恶性疟原虫抗独特型抗体的初步研究

用抗恶性疟原虫单克隆抗体１３Ａ２，制备兔抗血清，经ＢＡＬＢ／ｃ－ＩｇＧ－Ｓｅｐｈａｒｏｓｅ４Ｂ和１３Ａ２ＭｃＡｂ－Ｓｅｐｈａｒｏｓｅ４Ｂ柱提纯，得到抗独特型抗体（Ａｂ２）。免疫双扩散试验和ＥＬＩＳＡ试验表明：抗独特型抗体仅与１３Ａ２ＭｃＡｂ反应。４ｉ试验显示：当恶性疟原虫抗原浓度为８０μｇ／ｍｌ，对１３Ａ２Ｉｄ－Ａｎｔｉｌ３Ａ２Ｉｄ系统产生８１％的抗体结合抑制率，而伯氏疟原虫和诺氏疟原虫抗原检测，抗体结合抑制率分别为２７％和２．５％。兔抗血清中抗Ａｂ２抗体（Ａｂ３）滴度为１：６４０，提示此抗独特型抗体可替代原始抗原诱生抗恶性疟抗体。     

抗间日疟原虫红内期单克隆抗体的研制和鉴定

用Ｐｅｒｃｏｌｌ密度梯度离心法从志愿者血液中分离出感染间日疟原虫的红细胞。以此作为免疫原，免疫Ｂａｌｂ／ｃ小鼠，获得５株杂交瘤细胞系。融合率平均为４５％，抗体阳性率为０．６６％，用ＩＦＡ和ＩＰＡ染色进行免疫组化分析，４Ｂ２和８Ｅ３分泌的单克隆抗体呈较强的阳性反应，与约氏疟原虫红内期无交叉反应。     

血清HBcAg检测与e系统意义的再研究

用商品试剂ＲＩＡ法检测５２１份乙肝患者血清ＨＢｃＡｇ，并与ＨＢｅＡｇ，抗ＨＢｅ及ＰＣＲＨＢＶＤＮＡ比较。结果表明ＨＢｃＡｇ的检出率：ＨＢｃＡｇ阳性组为８５．１２％；抗ＨＢｅ阳性组为８１．３６％，二者经统计学处理无显著差异。而ＰＣＲ检测两组ＨＢＶＤＮＡ阳性率依次为８８．８９％，４０．００％。提示：用ｅ系统衡量ＨＢＶ复制与否已不完全，应将ＨＢｃＡｇ与其它免疫学指标同时作常规测定。     

恶性疟原虫多价重组疫苗的研制及其免疫活性的初步鉴定

为了探讨含Ｔ、Ｂ淋巴细胞双重激活表位的重组复合症疾抗原的免疫原性和保护作用，作者设计合成了编码恶性疟原虫红内期３个保护性抗原位点和２个外源性Ｔ细胞激活位点的复合基因ＨＧＦＣ，并构建了多连体复合基因ＨＧＦＣＡＣ。两种复合基因分别与表达载体ｐＷＲ４５０－１重组，并在大肠杆菌中表达出含外源蛋白和β－半乳糖苷酶部分氨基酸的融合蛋白（分子量分别为６５０００和７７０００），表达量为３５％。表达产物可与小鼠及兔抗恶性疟原虫抗体发生特异性免疫反应。用纯化的融合蛋白免疫家兔制备的免疫血清可特异地识别恶性疟原虫抗原，并对疟原虫体外生长有显著抑制作用。抑制程度与免疫血清浓度及作用时间呈正相关，在２０％浓度下作用７２小时，抑制率可达８２％，并引起疟原虫发育不良和死亡。研究结果说明，制备的恶性疟原虫重组复合抗原具有免疫原性和一定的免疫保护作用，可作为恶性疟疫苗候选物。     

PCR与ELISA法检测乙肝血清标志物对比分析

用ＰＣＲ技术检测４８例肝病患者，５５例非肝病患者血清ＨＢＶＤＮＡ，ＥＬＩＳＡ法检测其血清五项标志物，肝病组ＰＣＲ阳性率为３１．３％，非肝病组ＰＣＡ阳性率为１１．４％。ＨＢｓＡｇ、ＨＢｃＡｂ阳性组ＨＢＶ－ＤＮＡ１００％阳性；ＨＢｓＡｇ、ＨＢｃＡｂ阳性、ＨＢｅＡｂ阳性或阴性组ＨＢＶ－ＤＮＡ阳性５０％；单项ＨＢｓＡｂ阳性组ＨＢＶＤＮＡ阳性２０％；五项标志物全阴ＨＢＶ－ＤＮＡ阳性４．７％。结果表明，ＰＣＲ技术特异性强、敏感度高，但两种方法各有所长，不宜取代。     

