IgG4酶联免疫分析的建立及初步应用

用兔抗人IgG4的多克隆抗体(IgG4Ab)包被成固相板,以识别结合待测标本中的IgG4。用生物素标记IgG4Ab得BioIgG4Ab;用亲和素标记辣根酶得HRPA。在已包被IgG4Ab的固相板中加入IgG4标准品(或待测样品),反应、洗涤后,加入BioIgG4Ab,反应、洗涤后,加入HRPA,反应、洗涤后,板上形成IgG4Ab—IgG4—BioIgG4Ab—HRPA复合物,加酶底物显色,用酶标仪在490nm波长测定OD值,作标准曲线,根据标准曲线,查出标本中IgG4含量。该法测定范围:2.5～200ng/mL,最低检出量2.1ng/mL,批内和批间CV分别8.4%和11.2%。测得血清IgG4含量:青年人(30名)为37.7±2.3ng/mL;30名献血员为42.7±9.9ng/mL,49例重症监护患者明显升高为71.2±9.2ng/mL;49例肺部感染患者明显降低为28.4±9.2ng/mL。结果表明该法稳定,灵敏度适于检出人血清IgG4水平。     

IgG3酶联免疫分析方法的建立及初步应用研究

目的:建立IgG3酶联免疫分析(ELISA)方法,探讨IgG3在炎症性疾病患者血清中的水平变化.方法:用兔抗人IgG3的多克隆抗体包被成固相酶标板,以识别结合待测标本中的IgG3,再用此多克隆抗体标记生物素(Bio-Ab),用亲和素标记辣根过氧化物酶(HRP-A),向固相酶标板中加入标准品或待测品,反应、洗涤后,加入Bio-Ab,反应、洗涤后,加入HRP-A,反应、洗涤后,酶标板上形成Ab-Ag-Ab-Bio-A-HRP复合物,加酶底物显色,用酶标仪在相应波长下测定光密度(OD值),根据标准曲线,计算出待测标本中的IgG3含量.结果:该法测定范围为(1.875～60)ng/ml,最低检出量1.5ng/ml,批内和批间误差分析＜8%和10%.用该法测得正常青年人血清IgG3含量为(12.91±4.25)ng/ml(n=64),风湿病患者为(11.23±4.64)ng/ml(n=64),肺部感染患者为(9.32±2.00)ng/ml(n=24),后两组血清IgG3水平均显著低于正常青年人(P＜0.05).结论:我们建立的IgG3 ELISA方法稳定、简便,特异性和灵敏度高,适于检测人血清IgG3水平的变化.     

人血清脂联素酶联免疫吸附试验测定方法的建立及临床应用研究

利用自行制备的抗脂联素单克隆抗体（McAb）,建立双抗体夹心酶联免疫（ELISA）分析人血清脂联素水平,优化实验条件,评价检测方法,并将该方法进行初步的临床应用。用抗脂联素McAb包被酶标板,加入待测抗原,再加入酶标记抗体,DAB显色。方法的线性范围为1.25~40μg/mL,灵敏度为0.2μg/mL,批内、批间变异系数分别为5.4%和7.1%,回收率为92.0%。与进口试剂盒比较,相关性良好（r=0.921）。测定了150份正常人血清脂联素含量,测定值为10.2±3.9μg/mL;66例2型糖尿病患者血清,含量为7.3±4.2μg/mL。结果表明,该方法简便可行,标准曲线范围、灵敏度、特异性、重复性、准确性均符合临床质控要求,适用于基础研究和临床检验。     

白介素8酶联免疫分析方法及其初步应用

建立高灵敏度和特异性的白介素8(IL-8)ELISA法.用人工重组的IL-8多次免疫兔,获取高效价抗体.纯化抗体经生物素标记,同辣根过氧化酶(HRP)标记的亲和素可特异结合.采用纯化的抗体包被96孔板,形成固相抗体.加入不同浓度标准品后,反应1.5h,洗净后再加入生物素标记抗体,以酶标记亲和素作为探针,经酶底物显色反应可获得良好的ELISA标准曲线.该方法测定范围为0.2-10.8ng/mL,最低测出值为0.02ng/mL,批内和批间CV小于8%和10%.用该法测得18名正常人血浆中IL-8水平为0.15±0.06ng/mL.18份高低不同浓度IL-8水平的血清样品经本法和RIA测定其均值为0.54±0.35ng/mL和0.63±0.59ng/mL,相关性强(r=0.819,t=5.89).在U937细胞系体外培养液中,LPS刺激24h和48h可测出IL-8水平明显上升;而经TNF-α刺激24h和48h后同对照没有明显变化.该方法操作简便,具有较强灵敏度和特异性,可满足人血清和细胞培养液中IL-8水平的检测.     

