删除CD4^＋CD25^＋T细胞对OVA诱导的小鼠体液免疫应答的影响

目的观察删除CD4＋CD25＋T细胞对OVA免疫小鼠体液免疫应答的影响。方法小鼠腹腔注射抗CD25单克隆抗体,分别于3天、10天、27天采血流式检测外周血中CD4＋CD25＋T细胞的比例;注射抗CD25单克隆抗体3天后,OVA加铝佐剂免疫未删除和删除CD4＋CD25＋T细胞小鼠,7天后加强免疫一次,分别于初次免疫后7天,加强免疫后14天采血制备血清,ELISA法检测血清中OVA特异性IgE和IgG1的浓度。结果注射抗CD25单克隆抗体3天和10天,外周血中无CD4＋CD25＋T细胞;27天后CD4＋CD25＋T细胞的比例部分恢复。初次免疫7天后,删除CD4＋CD25＋T细胞小鼠总IgE、OVA特异性IgE和IgG1浓度较未删除组升高;加强免疫14天后,删除CD4＋CD25＋T细胞小鼠OVA特异性IgE较未删除组升高。结论删除CD4＋CD25＋T细胞能够影响小鼠OVA特异性体液免疫应答。     

A role for complement in feedback enhancement of antibody responses by IgG3

IgG1, IgG2a, and IgG2b, passively administered with soluble Ags, enhance specific Ab responses. The effect of IgG3 in this type of feedback regulation has not been studied previously. We immunized mice with trinitrophenyl (TNP)-coupled carrier proteins (bovine serum albumin [BSA] or ovalbumin [OVA]) alone or complexed to monoclonal TNP-specific IgG3. The carrier-specific Ab responses were enhanced by several hundred-fold by IgG3. Enhancement was significantly impaired in mice depleted of complement factor C3 and in mice lacking complement receptors 1 and 2 (Cr2-/-). In contrast, mice lacking the common Fc-receptor gamma chain (FcR gamma -/-), resulting in reduced expression of Fc gamma RI and lack of Fc gamma RIII, and mice lacking Fc gamma RIIB (Fc gamma RIIB-/-), responded equally well to immunization with IgG3-complexed Ag as wild-type controls. These findings demonstrate that IgG3 can induce feedback enhancement and that IgG3, in analogy with IgM, uses the complement system for this function.     

Adjuvant effect of cationic liposomes and CpG depends on administration route

In this study we explored the immunization route-dependent adjuvanticity of cationic liposomes loaded with an antigen (ovalbumin; OVA) and an immune potentiator (CpG). Mice were immunized intranodally, intradermally, transcutaneously (with microneedle pre-treatment) and nasally with liposomal OVA/CpG or OVA/CpG solution.In vitro, OVA/CpG liposomes showed enhanced uptake by DCs of both OVA and CpG compared to OVA + CpG solution. A similar enhanced uptake by DCs was observed in vivo when fluorescent OVA/CpG liposomes were administered intranodally. However, after transcutaneous and nasal application a lower uptake of OVA/CpG liposomes compared to an OVA + CpG solution was observed. Moreover, the IgG titers after nasal and transcutaneous administration of OVA/CpG liposomes were reduced compared to administration of an OVA + CpG solution. Although serum IgG titers may suggest limited added value of liposomes to the immunogenicity, for all routes, OVA/CpG liposomes resulted in elevated IgG2a levels, whereas administration of OVA + CpG solutions did not.These data show that encapsulation of antigen and adjuvant into a cationic liposome has a beneficial effect on the quality of the antibody response in mice after intranodal or intradermal immunization, but impairs proper delivery of antigen and adjuvant to the lymph nodes when the formulations are administered transcutaneously or nasally.     

Fucose‐deficient hematopoietic stem cells have decreased self‐renewal and aberrant marrow niche occupancy