乙型病毒性肝炎的基因诊断

对１２０例乙肝病毒（ＨＢＶ）感染者血清中ＨＢＶ－ＤＮＡ用ＰＣＲ技术进行检测，结果表明，ＰＣＲ法的敏感性阳性检出率７５．８％，明显高于ＥＬＩＳＡ法（ＨＢｓＡｇ、ＨＢｅＡｇ和ＨＢＣＡｂ）的阳性率分别为６１％、３８％和６２％）；ＰＣＲ检测ＨＢｓＡｇ和ＨＢＡｂ一血清的ＨＢＶ－ＤＮＡ，阳性率分别为２６％、５２％和２４％，ＰＣＲ检测三者均为阳性血清的ＨＢＶ－ＤＮＡ，阳性率为１００％慢性迁延肝炎、慢性活动性肝炎     

表达疟原虫裂殖子表面蛋白1减毒鼠伤寒杆菌的稳定性

目的：检测含恶性疟原虫裂殖子表面蛋白１基因片段的重组减毒鼠伤寒杆菌Ｘ４０６４（ｐＱＥＭ１）的入侵宿主能力，所含质粒的稳定性和再表达能力。方法：用含恶性疟原虫裂殖子表面蛋白１第１号基因片段的重组减毒鼠伤寒杆菌Ｘ４０４６（ｐＱＥＭ１）灌胃接种ＢＡＬＢ／ｃ...     

疟原虫感染小鼠血清对L_(929)细胞的毒性作用

采用Ｌ９２９细胞作为靶细胞，对约氏疟原虫感染小鼠血清的细胞毒活性（ＣＡ）进行了测定。结果：持续疟原虫感染鼠血清在感染后第３ｄ即出现轻度的ＣＡ，至感染后第８～１５ｄＣＡ达到较高水平（３５％～４０％），以后逐渐下降，１个月时仍有２０％的ＣＡ。仅感染疟原虫７ｄ的鼠血清，高水平ＣＡ维持时间较短，感染后１５ｄＣＡ尚存留１７．８％，１个月后仅有６％。在小鼠感染疟原虫后第８ｄ注射大肠杆菌内毒素（ＬＰＳ），可迅速增强感染鼠血清的ＣＡ，此活性在注射ＬＰＳ后６０ｍｉｎ达高峰，１５０ｍｉｎ后下降至未注射ＬＰＳ的持续感染鼠水平。提示：疟原虫感染鼠血清具有一定的抗肿瘤细胞活性，此活性可以被ＬＰＳ协同增强。     

直肠癌脱落细胞PNA受体表达的研究

用荧光素标记的ＰＮＡ和ＤＢＡ检查直肠癌脱落细胞是否存在ＰＮＡ受体，共取直肠癌脱落细胞６５例，非直肠癌病人的正常直肠脱落细胞３２例，均用ＰＮＡ和ＤＢＡ染色，结果６５例直肠癌脱落细胞５９例ＰＮＡ阳性（９０．７７％），其中，强阳性２７例（４１．５４％），阳性２４例（３６．９２％）弱阳性８例（１２．３％）ＤＢＡ染色４例阳性（６．１５％），正常直肠脱落细胞３２例ＰＮＡ和ＤＢＡ染色均为阴性。表明直肠癌脱落细胞     

γ－氨基丁酸及荷包牡丹碱对大鼠脑片旁巨细胞外侧核神经元自发放电的影响

本研究用多管微电极技术在大鼠脑片旁巨细胞外侧核（ＰＧＣＬ）区观察了微电泳γ－氨基丁酸（ＧＡＢＡ）及其拮抗剂荷包牡丹碱（ＢＩＣ）对神经元自发放电的效应及ＢＩＣ对ＧＡＢＡ效应的影响。ＧＡＢＡ和ＢＩＣ对自发放电均有兴奋、抑制和无影响三种效应，各占所测试神经无数的５６．１０％，３１．７０％，１２．２０％和６６．６７％，５．８８％，２７．４５％。ＧＡＢＡ的抑制效应和ＢＩＣ的兴奋效应显示量效依赖关系。ＢＩＣ可阻断大多数被试神经元（７８．２６％）对ＧＡＢＡ的反应，本结果从细胞水平上提示ＰＧＣＬ区有内源性ＧＡＢＡ，其神经元上有ＧＡＢＡ＿Ａ受体。     