肿瘤相关胰蛋白酶原-2免疫放射分析

本文建立了肿瘤相关胰蛋白酶原-2(TAT-2)IRMA,并初步应用于临床, 以探讨其对肿瘤早期诊断的可能性.以抗TAT-2 McAb T2C和T2L分别作为捕捉抗体和标记抗体.在pH9.6的碳酸缓冲液中, 将McAb T2C包被小珠, 成为固相抗体;用氯胺T方法将McAb T2L制成125I-标记抗体;并将两株McAb与不同量的TAT-2标准品, 在pH7.5的磷酸缓冲液中保温反应, 制作TAT-2标准曲线.检测肿瘤患者及正常人血清或尿液TAT-2含量, 并作对比.结果表明, 测定的TAT-2标准曲线(N=10):Bmax/T=(22±3.2)%, Bmax/B0=689.2±35.6倍, 非特异结合率小于0.5%, 检测下限在0.5ng/mL;批内CV为(4.3±0.26)%, 批间CV为(10.6±1.2)%;平均回收率为(98.5±3.1)%和(97.9±2.3)%.正常人血清(87份)和尿(20份)TAT-2浓度为7.58±5.46ng/mL和3.11±1.62ng/mL; 胃癌患者(45例)的血清TAT-2浓度63.2±62.9ng/mL, 胰腺癌和胆管癌患者(12例)和其它各种转移癌患者(16例)尿TAT-2浓度为236.6±185.3ng/mL, 104.9±171.2ng/mL;3组癌症患者血清、尿TAT-2浓度与正常人相比, 差别非常显著(P值均小于0.001).因此, TAT-2有希望成为一项肿瘤辅助诊断指标.     

弓形虫IgG时间分辨免疫荧光分析法的建立

建立了高灵敏弓形虫-IgG的时间分辨荧光免疫分析(TRFIA)检测方法.样品或参考标准加入到由弓形虫抗原包被的96孔微孔板上,用Eu3+标记羊抗人IgG,建立间接弓形虫-IgG TRFIA检测法.本法检测范围为0.7～200 U/mL, 灵敏度为0.7U/mL;方法的平均批内和批间CV分别为4.5%和5.6%;标准曲线可测范围为0.7～14U/mL. 本文建立的弓形虫-IgG的TRFIA法灵敏度高、稳定性好,具有良好的应用前景.     

血清白介素8固相放射免疫法的建立及应用

本方法将兔抗人白介素8(IL-8)特异性抗体IgG包被塑料试管内壁,建立了一种快速、简便、高灵敏、特异的固相竞争放射免疫分析法(RIA)测定人血清中的IL-8.标准曲线范围为0.2～9.6ng/mL,最小检出值和正常值分别为0.16 ng/mL和0.30±0.05ng/mL.批内CV%为6.8%,批间CV%为9.6%.本文所建的固相RIA法与液相RIA法平行测定58份样品的IL-8浓度(0.26～3.56 ng/mL),经统计学处理,两者之间具有良好的直线相关(r=0.954,P<0.001,Y=0.994X).表明本文建立的固相RIA法可以代替液相RIA法测定血清IL-8浓度,并适用于大批样品和自动化检测.     

脂肪酸结合蛋白放射免疫分析方法的建立及初步应用

用人工重组的人脂肪酸结合蛋白(FABP)多次免疫大耳白兔,获取高效价的FABP抗体.用氯胺T法制备125I-FABP,再经Sephdex G-25纯化;抗原抗体反应采用平衡一步法,4℃培养24h后,经PR试剂分离结合和游离的标记抗原.该法测定范围5-405ng/mL,最低检出量为9ng/mL,批内和批间CV分别小于6.4%和8.5%.用该法测得人血清FABP含量:健康人(45名)为15.68±2.9ng/mL;重症监护病人(46例)为72.08±32.64ng/mL;心脏病患者(73例)为78.95±24.83ng/mL,明显升高.结果表明,本法稳定,简便特异,其灵敏度适于检出人血清FABP水平的变化.     