BACKGROUND: Modification of Notch receptors by O‐linked fucose and its further elongation by the Fringe family of glycosyltransferase has been shown to be important for Notch signaling activation. Our recent studies disclose a myeloproliferative phenotype, hematopoietic stem cell (HSC) dysfunction, and abnormal Notch signaling in mice deficient in FX, which is required for fucosylation of a number of proteins including Notch. The purpose of this study was to assess the self‐renewal and stem cell niche features of fucose‐deficient HSCs. STUDY DESIGN AND METHODS: Homeostasis and maintenance of HSCs derived from FX^?/? mice were studied by serial bone marrow transplantation, homing assay, and cell cycle analysis. Two‐photon intravital microscopy was performed to visualize and compare the in vivo marrow niche occupancy by fucose‐deficient and wild‐type (WT) HSCs. RESULTS: Marrow progenitors from FX^?/? mice had mild homing defects that could be partially prevented by exogenous fucose supplementation. Fucose‐deficient HSCs from FX^?/? mice displayed decreased self‐renewal capability compared with the WT controls. This is accompanied with their increased cell cycling activity and suppressed Notch ligand binding. When tracked in vivo by two‐photon intravital imaging, the fucose‐deficient HSCs were found localized farther from the endosteum of the calvarium marrow than the WT HSCs. CONCLUSIONS: The current reported aberrant niche occupancy by HSCs from FX^?/? mice, in the context of a faulty blood lineage homeostasis and HSC dysfunction in mice expressing Notch receptors deficient in O‐fucosylation, suggests that fucosylation‐modified Notch receptor may represent a novel extrinsic regulator for HSC engraftment and HSC niche maintenance.     

江苏省赣榆县1～9岁儿童脊髓灰质炎、麻疹、风疹、流行性乙型脑炎抗体水平调查

目的 了解江苏省赣榆县1～9岁儿童脊髓灰质炎、麻疹、风疹和流行性乙型脑炎4种传染病特异性IgG抗体水平,为免疫规划工作提供参考数据. 方法 采用整群抽样的方法,共采集1～9 岁儿童手指血样39 363份,分离血清;采用酶联免疫吸附试验进行4 种病毒特异性IgG抗体水平检测,并进行统计分析. 结果 脊髓灰质炎病毒、麻疹病毒、风疹病毒和流行性乙型脑炎病毒IgG 抗体阳性率分别为97.5%、96.3%、 97.9%和95.9%.阳性率与调查对象的年龄、性别之间差异无统计学意义. 结论 赣榆县1～9岁4类传染病特异性IgG抗体均维持在较高水平,对控制传染病的暴发和流行起到了较好的免疫屏障作用.     

Fucosylation of IgG heavy chains is increased in rheumatoid arthritis

Objectives: Glycosylation of IgG was suggested to be important in the etiology of rheumatoid diseases. Most studies addressed the amount of galactose, but recently we showed that fucose is highly increased in the juvenile chronic arthritis. The objective of this study was to determine fucosylation of IgG heavy chains in patients with rheumatoid arthritis (RA).

Design and methods: IgG was purified from sera of 29 RA patients and 17 matching controls using ammonium sulfate precipitation and ion exchange. Heavy chains were separated by denaturing polyacrylamide gel electrophoresis and their fucosylation analysed using fucose-specific UEA I lectin.

Results: Fucose was found to be approximately 40% increased in RA patients with very high statistical significance (p = 0.00095).

Conclusions: Fucose on IgG heavy chains is significantly increased in patients with rheumatoid arthritis.     

Quantitation of IgG subclass antibodies to pneumococcal capsular polysaccharides by ELISA, using Pneumovax-specific antibodies as a reference

A quantitative enzyme-linked immunosorbent assay (ELISA) method has been developed to assay the levels of IgG subclasses to pneumococcal capsular polysaccharides (PCP) by using a reference standard. This standard solution containing specific antibodies to a polyvalent pneumococcal vaccine (Pneumovax) was purified from the serum of an immunized healthy adult by affinity chromatography. In order to determine the predominant response to Pneumovax in the four IgG subclasses, specific IgG subclasses in preimmune and postimmune sera from six healthy adults were assessed quantitatively by the ELISA. With regard to peak concentrations after immunization, there was a marked increase in the IgG2 subclass, compared with those of IgG1 and IgG3. Such a quantitative assay of Pneumovax-specific IgG subclass antibodies is useful for the direct evaluation of immune responses to immunization with a polyvalent pneumococcal vaccine, and at the same time, for estimating the IgG2 response to PCP antigens in individuals.     