B细胞系特异性单克隆抗体ZCH－4－2E8的制备鉴定及其初步应用

本文报告一种识别Ｂ细胞系白血病相关抗原的鼠单克隆抗体ＺＣＨ－４２Ｅ８（ＩｇＧ１），免疫印迹试验示其所识别的抗原分子量为２４５ｋｄ。经多种细胞系、正常外周血细胞成分、骨髓单个核细胞、胚胎肝、脾、胸腺和１３８例急、慢性白血病细胞的鉴定结果表明，２Ｅ８为Ｂ细胞系特异性单抗，其反应谱与ＣＤ１９类似，但其尚能识别早至１８周龄的胚胎脾（Ｂ）细胞，阳性率为３１．９％（ＣＤ１９为０％）。作者认为：２Ｅ８是一种能用于分化早期Ｂ细胞系急性淋巴细胞白血病（ＡＬＬ）的诊断和急性未分化型白血病（ＡＵＬ）鉴别诊断的Ｂ细胞系特异性单抗。     

问号状钩端螺旋体外膜单克隆抗体凝集反应特性的研究

Three McAb were produced against an outer envelope preparation from Leptospira, interrogans, serovar Lai by fusion of SP2/0 myeloma cells with immune BALB/c mice spleen cells. The fusion rate was 96% and the antibody positive rate was 50%. One of the hybridomas, E4B11C9, reacted with 13 of the 13 serovars of the Icterohaemorrhagiae serogroup in microscopic agglutination test (MAT) but did not react with the 18 representative serovars of L. interrogans and L. biflexa serovar patoc and Leptonema illini. For all non-reactive serovars the MAT titres were greater than 1:25. The McAb, E4B7G5, reacted similarly with all serovars except smithi and tonkini. E4B7D4 reacted also similarly with all serovars except serovars birkini, ndambari, bogvere, smithi and tonkini. Therefore, 3 McAb showed serogroup specificity and partial serogroup specificity by agglutination. The agglutination titres were high and hybridomas were stable, so it might be useful in providing a simple, rapid method for the classification and identification of clinical isolates such as pathogenic L. interrogans in place of the complicated and time-consuming conventional methods.     

抗—P^30单克隆抗体杂交瘤细胞株的建立

BALB/c mice were immunized with the purified p30 antigen. SP2/0 myeloma cells and immune spleen cells were fused with 50% PEG (Sigma, MW 3,350-4,000). The cell fusion rate was 93.95%, and the antibody producing rate 21.01%. The technique of limiting dilution was used for cloning of the hybridoma cells. Two hybridoma cell lines E8 and G1 secreting McAb against p30 antigen were obtained. The number of chromosome of both E8 and G1 cell lines was 98.7 +/- 6.54 and 97.7 +/- 7.77, respectively. Results of the PAGE of the ascites generated by the hybridoma cell lines E8 and G1 showed a thick protein band at the gamma-region which was absent from the ascites generated by the SP 2/0 myeloma cells. The immunoglobulin of the McAb E8 and G1 belonged to IgG2 subclass. The results of the immunohistochemical method using the horseradish peroxidase conjugated antibody showed that both E8 and G1 McAb only reacted with epithelial cells of the normal human prostatic glands and their ducts, but did not cross-react with other thirty-four different kinds of normal human tissues. The results of inhibiting ELISA test showed that both E8 and G1 McAb were only inhibited by the human seminal plasma and p30, weakly inhibited by the adult male's urine, but were not inhibited by other eight different kinds of human body fluids and secretions as well as semen from eight different species of animals. It was concluded that McAb of E8 and G1 were organ and species specific.(ABSTRACT TRUNCATED AT 250 WORDS)     

抗人结肠癌单克隆抗体细胞株的建立及其免疫组化研究

本文报道了抗人结肠癌单克隆抗体3E_4细胞株的建立方法。其上清液与各种人癌细胞株及正常组织细胞等反应均阴性,仅与AFP似有弱交叉反应;经免疫组化研究,3E_4株单克隆抗体对结肠癌粘膜组织有一定选择性,但与癌旁正常结肠组织仍有一定交叉反应,属于抗结肠癌相关抗原的单克隆抗体。     

抗腺病毒载体单克隆抗体的制备及其初步鉴定

目的 研制抗腺病毒载体(Adv)表面蛋白的单克隆抗体(McAb).方法 将Adv以Al(OH)3佐剂化,常规免疫BALB/c小鼠.采用细胞融合技术制备抗Adv McAb;采用ELISA、免疫细胞化学及Western blot法对McAb进行筛选与鉴定.结果 共获得6株可稳定分泌特异性McAb的细胞(A4H11、A8C7、F1H5、G1D2、G4E3、H2G8),其染色体数目为102～106条,所分泌McAb均属IgG1亚类,持续3个月适应培养仍能保持其稳定的抗体分泌能力.腹水抗体的ELISA效价在1∶106～1∶108之间;Western blot结果显示6株McAb均与来自腺病毒载体及野生3、5、7型腺病毒抗原提取物中分子量约为114 kD的多肽反应,据此推测它们均是抗ADV-TK六邻体蛋白的McAb.结论 成功制备出6株抗腺病毒载体六邻体单克隆抗体,为腺病毒载体应用于肿瘤基因治疗的临床前研究提供物质基础.     

