological function, IM-9 cells bearing surface IgG (sIgG) effectively promoted IgG-secreting activity underlying the homotypic cell aggregation induced by mNKES. The surface antigen recognized by mNKES has a molecular size of about 55 kDa, as determined by immunoblotting analysis. These findings indicate that mNKES recognizes a novel adhesion-associated antigen distinct from any previously reported adhesion-associated antigens in terms of pattern of cellular distribution and biological function and that mNKES is the first human mAb found that rapidly induces homotypic cell aggregation and effectively promotes the IgG-secreting activity of human B lymphoblastoid cell line IM-9.     

Differences in the induction of CD8+ T cell responses by subpopulations of dendritic cells from afferent lymph are related to IL-1 alpha secretion

The major subset of dendritic cells (DC) from bovine afferent lymph expresses the SIRP alpha MyD-1 antigen, but not CD11a or the antigen recognized by mAb CC81, and potently stimulates CD4+ and CD8+ T lymphocyte proliferation. The minor subpopulation, that is CD11a+ CC81+ MyD-1-, effectively stimulates CD4+ but not CD8+ T lymphocyte proliferation. CD11a+ CC81+ MyD-1- DC did not induce anergy or death or secrete an inhibitory factor. However, supernatant from cultures of CD8+ T cells with CD11a- CC81- MyD-1+ DC significantly enhanced proliferation of CD8+ T cells in response to CD11a+ CC81+ MyD-1- DC, an effect that was blocked by interleukin (IL)-1alpha, but not IL-1beta, specific mAb. The proliferation of CD8+ T cells with CD11a+ CC81+ MyD-1- DC was also enhanced by adding IL-1alpha. IL-1beta slightly enhanced proliferation, whereas IL-2, IL-6, IL-12, and IL-15 had no effect. We conclude that the failure to stimulate CD8+ T cell proliferation results from the lack of IL-1alpha synthesis by this population, which may have important consequences in vivo.