抗肾综合征出血热病毒单克隆抗体纯化制剂的免疫学活性鉴定

采用多种方法对两株经辛酸－硫酸铵沉淀法纯化的抗ＨＦＥＳ病毒ＭｃＡｂ进行了免疫学鉴定。结果表明，纯化后的ＭｃＡｂ的免疫学活性（抗体效价）无明显变化，特别是仍保留了很高的中和活性、血凝抑制活性及对感染乳鼠的保护活性。纯化ＭｃＡｂ制剂置４℃保存，至少一年内抗体效价未见降低。     

抗肾综合征出血热病毒单克隆抗体纯化制剂的免疫学活性鉴定

采用多种方法对两株经辛酸－硫酸按沉淀法纯化的抗ＨＦＥＳ病毒ＭｃＡｂ进行了免疫学鉴定．结果表明，纯化后的ＭｃＡｂ的免疫学活性（抗体效价）无明显改变，特别是仍保留了很高的中和活性、血凝抑制活性及对感染乳鼠的保护活性．纯化ＭｃＡｂ制剂置４℃保存，至少一年内抗体效价未见降低     

肾综合征出血热病毒血凝素纯化及免疫学性状分析

应用高压液相色谱法对粗制肾综合征出血热病毒（ＨＦＲＳＶ）血凝素分离纯化获得具有血凝活性的蛋白，效价１８０，分子量为５６６００。ＳＤＳ－ＰＡＧＥ和蔗糖密度梯度超速离心分析显示具有均一的分子量和浮密度，多株位点特异性ＭｃＡｂ分析证实，该蛋白有血凝抗原和中和抗原决定簇，免疫小鼠可产生特异性抗－ＨＦＲＳＶ抗体，其中和效价４０，血凝抑制效价６４。     

肾综合征出血热病毒大鼠单克隆抗体亲和力及抗原结合位点测定

应用非竞争ＥＬＩＳＡ和ＥＬＩＳＡ相加试验对８株抗肾综合征出血热病毒（ＨＦＲＳＶ）大鼠单克隆抗体（ＭｃＡｂ）进行了抗体亲和力和抗原结合位点的测定。结果提示这些ＭｃＡｂ至少可识别ＨＦＲＳＶ核壳蛋白（ＮＰ）上６种不同抗原决定簇，并且表明ＮＰ上可能存在ＨＦＲＳＶ组、亚组、型或亚型抗原决定簇。ＭｃＡｂ亲和常数测定的高低排序结果为ＫＦ１２＞ＤＥ１１＞Ｘ１０＞ＣＨ１０＞ＩＧ４。     

人巨细胞病毒单克隆抗体的研制检定和应用

采用人巨细胞病毒（ＨＣＭＶ）ＡＤ１６９株作为免疫原，制备出１３株鼠－鼠杂交瘤细胞系。对其中的６株进行了检定．免疫印迹试验结果表明：单克隆抗体（ＭｃＡｂ）７Ｂ４、７Ｄ７、７Ｅ１１、８Ｅ８和８Ｄ６相对应的ＨＣＭＶ多肽分子量分别为４６、１５０、３８、５１７２和６５ｋＤ．ＨＣＭＶ感染人胚肺二倍体细胞（２ＢＳ）后不同时间制成抗原片，与ＭｃＡｂ作间接免疫荧光试验。结果表明：ＭｃＡｂ８Ｂ８相应的病毒多肽为即刻早期抗原，其它５株ＭｃＡｂ相应的病毒多肽均为晚期抗原，６株ＭｃＡｂ等量混合后，标上辣根过氧化物酶，用于ＩｇＭ抗体捕获法ＥＬＩＳＡ（ＭａｃＥＬＩＳＡ）中，并与间接ＥＬＩＳＡ（ＩＥＬＩＳＡ）同时检测ＨＣＭＶ－ＩｇＭ．在未经选择的１００份脐带血中，两法均为阳性的３份，两法均为阴性的９４份；ＭａｃＥＬＩＳＡ阳性而ＩＥＬＩＳＡ阴性的２份血清的特异性试验证明，ＨＣＭＶ－ＩｇＭ确为阳性．ＩＥＬＩＳＡ阳性而ＭａｃＥＬＩＳＡ阴性的１份血清的特异性试验证明，它是由ＲＦ引起的假阳性。     