钩端螺旋体澳洲群体特异性McAb的建立,鉴定及群特异性...

Spleen cells of BALB/c mice immunized with whole Leptospira interrogans serogroup Australis serovar australis strain 620 were fused with myeloma cells line SP2/0. Specificities of four McAbs determined by MAT. 2E1 McAb (IgG3) reacted with 11 serovars, of the Australis serogroup, but did not react with 22 representative serovars of L. interrogans in 20 serogroups, L. biflexa strain patoc I and Leptonema illini strain 3055. 2E1 McAb showed serogroup specificity for Australis by agglutination and the other 3 McAbs showed partial serogroup specificity. We compared the outer envelope (OE) protein profiles of serovar australis strain 620 with those of two pathogenic L. interrogans serovar lai strain 601 and serovar hebdomadis strain 156 by SDS-PAGE. 63kd protein profile was only found in the OE of strain 620, and the quantity of 42kd protein of strain 620 was greater than that of strain 601 and 156. The immunoblotting revealed that 2E1 McAb reacted with a 34kd band in the OE preparation of serovar australis strain 620, but did not react with that of other two L. interrogans. 2E1 McAb also did not react with OE of non-pathogenic leptospires. It was suggested that 34kd protein might contain the antigenic determinants which were shared by leptospires of Australis serogroup.     

抗食管癌单克隆抗体的免疫组织化学研究

以人食管癌109细胞株免疫BALB/c小鼠获得两株抗食管癌杂交瘤,分别分泌单克隆抗体(McAb)2GF_3和2GF_4。以上述两种McAb的一种或将该二者混合,用ABC染色法检测食管癌和正常组织的石蜡切片,发现该两种McAb对不同组织类型或分化程度不同的食管癌细胞有不同的反应性;而且,应用两种McAb混合液的,可增高在食管癌总体标本中的检测阳性率。     

钩端螺旋体七日热群部分群特异McAb的建立与鉴定

Hebdomadis is one of the major serogroups found in China. In Sichuan, this serogroup appears to have a close relationship with local outbreak of leptospirosis. BALB/c mice were immunized intrasplenically with outer envelope of serogroup Hendomadis serovar hebdomadis strain 245. Spleen cells were fused with SP2/0 myeloma cells, two monoclonal antibodies Af2 and Bb2 were produced by hybridoma technique. McAb Af2 and McAb Bb2 were identified to be IgG1 and IgG3 by immunodiffusion respectively. Specificities of these two McAbs were determined by MAT; both reacted to 8 serovars (hebdomadis, nona, kambale, kremastos, worsfoldi, jules, maruborincana) of Hebdomadis serogroup. The agglutination titres of McAb Af2 and McAb Bb2 were 1:640-1:2,500,000 and 1:320-1:2,500,000, respectively. The two McAbs did not agglutinate with serovar kabura of Hebdomadis serogroup, they did not agglutinate with 11 serovars of Sejroe serogroup, 4 serovars of Mini serogroup, 18 representive serovars of L. interrogans in 18 serogroups, L. biflexa strain Patoc I and Leptonema illini. So it was found that McAb Af2 and McAb Bb2 showed partial serogroup specificity for Hebdomadis by agglutination.     

抗EB病毒膜抗原单克隆抗体的制备及鉴定

本文将sp_2/o细胞与经脾内注射用0.2mMH2O2激活的B95—8全细胞免疫的BALB/C小鼠脾细胞,用50％PEG4000进行融合,其平均融合率为51.79%,抗体分泌为43.48％。经亚克隆后获得一株能持续分泌抗EB病毒脱抗原的单克隆抗体(以下简称抗EBV—MA单抗)的杂交瘤细胞—B3B7,该细胞的染色体数为111.7±4.76。经ELISA法,间接免疫酶法和间接免疫荧光法证实该单抗不能与EBV—MA(一)的人Rajì细胞发生特异性免疫反应,只能与EBV—MA(+)的人P3HR—1和绒猴的B95—8细胞的反应,而且与H_2O_2激活的B95—8和P_3HX—1细胞的反应强度非常显著地高于与未经H_2O_2激活的B95—8和P_3HR—1细胞的反应强度(P<0.001),同时亦证实特异性荧光主要集中在EBV—MA(+)细胞的细胞膜上,从而表明B_3B_7确实是能分泌抗EBV—MA单抗的杂交瘤细胞。