Intranasal immunization with poly(γ-glutamic acid) nanoparticles entrapping antigenic proteins can induce potent tumor immunity

We previously reported strong induction of ovalbumin (OVA)-specific tumor immunity in mice injected subcutaneously with OVA-entrapping nanoparticles comprising amphiphilic poly(γ-glutamic acid) (OVA/γ-PGA NPs). In the present study we investigated antitumor efficacy and associated immune responses in mice vaccinated with OVA/γ-PGA NPs via the nasal cavity. Mice vaccinated intranasally with OVA/γ-PGA NPs resisted challenge by E.G7-OVA tumor cells, and lung metastasis of B16-OVA cells were significantly suppressed by three intranasal doses of OVA/γ-PGA NPs. Although the total serum anti-OVA IgG titer was equivalent between the OVA/γ-PGA NP- and OVA solution-immunized groups, intranasal vaccination with OVA/γ-PGA NPs efficiently induced cytotoxic T lymphocytes (CTLs) and interferon-γ-secreting cells specific for OVA in the spleen and lymph nodes. The antitumor activity induced by intranasal vaccination of OVA/γ-PGA NPs mainly required CD8⁺ CTLs, and the development of long-term specific immunity was confirmed in rechallenge experiments. OVA/γ-PGA NPs administered via the nasal cavity were rapidly taken up by nasopharyngeal-associated lymphoid tissue and delivered to the cervical lymph nodes. Thus, nasal vaccination with antigen-entrapping γ-PGA NPs evokes tumor immunity by eliciting antigen-specific CTLs. γ-PGA NPs are a promising antigen delivery carrier for the development of non-invasive cancer vaccines.     

Nonmyeloablative chemotherapy followed by T-cell adoptive transfer and dendritic cell-based vaccination results in rejection of established melanoma

We demonstrated previously that dendritic cell (DC)-based vaccines could mediate a specific and long-lasting antitumor immune response during early lymphoid reconstitution after lethal irradiation and bone marrow transplant. The purpose of this current study was to examine the potential therapeutic efficacy of DC-based vaccines in combination with sublethal lymphodepletion and T-cell transfer. In an aggressive model of melanoma, treatment with the combination of 200 mg/kg cyclophosphamide (Cy) and 100 mg/kg fludarabine (Flu) led to a lymphopenic state lasting approximately 14 days, but had no effect on the growth of an established M05 melanoma. Addition of ovalbumin (OVA) peptide-pulsed DC-based immunization resulted in a delay in tumor growth but did not enhance overall survival in this model. To improve treatment, adoptively transferred naive T cells were added. After induction of lymphopenia with Cy and Flu, transferred T cells demonstrated an activated memory phenotype including high expression of CD44 and low expression of CD62L. Induction of lymphopenia with Cy and Flu in combination with adoptive transfer of naive T cells and OVA peptide-pulsed DCs immunization led to an enhancement in the number of OVA specific, CD8 T cells that demonstrated specific cytotoxic activity, proliferation, and interferon-gamma production in response to the OVA expressing M05 melanoma. This combination therapy also led to tumor regression and enhanced survival in mice bearing M05 melanoma.     

HIV-1中国流行株gag-hIL-2重组痘苗病毒与pIRES-gag-hIL-2核酸疫苗联合免疫的实验研究

目的　将HIV 1中国流行株B亚型gag和hIL 2基因在天坛株痘苗病毒中进行共表达 ,与核酸疫苗联合免疫 ,评价免疫效果 ,为新型艾滋病疫苗开发研制奠定基础。方法　将HIV 1中国流行株gag hIL 2基因片段插入到痘苗病毒载体pJ38启动子下游 ,经同源重组和血凝素阴性空斑筛选重组痘苗病毒 ,SDS PAGE、Westernblot检测目的蛋白。以重组病毒和核酸疫苗免疫BALB c小鼠 ,进行淋巴细胞转化实验及血清抗体的细胞免疫和体液免疫指标检测。结果　获得了重组痘苗病毒vJ38gag IL 2 ,具有更好的免疫原性 ,3rVV效果好于 2rVV ,以 2rVV DNA混合免疫效果最好。结论　重组痘苗病毒vJ38gag IL 2能够表达外源蛋白并诱导机体产生细胞免疫和体液免疫     

Evidence for a regulatory idiotypic network in the in vivo response to H-2 antigens

Treatment of BALB/c mice with purified pig antiidiotype to 11-4.1 (anti- H-2Kk) monoclonal antibody has been found previously to induce the appearance of idiotype-bearing molecules (Id') in the serum of these mice, in the absence of detectable antigen-binding activity. In the present study we examined the effect of subsequent immunization of such antiidiotype-primed mice with the original H-2Kk antigen. Skin grafting of virgin BALB/c mice with BALB.K skin did not generate any detectable Id' antibodies when tested by enzyme-linked immunosorbent assay (ELISA). In contrast, grafting of antiidiotype-primed mice with BALB.K skin specifically boosted ther serum level of Id' molecules. Challenge of antiidiotype-primed mice with either B10.D2 or rat skin had no effect on the production of such Id' molecules. Absorption studies demonstrated that the majority of Id' molecules induced by H-2Kk antigenic stimulus and detected in ELISA are antigen-nonbinding molecules, thus indicating specific restimulation by the original H-2Kk antigen of nonbinding idiotype-positive B cell clones. The relevance of these findings to the existence of network interactions in the immune response to H-2 antigens is discussed.     