肾综合征出血热病毒结构蛋白的纯化及免疫学特性分析

应用从肾综合征出血热（ＨＦＲＳ）患者血清中提取的ＩｇＧ与活化的Ｓｅｐｈａｒｏｓｅ４Ｂ偶联，制备亲和层析柱，用此亲和层析柱从ＨＦＲＳＶ感染的小白鼠乳鼠鼠脑中提取出２种分子量为６７０００和５５０００的病毒结构蛋白。经ＥＬＩＳＡ证明，此病毒结构蛋白能与ＨＦＲＳ患者血清ＩｇＧ及抗ＨＦＲＳ病毒的ＭｃＡｂ反应，效价为１６０。与６株抗ＨＦＲＳＶＭｃＡｈ试验表明，此病毒结构蛋白能与Ｈ７株反应。血凝试验证明，此病毒蛋白不能凝集鹅红细胞、鸽血球和人＂Ｏ＂型血红细胞。将纯化的病毒结构蛋白免疫家兔，证明其有较强的免疫原性，可刺激家兔产生特异性抗体；微量中和试验及乳鼠中和试验证明，中和效价为２５６，说明纯化的病毒结构蛋白具有中和抗原位点；免疫血清对感染乳鼠有一定保护作用。     

抗肾综合征出血热病毒人单克隆抗体杂交瘤细胞株的建立及初步鉴定

以亲和层析技术纯化的肾综合征出血热病毒（ＨＦＲＳＶ）结构蛋白（５５０００，６７０００）为特异性抗原，体外免疫正常人外周血淋巴细胞（ＰＢＬ），然后将体外免疫的淋巴细胞与人－鼠种间杂交瘤细胞（Ｋ６Ｈ６／Ｂ５）融合，经ＥＬＩＳＡ间接法筛选出２株（２Ｄ５，１Ｂ７）分泌抗－ＨＦＲＳＶ人单克隆抗体（Ｈ－ＭｃＡｂ）杂交瘤细胞株，４次克隆化后，１００％阳性。对２Ｄ５株初步鉴定表明，其抗体类型为ＩｇＧ１；培养上清中抗体浓度为２０～３０μｇ／ｍｌ；特异性免疫荧光反应证明２Ｄ５株Ｈ－ＭｃＡｂ是ＨＦＲＳＶ特异性；中和效价为１∶１６０；体外连续传代６个月仍稳定分泌抗体。     

免疫亲和层析纯化汉坦病毒核壳蛋白实验条件的研究

为了建立免疫亲和层析纯化汉坦病毒核壳蛋白（ＮＰ）的方法，应用酶联免疫吸附试验（ＥＬＩＳＡ）洗脱模型筛选出一种高效而温和的ＮＰ洗脱液，即ｅｔ－ＭｇＣｌ２（乙二醇－ＭｇＣｌ２）。ｅｔ－ＭｇＣｌ２能高效洗脱ＮＰ，可在数分钟内将结合于ＥＬＩＳＡ板固相与Ａ３５单克隆抗体（ＭｃＡｂ）结合的ＮＰ几乎完全解离，在用Ａ３５ＭｃＡｂ－Ｓｅｐｈａｒｏｓｅ４Ｂ免疫亲和层析柱纯化ＮＰ的实验中观察到同样的洗脱效果，经ＳＤＳ－     

抗淋球菌PIA单克隆抗体的制备和应用分析

用淋球菌全菌体免疫ＢＡＬＢ／ｃ小鼠，取脾细胞与Ｓｐ２／０骨髓瘤细胞融合，以纯化ＰＩＡ做ＥＬＩＳＡ间接法筛选，获得５株稳定分泌抗ＰＩＡ的ＭｃＡｂ的杂交瘤细胞株。５株ＭｃＡｂ中３株为ＩｇＭ类（２Ｈ１１，４Ｈ８，４Ｅ１０），另２株分别为ＩｇＧ１（１Ｃ２）和ＩｇＧ２ｂ（５Ａ５）亚类。２Ｈ１１和１Ｃ２为高亲和力抗体，１Ｃ２与５Ａ５识别的抗原表位相同。Ｗｅｓｔｅｒｎ ｂｌｏｔｔｉｎｇ试验表明，５株ＭｃＡｂ均能     

肾综合征出血热病毒大鼠单克隆抗体亲和力及抗原…

应用非竞争ＥＬＩＳＡ和ＥＬＩＳＡ相加试验对８株抗肾综合征出血热病毒大鼠单克隆抗体进行了抗体亲和力和抗原结合位点的测定。结果提示这些ＭｃＡｂ至少可识别ＨＦＲＳＶ核壳蛋白上６种不同抗原决定簇，并且表明ＮＰ上可能存在ＨＦＲＳＶ组、亚组、型或亚型抗原决定簇。ＭｃＡｂ亲和常数测定的高低排序结果为ＫＦ１２＞ＤＥ１１＞Ｘ１０＞ＣＨ１０＞ＩＧ４。     

Purification of equine infectious anemia virus antigen by affinity chromatography.

