应用双抗体夹心ELISA定量检测鼠疫F1单克隆抗体IgG含量

目的建立一种用于定量检测鼠疫单克隆抗体样品中IgG含量的方法 ,以更好地指导杂交瘤细胞的筛选和抗体的生产、纯化。方法采用羊抗鼠IgG为包被抗体、辣根过氧化物酶标记的羊抗鼠IgG为标记抗体,纯化鼠IgG作参考品,制作标准曲线,测定未知样品中的IgG含量。结果该方法与BCA蛋白浓度测定法测得的结果基本一致,且重复性好,特异性强,灵敏度高。结论双抗体夹心ELISA法可用于测定鼠疫F1单克隆抗体样品中的IgG含量。     

鼠疫F1McAb酶免疫染色法快速检验鼠疫苗

为缩短检验鼠疫菌的时间，建立了酶免疫染色法检验鼠疫菌的方法，被检涂片标本用含过氧化物酶底物的固定液，固定标本同时，内源性过氧化物酶被消耗，辣根过氧化物酶标记鼠疫F1单克隆抗体和鼠疫菌菌体表面的特异性F1抗原结合，底物被酶催化生成有色沉淀物使鼠疫菌着色，用普通光学显微镜观察结果，常温1h出结果，Kaplow法鼠疫菌被蓝色晶体覆盖，DAB法鼠疫菌呈棕黄色，不含F1抗原的8株细菌均为阴性，酶免疫染色法具有特异，快速，简便的特点，可用于鼠疫菌的快速检验。     

鼠疫F1McAb酶免疫染色法快速检验鼠疫菌

为缩短检验鼠疫菌的时间 ,建立了酶免疫染色法检验鼠疫菌的方法 ,被检涂片标本用含过氧化物酶底物的固定液 ,固定标本同时 ,内源性过氧化物酶被消耗 ,辣根过氧化物酶标记鼠疫 F1单克隆抗体和鼠疫菌菌体表面的特异性 F1抗原结合 ,底物被酶催化生成有色沉淀物使鼠疫菌着色 ,用普通光学显微镜观察结果 ,常温 1h出结果 ,Kaplow法鼠疫菌被蓝色晶体覆盖 ,DAB法鼠疫菌呈棕黄色 ,不含 F1抗原的 8株细菌均为阴性 ,酶免疫染色法具有特异、快速、简便的特点 ,可用于鼠疫菌的快速检验     

分泌抗鼠疫耶尔森菌F1抗原单克隆抗体杂交瘤株的建立

目的应用杂交瘤技术,制备分泌抗鼠疫F1抗原单克隆抗体的杂交瘤细胞株。方法用鼠疫菌F1抗原免疫雌性BALB/c小鼠,检测小鼠血清中的F1抗体,选抗体滴度最高的小鼠用于细胞融合,融合前3天F1抗原经腹腔注射加强免疫1次,然后取其脾脏细胞与处于对数生长期的SP2/0骨髓瘤细胞混合,50%的PEG作为细胞融合剂,HAT培养液选择培养融合后的细胞,间接ELISA法检测细胞培养上清中的F1抗体,阳性杂交瘤细胞用有限稀释法进行克隆及再克隆。结果获得共计100余株分泌F1单克隆抗体的杂交瘤细胞株,3株进行了4次克隆化,经8个月保存后抗体分泌稳定。结论应用杂交瘤技术,在云南首次成功制备了3株能稳定分泌抗鼠疫杆菌F1抗原的单克隆抗体杂交瘤细胞株,建立了杂交瘤技术的实验程序。     