Fucosylation of serum glycoproteins in lung cancer patients.

Increased expression of sialyl Lewis X or A antigens on metastatic cancer cells leads to their selectin-mediated extravasation. Profound fucosylation of the serum microenvironment may be a factor that interrupts adhesion and influences the formation of metastases. In this study we quantitatively analyzed fucosylation of serum glycoproteins in small-cell and non-small-cell lung cancer patients. Fucosylation of four chosen glycoprotein bands was measured as the reactivity with Aleuria aurantia lectin on nitrocellulose blots, preceded by polyacrylamide gel electrophoresis. Relative fucosylation and fucosylation coefficients were calculated by densitometric analysis. Fucosylated oligosaccharides were observed in higher amounts in cancer sera when compared to sera from healthy individuals in all bands analyzed. Glycoproteins of a molecular mass of 29 kDa appear to carry more fucose residues than the 42-kDa band, comprising alpha(1)-acid glycoprotein and haptoglobin. Glycans of the 26-kDa band were fucosylated to a higher extent in non-small-cell vs. small-cell lung cancer. The results suggest that the extent of fucosylation could be a useful marker for estimation of the glycosylation status of serum proteins in cancer patients. Cluster analysis leads to the preliminary suggestion that the fucosylation status could serve as a predictive factor for patient survival.     

Development and transfer of immediate cutaneous hypersensitivity in mice exposed to aerosolized antigen.

We previously showed that BALB/c mice sensitized to ovalbumin (OVA) by brief daily inhalations of antigen over 10 consecutive days exhibit elevated antigen-specific serum IgE antibody levels and increased airways responsiveness. For the first time, we now show that animals sensitized in this fashion to either OVA or ragweed (RGW) develop immediate hypersensitivity skin test reactions when challenged 2 d after completion of the sensitization protocol. Skin testing, performed by direct assessment of wheal formation after intradermal injection of allergen, was sensitive and specific, since animals exposed to RGW by inhalation only responded to RGW, and OVA-sensitized animals responded only to OVA. Positive reactions were associated with mast cell degranulation, whereas control injections were not. Since only sensitized IgE high responder BALB/c mice but neither nonsensitized BALB/c mice nor OVA-sensitized IgE low responder SJL/J mice exhibited wheal responses, induction of OVA-specific IgE appeared to be essential for the mediation of OVA-specific immediate hypersensitivity reactions of the skin in this model. Passive cutaneous anaphylaxis (PCA) testing confirmed the presence of antigen-specific IgE in the serum. Mice that developed IgG (predominantly IgG2b) anti-OVA antibodies did not respond to OVA injection, indicating that OVA-specific IgG was not involved in this system. Further support for the role of IgE in the immediate hypersensitivity response included the wheal response to intradermal injection of anti-IgE antibody that occurred in OVA- and RGW-sensitized mice at 10-fold lower concentrations than in nonsensitized BALB/c mice and not in sensitized SJL/J mice. After transfer of mononuclear cells from peribronchial lymph nodes of OVA- or RGW-sensitized BALB/c mice, naive, syngeneic recipients developed antigen-specific IgE and specific immediate hypersensitivity responses, indicating that the local lymphoid tissue at the site of sensitization can transfer responsiveness to these allergens. These results demonstrate for the first time the ability to elicit and study IgE-mediated immediate skin hypersensitivity responses in the mouse and illustrate the association of increased antigen-specific and total serum IgE levels, airways hyperresponsiveness, and antigen-specific immediate cutaneous reactivity after sensitization to allergen via the airways.     