Affinity chromatography was performed to obtain highly purified antigen from equine infectious anemia (EIA) virus. After crude antigen was concentrated by polyethylene glycol precipitation of culture fluids from equine dermal cells persistently infected with EIA virus, and after the virus was disrupted with ether, it was added to a column of cyanogen bromide-activated Sepharose 4B to which EIA-specific antibody had been conjugated. The antigen was effectively released from the column with 5M MgCl2 and proved to be highly purified. Passive hemagglutination tests on sera from EAI infections were carried out, using the purified antigen. Results indicated that the passive hemagglutination test with the antigen was a specific laboratory test with high sensitivity for EIA infection.     

汉滩病毒核蛋白纯化及其免疫学特性的研究

目的　研究汉滩病毒核蛋白及其免疫学特性,研制新一代的肾综合征出血热(HFRS)亚单位疫苗。方法　用亲和层析法纯化重组杆状病毒表达的汉滩病毒核蛋白及野生汉滩病毒76-118的核蛋白,以聚丙烯酰胺凝胶电泳(SDS-PAGE)和免疫印迹实验(Westernblot)鉴定纯化蛋白。用纯化蛋白免疫BALB/c小鼠,分别以福氏佐剂(FCA)和氢氧化铝［Al(OH)     

Use of acute-stage-specific antigens of Toxoplasma gondii for serodiagnosis of acute toxoplasmosis.

Antisera to acetone-fixed tachyzoites were prepared by immunizing rabbits intravenously with the fixed organisms. Immunoblots in which purified immunoglobulin G (IgG) of the antisera was used recognized four antigens of a supernatant (after centrifugation at 10,000 x g for 30 min) fraction of sonicated tachyzoites. A purified toxoplasma antigen fraction, referred to as acute-stage-specific (AC) antigen, that contained mainly three antigens was obtained by affinity chromatography by using the purified IgG. One antigen had a molecular weight of approximately 52,000, and two antigens had molecular weights of approximately 6,000. An enzyme-linked immunosorbent assay (AC-ELISA) in which this purified antigen fraction, obtained by affinity chromatography, was used for detection of IgG antibody detected early acute infection in sera of patients with toxoplasmic lymphadenopathy. Ninety-two percent of sera obtained within 2 months after onset of lymphadenopathy were positive in the AC-ELISA, whereas only 9% of sera obtained later than 5 months after onset of the clinical illness were positive. The AC-ELISA appears highly sensitive and specific for diagnosis of acute toxoplasmosis.     

Identification of measles virus-specific hemolysis-inihibiting antibodies separate from hemagglutination-inhibiting antibodies.

The occurrence of antibodies giving hemolysis inhibition (HLI) but not hemagglutination inhibition (HI) was examined in human convalescent and rabbit hyperimmune sera. HI antibodies, which through their interaction with hemagglutinin components display HLI activity, were removed by absorption with Tween 80-ether (TE)-treated measles virus material. This absorption did not change the titer of non-HI HLI antibodies. After removal of HI antibodies from 16 late measles convalescent sera and three batches of gamma globulin. HLI antibody titers showed a two- to eightfold reduction. The titers of neutralizing antibodies were reduced from 1/4 to 1/20 of the original titers. There was a good correlation between the titers of neutralizing and HLI antibodies both in sera from which HI antibodies had been removed by absorption and in sera spontaneously showing markedly higher HLI than HI antibody titers. HLI antibodies with these characteristics could be identified in HI tests when whole virus instead of TE-treated material was used an antigen and anti-antiserum was added to the tests. In contrast to the situation in human sera, antibodies remaining after removal of HI antibodies from rabbit hyperimmune sera against purified virus particles were detectable in neutralization and HLI tests only in the presence of anti-antiserum. However, virus particles from which the major fraction of all envelope projections had been removed by treatment with 0.004% trypsin induced the production of non-HI HLI antibodies active also in the absence of anti-antiserum. TE and formalin treatment destroyed the hemolytic activity of virus preparations and also their capacity to induce a production of non-HI HLI antibodies.     

K99 surface antigen of Escherichia coli: antigenic characterization.