广谱检测磺胺类药物的酶联免疫分析方法的建立

目的 利用抗磺胺类药物多抗建立酶联免疫分析方法,测定磺胺类药物在动物源食品中的残留.方法 以抗磺胺类药物抗体包被微孔板,辣根过氧化物酶(HRP)标记磺胺类半抗原磺胺噻唑(ST),磺胺间甲氧嘧啶为标准品建立酶联免疫分析方法.结果 本方法的标准曲线测定范围为0.3～ 100ng/mL.方法学鉴定结果表明,灵敏度为0.2ng/mL,批内、批间变异分别为9.2％ ～ 14.4％、10.5％～ 12.7％.牛奶、蜂蜜、水产品及鸡肉、鸡蛋样品的添加回收率分别在79.3％～88.9％、65.5％～136.4、63.0％～83.3％、59.3％～78.1％之间.牛奶稀释实验表明,测定值与稀释度呈线性相关,相关系数r=0.997.结论 本方法可同时快速测定动物源性食品中23种磺胺类药物的残留,对19种磺胺类药物的检测灵敏度高于10ng/mL.     

老年男性肾脏雄激素调节蛋白检测方法的建立及参考范围

目的 探索建立检测人尿和血清中肾脏雄激素调节蛋白(KAP)含量的ELISA试剂盒,并初步提出在老年男性中的参考值范围.方法 采用间接竞争ELISA法,筛选酶标亲和素及生物素化二抗的最适滴度.以KAP标准品对数值作为横坐标,加入抗体后测得的A450值作为纵坐标,绘制标准曲线.利用此试剂盒检测100例正常老年男性的尿和血清中KAP的含量,每组做3个复孔,间隔6个月后再重复1次.结果 包被原为0.5μg,生物素化抗体为1∶3 200,酶标亲和素的稀释度为1∶2000时所获的A450值最佳.间接竞争ELISA法标准曲线的绘制:以0.008～0.500 mg/L、0.250 mg/L、0.125 mg/L、0.063 mg/L、0.031 mg/L、0.016 mg/L、0.008 mg/L的对数值为横坐标,其对应的A450值为纵坐标绘制标准曲线,其线性关系良好(r=0.980).本试剂盒的灵敏度为0.008 ～ 0.5 mg/L.100例样本中尿KAP的含量为(0.042±0.031) mg/L,血清KAP的含量为(0.014±0.011)mg/L.老年男性尿及血清KAP参考值范围分别为(0.000～ 0.103) mg/L及(0.000～ 0.041) mg/L.组内差异为1.7％,组间差异为8％,每次检测耗时小于2h.结论 本试剂盒具有较高的灵敏度及特异性,检测速度较快.同时,我们提供了部分正常老年男性尿及血清中KAP的含量,为今后对KAP与疾病关系的研究提供了依据.     

Schizophrenia in a North Swedish geographical isolate, 1900–1977. Epidemiology, genetics and biochemistry

Over 200 schizophrenic patients belonging to three major and interrelated pedigree complexes have been investigated over the past 30 years in a North Swedish geographically isolated population, presently numbering about 6,000. An intensive investigation of a number of biochemical correlates and genetic markers in a few selected families belonging to one of the major pedigrees has indicated new strategies for the current research program.
Schizophrenia, as defined operationally, is significantly associated with decreased activities of two enzymes (1) blood platelet monoamine oxidase, (2) plasma dopamine-β-hydroxylase, and (3) with the genetic marker Gc² (group specific antigen). Both enzymes are subject to genetic variation. A positive score for linkage between schizophrenia and low plasma DBH activity has been calculated, but, so far, available data are insufficient for discrimination between linkage and partial contribution of genetically controlled low plasma DBH to the pathogenesis of the disease. Alternatively, both mechanisms could be involved.
As a model for continued research, schizophrenia is explained as based on a double dominant-recessive genotype (Aabb), representing a vulnerability which in about 50 % of cases develops into clinical schizophrenia. It is suggested that the dominant mutation (A) operates on or affects MAO activity, and that the recessive genotype (bb) is instrumental in low variates of DBH activity and very likely such variates within the normal range of physiological variation. Moreover, it is suggested that the combined effects of MAO- and DBH-reduced efficiency on the metabolism of e.g. dopamine could be an essential pathogenic mechanism for the schizophrenic illness which is segregating in this population.     