双抗夹心ELISA检测犬粪贾第虫抗原方法的建立及应用

目的建立简便、特异的检测犬粪贾第虫抗原贾第素α-12的双抗夹心ELISA方法。方法利用辣根过氧化物酶标记抗贾第素α-12单克隆抗体4C9,通过过滤、超声的方法处理犬粪样品。以抗贾第素α-12单克隆抗体3H10为捕获抗体,5%脱脂奶粉作为封闭剂,辣根过氧化物酶标记的抗贾第素α-12单克隆抗体4C9为检测抗体,经底物显色后,用酶标仪测量490nm处的吸光度（A）值。通过棋盘滴定法确定抗体最适包被浓度、最佳封闭剂、待检粪液最佳稀释度及酶标抗体最佳稀释度。应用建立的ELISA方法对122份犬粪样品进行检测。结果建立的双抗夹心ELISA检测标准阳性样品的A490为0.465,标准阴性样品的A490为0.098;优化的最佳检测条件为：抗体包被浓度为0.01mg/ml,封闭剂为1%BSA,粪液稀释度为1∶2,酶标二抗稀释度为1∶400,该试验与犬等孢球虫、犬隐孢子虫、犬蛔虫样品均无交叉反应,批间和批内变异系数较小,重复性好。应用本方法对122份犬粪进行检测,阳性率为36.9%。结论建立的双抗夹心ELISA方法特异性高,重复性好,为贾第虫流行病学调查提供了一种更简便、快速、特异的免疫学检测方法。     

抗辣根过氧化物酶和猪囊尾蚴的双特异单克隆抗体的研制

抗辣根过氧化物酶和猪囊尾蚴的双特异单克隆抗体的研制李达，沈喜，任永卫双特异单克隆抗体（Ｂｉｓｐｅｃｉｆｉｃｍｏｎｏｃｌｏｎａｌａｎｔｉｂｏｄｉｅｓ）是采用杂交瘤技术、化学偶联或基因工程方法制备的能够识别两种抗原的抗体或抗体复合物。近年来双特异单克隆抗...     

人血清真胰岛素酶联免疫法的建立及初步应用

人血清真胰岛素酶联免疫法的建立及初步应用黎明吴从愿宋爱玲张葵摘要选用两种可相互配对的抗人胰岛素（ｈＩｎｓ）单克隆抗体，其中一种包被塑料孔条作固相抗体，另一种经生物素化后作液相抗体，再以辣根过氧化物酶标记的亲和素放大检测信号，建立了高度特异和灵敏的人血...     

鼠疫F1单克隆抗体的胶体金标记及斑点渗滤法检测F1抗原方法的建立

目的建立斑点渗滤法快速检测鼠疫F1抗原的方法。方法粗提鼠疫F1单克隆抗体,在pH8.2下对其进行胶体金标记,以兔抗F1抗体与金标记的单克隆抗体建立斑点渗滤快速检测鼠疫F1抗原的方法,并对该方法进行特异性与敏感性试验。结果成功建立斑点渗滤法快速检测鼠疫F1抗原的方法,该检测可在15min内完成,敏感性为3.12mg/L,对假结核、小肠结肠炎等近缘菌的特异性实验未发现交叉反应。结论该方法可作为鼠疫快速诊断的手段,但实验敏感性有待进一步提高。     

慢性肾功能衰竭患者血清氧化型低密度脂蛋白含量的测定及与血脂的相关性(摘要)

１　对象和方法慢性肾功能衰竭患者３０例，男性１８例，女性１２例，平均年龄３５．８±１０．５岁。正常对照者３０例，按年龄、性别与病例组相匹配。取隔夜空腹血清作血脂测定，其余血清加适量ＥＤＴＡ，冰箱冻存，用以测定ｏｘＬＤＬ含量。抗人氧化型低密度脂蛋白单克隆抗体（ＨＯＬ１）包被聚乙烯板，４℃冰箱过夜，３７℃封闭２ｈ。按改良过碘酸钠法，用辣根过氧化物（ＨＲＰ）标记ＨＯＬ１。将１００μＬ血清加入已包被ｏｘＬＤＬ单克隆抗体的酶标反应板中，３７℃水浴２ｈ，用０．０５％Ｔｗｅｅｎ２０－ＰＢＳ洗３遍，每遍３ｍ…     

酶标记抗原对流免疫电泳试验诊断棘球蚴病的研究

用戊二醛一步法以辣根过氧化物酶标记纯化人棘球蚴液抗原,对80例棘球蚴病,97例其它寄生虫病和其它疾病以及74例健康人进行了酶标记抗原对流免疫电泳试验(E-CIEP),其敏感性为85.0％,特异性为100％。E-CIEP敏感性显著高于CIEP 而低于PPA-ELISA,但其特异性显著高于PPA-ELISA。用国产辣根过氧化物酶与进口酶标记的抗原,试验敏感性相同。凝胶片先烘干后染色并不影响抗体活性。     