K99 prepared by acid precipitation hemagglutinated guinea pig erythrocytes, whereas K99 prepared by chromatography on diethylaminoethyl-Sephadex did not. K99 purified by either procedure hemagglutinated horse erythrocytes. K99 prepared by acid precipitation contained a second antigen not presnet in the K99 prepared by chromatography on diethylaminoethyl-Sephadex. This antigen could be detected by immunoprecipitation with some, but not all, sera prepared against K99-positive Escherichia coli strains. It was assumed that this second antigen is not K99 and is responsible for the guinea pig erythrocyte hemagglutination reaction. Furthermore, the second antigen has an isoelectric point of 4.2, which has been reported by Morris and co-workers to be the isoelectric point of K99.     

Purification and characterization of a rhamnose-containing cell wall antigen of Streptococcus mutans B13 (serotype d).

A rhamnose-containing polysaccharide (RCP) was extracted and purified from cell walls of Streptococcus mutans B13 (serotype d) and was chemically and immunologically characterized. Walls were initially extracted with 5% trichloroacetic acid at 4 degrees C to remove the serotype antigen and were then sequentially extracted with increasing concentrations of hot acid. Extracts lacking galactose were combined and chromatographed on a column of diethylaminoethyl--Sephadex A25. The purified RCP contained 90% carbohydrate, 1.4% protein, and 0.16% phosphorus. Analysis by gas chromatography indicated that the RCP was composed of rhamnose and glucose in a 1.6:1 ratio. RCP was immunogenic in rabbits when animals were immunized with whole cells or cell walls. Antisera prepared against partially extracted cell walls of B13 appeared specific for RCP. These sera were not reactive with purified serotype d antigen or lipoteichoic acid in passive hemagglutination assays or by agar gel diffusion. The RCP appeared to be a cell wall polysaccharide that was both chemically and immunologically distinct from the serotype d antigen.     

流行性出血热循环免疫复合物各组分的动态观察及意义

应用ＰＥＧ沉淀法对２７例１１４份流行性出血热病人血清中循环免疫复合物进行了检测，不同病期循环免疫复合物检出率分别为：发热期８８．８％，少尿期８２．５％，多尿期４７．８％，恢复期２１．４％。对流行性出血热循环免疫复合物组分的动态观察发现，在不同病期循环免疫复合物中绝大多数可检出流行性出血热病毒抗原及免疫球蛋白与补体Ｃ３，表明在流行性出血热患者体内检出的循环免疫复合物是特异性病毒抗原刺激机体免疫应答的     

Antigenic relationship between the common antigen (OEP) of Pseudomonas aeruginosa and Vibrio cholerae.

Antibodies were found by the OEP-passive hemagglutination test to cross-react with the common antigen (OEP) of Pseudomonas aeruginosa in sera of rabbits immunized with two serotype (Inaba and Ogawa) strains of Vibrio cholerae. The titer in the OEP-passive hemagglutination reaction rose later than did the agglutinin titer and reached a peak of 640 to 1,280. The titers of OEP antibody formation in rabbits immunized with V. cholerae were almost the same as that of P. aeruginosa. The common antigen of P. aeruginosa was confirmed to exist serologically in both strains of V. cholerae as determined by the indirect fluorescent antibody test and the agar gel precipitin test. Passive immunization with the V. cholerae immune rabbit serum significantly protected mice against P. aeruginosa infection. Purified antibodies cross-reacting with the OEP of P. aeruginosa derived from the V. cholerae immune rabbit sera by OEP-coupled affinity chromatography protected mice against P. aeruginosa infection as compared with the control group, which was injected with 100 microgram of immunoglobin G not containing OEP antibody. The purified antibodies (2.5 microgram per mouse) protected animals challenged with approximately 10,000 50% lethal doses in the control group. Consequently, the common antigen (OEP) of P. aeruginosa proved to be a common antigen of V. cholerae both serologically and in possessing infection protective properties.     

Comparison of hemagglutination inhibition test and enzyme-linked immunosorbent assay for determining antibody to rubella virus.

The hemagglutination inhibition test (HAI) and the enzyme-linked immunosorbent assay (ELISA) for detecting antibody to rubella virus were compared by testing 25 sets of paired sera taken before and after infection and 10 sets of sera taken during acute and convalescent stages of the disease and by screening 700 serum samples from the Collaborative Perinatal Project, NIH/NINCDS. The tests were found to be comparable in their ability to detect positive and negative sera, rises in titers, and seroconversions. When a purified antigen and carefully prepared reagents were used, ELISA was found to be as accurate and reliable as HAI. ELISA required no pretreatment of serum, could easily be automated, and was less time-consuming than HAI.