Renal dysplasia and asplenia in two sibs

A family is reported in which two sibs, one male and the other female, both died within 24 hours of birth with enlarged polycystic kidneys. Postmortem histology in the second child showed gross renal dysplasia. In both children the pancreas was enlarged, nodular and cystic but the liver appeared macroscopically normal. In the second child, histological examination confirmed pancreatic fibrosis with cystic dilation of ducts, but showed portal fibrosis with bile duct proliferation in the liver.
This combination of findings is very reminiscent of those in a girl and her brother reported by Ivemark et al. (1959). The children reported here also showed absence or hypoplasia of the spleen, cardiac anomalies and other features of the Ivemark syndrome (Ivemark 1955), a quite different, usually sporadic, congenital disorder. It is suggested that the children described here have a distinct lethal congenital disorder, probably inherited in an autosomal recessive manner.     

The dominant diseases of modernized societies as omega-3 essential fatty acid deficiency syndrome: Substrate beriberi

About 1900, modern food selection and processing caused widespread $\text{epidemics}$ of the B vitamin deficiency diseases of beriberi and pellagra which, for genetic reasons, often expressed as different diseases ranging from bowel and heart disease to dermatoses and psychoses. But the B vitamins merely help convert essential fatty acids (EFA) into the prostaglandin (PG) tissue regulators and it now turns out that, through hydrogenation, milling and selection of w3-poor southern foods, we have also been systematically depleting, by as much as 90%, a newly discovered trace Nordic EFA (w3) of special importance to primates and sole precursor of the PG3(4) series, even as a concurrent fiber deficiency increases body demand for EFA. Since substrate EFA is processed by many B vitamin catalysts, an EFA deficiency will mimic a $\text{panh}$ypovitaminosis B, i.e., a mixture of $\text{substrate}$ beriberi and $\text{substrate}$ pellagra resembling vitamin beriberi and pellagra but exhibiting as even more diverse $\text{endemic}$ disease. This would consitute a second stage of the Modern Malnutrition and explain why some workers now hold the dominant diseases of modermized societies to be new, nutritionally based, pellagraform yet lipid-related and to range, once again, from heart disease to psychosis. It is an assumption that our dominant diseases are unrelated to each other or are merely revealed by our diagnostic acumen and therapeutic success; and that hydrogenating millions of tons of food oils annually, to destroy the rancidity producing w3-EFA, is safe for $\text{primates}$. Extensive beriberiform disease is reported here in 32 typical cases taken from medical practice which responds strikingly to linseed oil supplements (60% w3-EFA) in confirmation of identical results in Capuchins.     

'Very sore nights and days': the child's experience of illness in early modern England, c.1580-1720

Sick children were ubiquitous in early modern England, and yet they have received very little attention from historians. Taking the elusive perspective of the child, this article explores the physical, emotional, and spiritual experience of illness in England between approximately 1580 and 1720. What was it like being ill and suffering pain? How did the young respond emotionally to the anticipation of death? It is argued that children’s experiences were characterised by profound ambivalence: illness could be terrifying and distressing, but also a source of emotional and spiritual fulfilment and joy. This interpretation challenges the common assumption amongst medical historians that the experiences of early modern patients were utterly miserable. It also sheds light on children’s emotional feelings for their parents, a subject often overlooked in the historiography of childhood. The primary sources used in this article include diaries, autobiographies, letters, the biographies of pious children, printed possession cases, doctors’ casebooks, and theological treatises concerning the afterlife.     

Guidelines for the Validation and Use of Immunoassays for Determination of Introduced Proteins in Biotechnology Enhanced Crops and Derived Food Ingredients

Recent advancements in agricultural biotechnology have created a need for analytical techniques to determine introduced proteins in crops enhanced through modern biotechnology techniques. These proteins are expressed in plant tissues and may be present in food ingredients. Immunoassays are ideally suited for protein detection and may be used as both quantitative and threshold methods. Microplate ELISA and lateral flow devices are two of the most commonly used immunoassay formats for agricultural biotechnology applications. This paper provides general background information and a discussion of criteria for the validation and application of immunochemical methods to the analysis of proteins introduced into plants and food ingredients using biotechnology methods. It is the result of a collaborative effort of members of the Analytical Environmental Immunochemical Consortium. This collaborative effort represents the combined expertise of several organizations to reach consensus on establishing guidelines for the validation and use of immunoassays. Further, the paper offers developers and users a consistent approach to adopting the technology as well as aid in producing accurate and meaningful results.     