胰岛素瘤的解剖分布特点分析

目的胰岛素瘤是最常见的胰腺神经内分泌肿瘤，因其临床表现多样，导致诊断困难。影像学诊断尤其是超声内镜(EUS)在胰岛素瘤的诊断中起着重要作用，拥有较高的敏感性和特异性。本研究拟通过明确胰岛素瘤的解剖分布特点，以期有助于提高影像学的诊断准确率和降低漏诊率，尤其是在教育和培训实践中对于EUS的学习者更具有指导价值。 方法回顾性分析解放军总医院第一医学中心病案资料数据库1993年1月至2019年11月经外科手术、病理确诊为胰岛素瘤的患者的临床资料，检索方法采取搜索术后病理诊断为"胰岛素瘤"的病例，通过查阅病例的方法，提取出胰岛素瘤的大小和解剖分布等数据，进一步分析其特点。 结果共检索到确诊为胰岛素瘤的患者116例，其中，男45例、女71例，年龄13～76岁，平均年龄(44.4±14.85)岁。胰岛素瘤单发110例(94.8%)、多发6例(5.2%)。位置分布：头颈部46例(39.7%)，单发45例、多发1例；体尾部68例(58.6%)，单发65例、多发3例；全胰腺多发2例(1.7%)。病变大小特点：最大径0.4～3.4 cm，平均大小(1.53±0.58)cm。≤1 cm 29例、>1 cm而≤1.5 cm41例、>1.5 cm而≤2.0 cm28例，≤3 cm 15例，>3 cm 3例。年龄与肿瘤的大小相关，≤44岁患者肿瘤平均大小为(1.36±0.51)cm、>44岁患者肿瘤平均大小为(1.70±0.60)cm，P<0.05。头颈部的肿瘤大于体尾部的肿瘤，头颈部肿瘤平均大小(1.66±0.63)cm，体尾部(1.42±0.52)cm，P<0.05。 结论胰岛素瘤在胰腺体尾部较头颈部更好发；绝大多数单发，但可以全胰腺多发；多数小于1.5 cm，肿瘤的大小与患者年龄和肿瘤的解剖分布相关。     

Pathogenesis of carcinoma of the papilla of Vater

Most adenomas and carcinomas of the small intestine and extrahepatic bile ducts arise in the region of the papilla of Vater. In familial adenomatous polyposis (FAP) it is the main location for carcinomas after proctocolectomy. In many cases symptoms due to stenosis lead to diagnosis at an early tumor stage. In about 80%, curative intended resection is possible. Operability is the most relevant prognostic factor. Most ampullary carcinomas resp. carcinomas of the papilla of Vater develop from adenomatous or flat dysplastic precursor lesions. They can be sited in the ampulloduodenal part of the papilla of Vater, which is lined by intestinal mucosa. They also can develop in deeper parts of the ampulla, which are lined by pancreaticobiliary duct mucosa. Intestinal-type adenocarcinoma and pancreaticobiliary-type adenocarcinoma represent the main histological types of ampullary carcinoma. Furthermore, there exist unusual types and undifferentiated carcinomas. Many carcinomas of intestinal type express the immunohistochemical marker profile of intestinal mucosa (keratin 7?, keratin 20+, MUC2+). Carcinomas of pancreaticobiliary type usually show the immunohistochemical profile of pancreaticobiliary duct mucosa (keratin 7+, keratin 20?, MUC2?). Even poorly differentiated carcinomas, as well as unusual histological types, may conserve the marker profile of the mucosa they developed from. These findings underline the concept of histogenetically different carcinomas of the papilla of Vater which develop either from intestinal- or from pancreaticobiliary-type mucosa of the papilla of Vater. Molecular alterations in ampullary carcinomas are similar to those of colorectal as well as pancreatic carcinomas, although they appear at different frequencies. In future studies, molecular alterations in ampullary carcinomas should be correlated closely with the different histologic tumor types. Consequently, the histologic classification should reflect the histogenesis of ampullary tumors from the two different types of papillary mucosa.     