S-adenosyl-L-homocysteine hydrolase inactivation curtails ovalbumin-induced immune responses

The reversible S-adenosyl-L-homocysteine (AdoHcy) hydrolase inhibitor methyl 4-(adenin-9-yl)-2-hydroxybutanoate (DZ2002) suppresses macrophage activation and function. The effects of DZ2002 on T cell function, however, are still unclear. Here, we examined whether DZ2002 alters type 1 helper T cell (Th1) and/or type 2 helper T cell (Th2) immune responses, and whether these effects are associated with both the inhibition of AdoHcy hydrolase and intracellular elevation of endogenous AdoHcy. Male C57BL/6 mice immunized with ovalbumin (OVA) were treated with DZ2002 (1, 5, and 25 mg/kg/day) after which lymphocyte proliferation, cytokine production, and IgG responses to OVA were monitored. Administration of DZ2002 dose dependently suppressed OVA-specific lymphocyte proliferation and anti-OVA IgG production compared with controls. Interleukin (IL)-2 and interferon (IFN)-gamma as well as anti-OVA IgG2a and IgG3, indicators of Th1 immune responses, were markedly decreased in mice treated with DZ2002, whereas IL-4 and anti-OVA IgG1, indicators of Th2 immune responses, were only mildly suppressed. AdoHcy hydrolase activity in spleens of DZ2002-treated mice was substantially blocked, and not surprisingly, AdoHcy levels were significantly elevated compared with controls. Finally, similar immunosuppressive effects were also observed in mice treated with AdoHcy. These data strongly indicate that DZ2002 suppresses antigen-induced specific immune responses, particularly Th1 responses, through inhibition of AdoHcy hydrolase and elevation of endogenous AdoHcy.     

Oligosaccharide-protein conjugate vaccines induce and prime for oligoclonal IgG antibody responses to the Haemophilus influenzae b capsular polysaccharide in human infants

The diversity of the IgG antibody induced by immunization of human infants and children with conjugate vaccines, composed of oligosaccharides prepared from the Haemophilus influenzae b capsular polysaccharide (CP) and covalently linked to diphtheria toxoids, was studied by analytical IEF. The antibody response was similar, in the degree of restriction, to that observed in the antibody response of older children to immunization with the CP alone. The booster responses induced by reimmunization with conjugate vaccines were accompanied by increases predominantly in the IgG antibody clonotypes expressed after the priming dose of vaccine. After a series of conjugate immunizations, immunization with isolated CP boosted the antibody titer and increased expression from all the clonotypes that were expressed after conjugate immunization. These findings suggest that the conjugate vaccines are acting on a limited number of human B cell clones that are preferentially restimulated after reimmunization. Little evidence of antigen-specific B cell recruitment was found. In addition, the ability of isolated CP immunization to restimulate the same B cell clone indicates that the responding B cell has matured and suggests a linear rather than a dual developmental pathway for the B cell participating in this human antibody response.     

2013年内蒙古自治区健康人群麻疹抗体水平调查分析

目的 了解内蒙古自治区健康人群麻疹抗体水平,及时发现高危人群,为制定消除麻疹防控措施提供依据。方法 按照全国麻疹监测方案健康人群免疫水平监测的要求,按健康人群分为8个年龄组,每组随机抽取不少于30人,应用酶联免疫吸附试验测定人体麻疹IgG抗体。结果 采集血清标本751份,麻疹IgG抗体阳性数699份,阳性率、保护率均为93.08%。不同地区、不同年龄组人群阳性率、抗体浓度差异均有统计学意义。结论 不同地区人群麻疹免疫水平不同,应根据该地区实际情况采取相应的措施,提高基础免疫的质量,加强补充免疫,提高人群抗体水平。     

Systemic tolerance and secretory immunity after oral immunization

Diminished systemic immune reaction after ingestion of antigen has been reported in several animal models. Conversely, it has been reported recently that oral immunization may lead to the production of secretory antibodies. To determine whether these events could occur concurrently, CBA/J mice were immunized intragastrically with varying doses of ovalbumin (OVA) and Streptococcus mutans. After 7 d, the animals were challenged systemically with antigen in complete adjuvant and 8 d later serum and saliva taken, and the draining lymph nodes assayed for a proliferative response. Intragastric doses of 1 mg OVA or 10(9) S. mutans led to significant suppression of the proliferative response, and intragastric doses of 10 mg OVA or 2.5 X 10(9) S. mutans led to the production of detectable salivary antibodies using hemagglutination. Serum antibodies were not detected after intragastric administration of OVA or S. mutans. Suppression of the proliferative response could be detected from 2-60 d after intragastric administration of OVA, and 2-21 d after S. mutans. Prior intragastric immunization with heterologous antigens did not suppress the response to OVA or S. mutans. Transfer of 40 X 10(6) mesenteric lymph node cells from mice given 20 mg OVA or 10(9) S. mutans led to suppression of the proliferative response in syngeneic recipients. Salivary antibodies wer removed by absorption with anti-IgA, but not anti-IgG or IgM, indicating that they were of the IgA class. It appears that intragastric administration of soluble or particulate antigens in mice may lead to the concurrent induction of salivary antibodies and systemic suppression.     