HLA in North Colombia Chimila Amerindians

HLA-A,-B,-C,-DRB1 and -DQB1 alleles have been studied in Chimila Amerindians from Sabana de San Angel (North Colombian Coast) by using high resolution molecular typing. A frequent extended haplotype was found:HLA-A*24:02-B*51:10-C*15:02-BRB1*04:07-DQB1*03:02 (28.7%) which has also been described in Amerinndian Mayos Mexican population (Mexico, California Gulf, Pacific Ocean). Other haplotypes had already been found in Amerindians from Mexico (Pacific and Atlantic Coast), Peru (highlands and Amazon Basin), Bolivia and North USA. A geographic pattern according to HLA allele or haplotype frequencies is lacking in Amerindians, as already known. Also, five new extended haplotypes were found in Chimila Amerindians. Their HLA-A*24:02 high frequencies characteristic is shared with aboriginal populations of Taiwan; also, HLA-C*01:02 high frequencies are found in New Zealand Maoris, New Caledonians and Kimberly Aborigines from Australia. Finally, this study may show a model of evolutionary factors acting and rising one HLA allele frequency (-A*24:02), but not in others that belong to the same or different HLA loci.     

Ultracryotomy: development and application (with examples from the yeast cytology) (author's transl)

The preparation steps usually necessary for obtaining ultrathin frozen sections of biological material (chemical prefixation, enclosing, cryoprotective treatment, freezing, sectioning, and post-staining the sections for transmission electron microscopy) are submitted to a critical analysis. The application of cryo-ultramicrotomy, in particularly for cytochemical purposes, is reviewed. Fundamental considerations of chemical prefixation and poststaining are supported by examples from yeast cytology. Furthermore, the efficiency of the cryo-ultramicrotomy (electron optical resolution of ultrastructural details) is demonstrated on yeast cells and protoplasts.     

The universal muscular chamber. A basic unit of the genitourinary, cardiovascular, gastro-intestinal, skeletal, cutaneous, respiratory and other systems, endocameral hypertension; and spasm, the key to their idiopathic diseases and arthropathies

Starting with the integument, we see many organs are contractile sacs or multiples thereof, which tubes or bags constitute the major part of the entire body. Recognition of this basic unit and its characteristics sheds new light, individually and collectively, on many disorders previously considered unrelated. Muscular tears and perforations develop in the walls of these chambers, being no way peculiar to those organs, wherein, hydrochloric acid occurs. So, it is not necessary to explain the absence of excessive acid from patients who exhibit holes in the gastric, uterine, aortic, duodenal, rectal, pulmonary, retina, and other walls. Muscle, not acid is the great common factor relating idiopathic disorders in the gastrointestinal tract to each other and to similar diseases in other systems. When the units are linked together, the lesions tend to appear as arthropathies, i.e. at the joints. Rephrasing common-place observations, frees us from conventional, conceptual cul-de-sacs. An observation is only as good as its interpretation, so all possibilities must be considered, otherwise, we will remain blinded by our misconceptions.     

Der Einfluß von Nahrungsmitteln und Rauchen auf die klinisch-chemische Diagnostik von Phäochromozytom,Neuroblastom und Karzinoid-Syndrom

Zusammenfassung Der Einfluß von verschiedenen Nahrungsmitteln auf Methoden zur Bestimmung von Adrenalin (AD), Noradrenalin (NA), Vanillinmandelsäure (VMS), Metanephrinen (MN), Homovanillinsäure (HVS) und 5-Hydroxyindolessigsäure (5-HIE) im 24 h-Harn zur Diagnose des Phäochromozytoms bzw. Karzinoid-Syndroms wurde untersucht. Die in die Untersuchung einbezogenen Nahrungsmittel waren: Tee, Kaffee, Mandeln, Ananas, Käse, Walnüsse, Vanillepudding, Bananen, Tomaten und Milchschokolade. Außerdem wurde der Einfluß des Zigarettenrauchens auf die Bestimmung von AD, NA, VMS und MN untersucht.Walnüsse führten zu einer starken Erhöhung der 5-HIE-Ausscheidung. Bananen erhöhten die Ausscheidung von AD, NA, VMS, MN und 5-HIE. Kaffee und Ananas bewirkten eine geringe Zunahme der MN-Werte. Rauchen von 20–30 Zigaretten/Tag beeinflußte keine der vier Variablen.Wenn die beschriebenen Methoden benutzt werden, sollte lediglich auf den Verzehr von Bananen und Walnüssen vor und während der Harnsammelperioden verzichtet werden, da die oberen Normgrenzen im Harn überschritten werden könnten. Ein Verzicht auf Kaffee und Ananas in normalen Mengen ist nicht erforderlich. Es besteht kein Anlaß, weiterhin die bisherigen umfangreichen Restriktionen der übrigen Nahrungsmittel beizubehalten.