Lack of correlation of degree of villous atrophy with severity of clinical presentation of coeliac disease

BACKGROUND AND AIM: Both the clinical presentation and the degree of mucosal damage in coeliac disease vary greatly. In view of conflicting information as to whether the mode of presentation correlates with the degree of villous atrophy, we reviewed a large cohort of patients with coeliac disease. PATIENTS AND METHODS: We correlated mode of presentation (classical, diarrhoea predominant or atypical/silent) with histology of duodenal biopsies and examined their trends over time. RESULTS: The cohort consisted of 499 adults, mean age 44.1 years, 68% females. The majority had silent coeliac disease (56%) and total villous atrophy (65%). There was no correlation of mode of presentation with the degree of villous atrophy (p=0.25). Sixty-eight percent of females and 58% of males had a severe villous atrophy (p=0.052). There was a significant trend over time for a greater proportion of patients presenting as atypical/silent coeliac disease and having partial villous atrophy, though the majority still had total villous atrophy. CONCLUSIONS: Among our patients the degree of villous atrophy in duodenal biopsies did not correlate with the mode of presentation, indicating that factors other than the degree of villous atrophy must account for diarrhoea in coeliac disease.     

Demonstration of the mechanism of action of biguanides

Summary Palmitic acid oxidation in rat diaphragm homogenate is depressed by biguanide concentrations that are still incapable of inhibiting oxidative phosphorylation. Glucose oxidation is not directly effected by the same biguanide concentrations: however, the inhibitory effect of palmitic acid on glucose oxidation is partly removed by biguanides. Inhibition of fatty acid oxidation, which accounts for most of the metabolic effects caused by these drugs, can be regarded as the fundamental mechanism of action of biguanides. There is some evidence suggesting that these drugs might interact with carnitine, thus preventing long-chain fatty acids from being transported across the mitochondrial membrane to the site of oxidation. Traduzione a cura degli AA.     

血吸虫童虫生长发育及其机制的研究进展

血吸虫童虫是宿主免疫系统攻击的重要靶标,包括皮肤型、肺型和肝门型童虫。宿主分子对童虫生长发育具有重要作用。童虫生长发育机制包括免疫调节、信号转导、性别发育及凋亡等。肌动蛋白、组织蛋白酶、烯醇化酶和葡萄糖基转移酶等分子为血吸虫童虫生长发育的重要分子。本文对血吸虫童虫生长发育及其机制的研究进展做一综述。     

124例耐多药结核分枝杆菌基因突变特征分析

目的对临床分离的耐多药结核分枝杆菌相关基因的突变特征进行分析。方法对124例耐多药结核分枝杆菌以及50株敏感株的耐药相关基因(包括异烟肼inh A、kat G、oxyR-ahp C间隔区以及利福平rpo B)进行序列测定,分析其基因突变情况。结果异烟肼耐药inh A基因突变率为14.5%;kat G基因突变率为70.2%(87/124),主要位于315位;oxyR-ahp C间隔区突变率为15.3%;inh A、kat G两种基因同时突变率75.0%,三种基因同时突变率为89.5%。利福平rpo B基因突变的检出率高达95.2%,突变主要发生在531、526、516位点。结论我省耐多药菌异烟肼耐药相关基因最常见突变为kat G 315、inh A C-T(-15)、axyR-ahp C间隔区(-10)C-T,利福平为rpo B531、526、516。结合MDR-TB耐药相关基因的特征分析,可以建立一种快速、准确、特异的适合于我省的检测结核菌耐多药性的新方法。     

Assessment of quality of life of parents of children with juvenile chronic arthritis