Cooperative roles of CTLA-4 and regulatory T cells in tolerance to an islet cell antigen

Adoptive transfer of ovalbumin (OVA)-specific T cells from the DO.11 TCR transgenic mouse on a Rag(-/-) background into mice expressing OVA in pancreatic islet cells induces acute insulitis and diabetes only if endogenous lymphocytes, including regulatory T cells, are removed. When wild-type OVA-specific/Rag(-/-) T cells, which are all CD25(-), are transferred into islet antigen-expressing mice, peripheral immunization with OVA in adjuvant is needed to induce diabetes. In contrast, naive CTLA-4(-/-)/Rag(-/-) OVA-specific T cells (also CD25(-)) develop into Th1 effectors and induce disease upon recognition of the self-antigen alone. These results suggest that CTLA-4 functions to increase the activation threshold of autoreactive T cells, because in its absence self-antigen is sufficient to trigger autoimmunity without peripheral immunization. Further, CTLA-4 and regulatory T cells act cooperatively to maintain tolerance, indicating that the function of CTLA-4 is independent of regulatory cells, and deficiency of both is required to induce pathologic immune responses against the islet self-antigen.     

Hochuekkito, a Kampo (traditional Japanese herbal) Medicine, Enhances Mucosal IgA Antibody Response in Mice Immunized with Antigen-entrapped Biodegradable Microparticles

The effect of oral administration of Hochuekkito (HET; Bu-Zhong-Yi-Qi-Tang in Chinese), a traditional Japanese herbal medicine, on mucosal IgA immune response was investigated. To induce the antigen-specific antibodies in mucosal site, ovalbumin (OVA)-entrapped biodegradable microparticles (OVA-microparticles) were used as an antigen. Mice were orally immunized with OVA-microparticles for 3 successive days with intragastric gavage. From 7 days after the onset of immunization, the mice were boosted twice a week with the same antigen for 2 weeks. HET or water alone was orally administered to the mice via the intragastric route from 7 days before to 27 days after the onset of immunization. Although no significant change in total secretory IgA antibody level was observed in intestinal and nasal washes, OVA-specific IgA titers in intestinal washes were significantly enhanced by oral administration of HET. When lymphocytes from spleen, peripheral blood and Payer's patches were investigated for cytokines production, it was found that the IFN-γ secretion from the lymphocytes was increased by the administration of HET. Microarray analysis of Peyer's patch cells revealed enhanced expression of L-selectin gene. The increase of L-selectin positive cells in B lymphocytes fraction was observed in Peyer's patch cells and peripheral blood mononuclear cells by flow cytometry. These results suggest that the enhanced IFN-γ secretion and increased population of L-selectin positive B lymphocytes by orally administered HET may partly contribute to enhancement of IgA immune response against intestinal antigens, and orally administered HET may strengthen defensive systems against various pathogens and food antigens in intestine.     

抗人ATⅢ单克隆抗体的研制

目的：建立人抗凝血酶Ⅲ（ Ａ Ｔ Ⅲ）的 Ｅ Ｌ Ｉ Ｓ Ａ 检测试剂盒及 Ａ ＴⅢ的纯化提供试剂准备。方法：用 Ａ ＴⅢ进行 ２ 次基础免疫、２ 次加强免疫的方法免疫 Ｂａｌｂ／ｃ 小鼠,经间接 Ｅ Ｌ Ｉ Ｓ Ａ 的方法筛选杂交瘤细胞株,通过吸收试验、免疫印迹试验等进行特异性分析。结果：筛选出１５ 株分泌抗人 Ａ ＴⅢ单克隆抗体的杂交瘤细胞株,对其中３ 株（ Ｂ４、 Ｂ５、 Ｂ７）作进一步研究,３ 株杂交瘤细胞分泌抗体亚型均为 Ｉｇ Ｇ１ 型,３ 株细胞培养上清滴度平均达到４×１０３,小鼠腹水滴度平均达到 ４×１０６,３ 株抗体均具抗人 Ａ ＴⅢ的特异性。结论：所筛选的单抗为 Ｅ Ｌ Ｉ Ｓ Ａ 检测试剂盒的制备及 Ａ ＴⅢ的纯化提供了试剂准备。