The aim of the study was to assess the quality of life (QOL) and the psychological status of parents of children with juvenile chronic arthritis (JCA). The QOL, anxiety and depression of the parents of 28 children with JCA were evaluated and compared to those of the parents of 28 healthy children. Mothers of JCA children and mothers of healthy children reported similar QOL. The reported anxiety and depression levels were similar for mothers and fathers in both groups. The parents of children with pauciarticular-type JCA reported lower QOL and higher levels of anxiety and depression than the parents of children with other types, namely polyarticular and systemic JCA. These findings may be explained by the fact that the pauciarticular patients had shorter disease duration and were less frequently seen in the outpatient clinic. The QOL of mothers of children with JCA was found to be slightly impaired in the group of children with pauciarticular JCA. Future larger studies are needed to confirm these results, as the number of subjects in the three groups was rather low. Received: 26 September 2001 / Accepted: 8 February 2002     

Numerical Simulation of the Effect of Rate of Change of Glucose on Measurement Error of Continuous Glucose Monitors

Background

A 5-day in-patient study designed to assess the accuracy of the FreeStyle Navigator® Continuous Glucose Monitoring System revealed that the level of accuracy of the continuous sensor measurements was dependent on the rate of glucose change. When the absolute rate of change was less than 1 mg•dl⁻¹•min⁻¹ (75% of the time), the median absolute relative difference (ARD) was 8.5%, with 85% of all points falling within the A zone of the Clarke error grid. When the absolute rate of change was greater than 2 mg•dl⁻¹•min⁻¹ (8% of the time), the median ARD was 17.5%, with 59% of all points falling within the Clarke A zone.

Method

Numerical simulations were performed to investigate effects of the rate of change of glucose on sensor measurement error. This approach enabled physiologically relevant distributions of glucose values to be reordered to explore the effect of different glucose rate-of-change distributions on apparent sensor accuracy.

Results

The physiological lag between blood and interstitial fluid glucose levels is sufficient to account for the observed difference in sensor accuracy between periods of stable glucose and periods of rapidly changing glucose.

Conclusions

The role of physiological lag on the apparent decrease in sensor accuracy at high glucose rates of change has implications for clinical study design, regulatory review of continuous glucose sensors, and development of performance standards for this new technology. This work demonstrates the difficulty in comparing accuracy measures between different clinical studies and highlights the need for studies to include both relevant glucose distributions and relevant glucose rate-of-change distributions.     

Angiographic diagnosis of the degree of serosal invasion of carcinoma of the colon

Angiography using Prostaglandin El^® was performed on 38 patients with carcinoma of the colon in order to diagnose the degree of serosal cancer invasion. The findings at angiography were classified into four groups:1) AG-S3, abnormal change (irregularity and/or encasement) up to marginal vessels; 2) AG-S2, abnormality up to vasa recta; 3) AG-S1, abnormality of penetrating branches of vasa recta within the wall of the colon; and 4) AG-S0, no distinct findings of abovementioned vessels. These angiographic findings were compared with both macroscopic and microscopic serosal cancer invasion. Angiographic diagnosis is in accord with the macroscopic findings in 84.2 percent of cases. Angiographic diagnosis is in accord with the microscopic findings in 32.4 percent of cases. Macroscopic findings confirm the angiographic diagnosis precisely but the conflict with microscopic findings should not be overlooked. This may be the result of inflammatory change, adhesion, and fibrosis around carcinoma of the colon.     

Study of constancy of hydrogen-consuming flora of human colon

The constancy of the hydrogen consuming flora of the human colon was studied in 15 healthy subjects via two measurements obtained 18 to 36 months apart. Hydrogen disappearance rate and the major products of H₂-consuming bacteria, methane and sulfide, were measured during incubation of fecal homogenates with excess hydrogen and sulfate. In 11/15, the hydrogen consumption rate and the predominant hydrogen-consuming pathway (methanogenesis, sulfate reduction, or neither) remained constant. However, major shifts in these pathways were observed in four subjects, with two losing and two gaining the ability to produce methane. Methanogenesis was associated with the highest hydrogen consumption rate. This study demonstrates that clinically unrecognizable, major alterations of the colonic flora occur in healthy subjects. Understanding of the factors responsible for these alterations might allow for therapeutic manipulation of the colonic flora.Supported in part by the Department of Veterans Affairs and NIDDKD RO1 DK 13309-25.