DEVELOPMENT AND EVALUATION OF A RAPID ENZYME-LINKED IMMUNOFILTRATION ASSAY (ELIFA) AND AN ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA) FOR THE DETECTION OF STAPHYLOCOCCAL ENTEROTOXIN B

An enzyme-linked immunofiltration assay (ELIFA) and a microtitre plate enzyme-linked immunosorbent assay (ELISA) were developed and compared for their ability to detect staphylococcal enterotoxin B (SEB). The double antibody capture format was used for both assays. Factors which improved the sensitivity of the ELIFA system were (1) addition of casein and thimerosal to the antigen dilution buffer; (2) addition of polyethylene glycol (MW 6000) to the detection and conjugate antibody dilution buffers; and (3) washing with diethanolamine buffer prior to addition of the substrate/chromogen. The ELIFA system had a turnaround time of approximately 1 h and a detection limit of 1 ng/mL of purified SEB. The ELISA had a total turnaround time of 21 h, or 3 h using plates pre-coated overnight with the capture antibody. The detection limit of the ELISA for purified SEB was 0.05 ng/mL. The detection limit of SEB in cheese samples spiked with purified enterotoxin and subjected to a simple extraction procedure was 1 ng/mL and 0.1 ng/mL of extract, with the ELIFA and the ELISA, respectively.     

脱落酸的酶标免疫测定

使用国产辣根过氧化物酶(HRP)合成酶标记物,用竞争法进行了植物内源激素脱落酸(ABA)的酶标免疫测定研究。测定范围为0.0125ng—100ng,在此范围内logitB/B。与ABA浓度的对数之间呈较好的线性关系。检测的灵敏度达到5×10~(-14)克分子。比较了两种酶标记方法对于测定的影响,结果发现直接使ABA共价结合到HRP上形成ABA-HRP酶标记物比先将ABA与牛血清白蛋白(BSA)结合后,然后进一步再与HRP反应形成ABA—BSA—HRP复合物灵敏度高,非特异性吸附小,而且合成步骤较少。酶标记物的稀释度直接影响测定的灵敏度和浓度对数与logit B/B。之间的线性关系好坏;在以每毫升3—6微克免疫球蛋白包埋免疫吸附板进行测定时,用每毫升5微克酶标记物的浓度获得了最佳结果。     

抗人IgA多克隆抗体ELISA法的建立和应用

用马抗人IgA抗体ELISA法,结合血清预处理,检测鼻咽癌(NPC)患者,其他肿瘤病人和正常人血清中的IgA/EA抗体结果表明,95.1％NPC病人阳性(抗体几何平均滴度为294.1),而健康对照组仅2.4～4.5％,ELISA与免疫酶染色(IE)法测得的血清抗体滴度之间有一定的相关性。但前者比后者测得的几何平均滴度(GMT)高9倍,本法敏感、特异、快速、简便,适用于现场大规模普查。     

PLASMA PROGESTERONE CONCENTRATIONS MEASURED USING AN ENZYME-LINKED IMMUNOSORBENT ASSAY USEFUL FOR DIAGNOSING PREGNANCY IN HARBOR SEALS (PHOCA VITULINA)

Wild-caught female harbor seals ( Phoca vitulina ) were classified as sexually mature or immature on the basis of standard body length (< 125 cm immature, > 125 cm mature) and plasma progesterone concentrations measured using an enzyme-linked immunosorbent assay (ELISA), a technique usable in the field. Sexually mature females were classified as pregnant or non-pregnant on the basis of their plasma progesterone concentrations. Of 28 wild mature female harbor seals caught in the Moray Firth, N.E. Scotland, between the end of February and the end of May, 79% had plasma progesterone concentrations greater than 60 nmol liter⁻¹, the lowest plasma progesterone concentration measured in one of eight females later observed with a pup, and were diagnosed as pregnant. A linear discriminant function, calculated to provide a method of distinguishing pregnant and non-pregnant females, predicted 100% of non-pregnant females and 95.8% of pregnant females using plasma progesterone concentration, standard length, and month of capture as parameters. Plasma progesterone concentrations were less than 30 nmol liter⁻¹ in all mature and immature males and immature females. In mature females plasma progesterone concentrations ranged from 0-318 nmol liter⁻¹.     

草鱼生长激素非竞争式酶联免疫吸附测定法的建立及鉴定

应用草鱼生长激素（ｇｃＧＨ）单克隆抗体及多价兔抗血清建立了草鱼ＧＨ非竞争式酶’联免疫吸附测定ＥＬＩＳＡ系统。用正辛酸法对腹水单抗进行了分离纯化,获得了高纯度的单抗制备物。聚丙烯酸胺凝胶电泳表明纯化的单抗由分子量分别为５５ｋＤ和２５ｋＤ的两条蛋白带组成。用纯化单抗铺底,用兔抗血清作后续抗体建立了一种测定草鱼ＧＨ的非竞争式双抗夹心ＥＬＩＳＡ方法。交叉试验表明该测定系统只与草鱼ＧＨ和基因重组鲤生长激素（ｒｃＧＨ）具有剂量依存的结合反应,而与大马哈鱼生长激素（ｓＧＨ）、牛生长激素（ｂＧＨ）、大马哈鱼促性腺激素（ｓＧｔＨ）、及黑鲢促性腺激素（ｂｓｃＧｔＨ）等均无交叉反应。该 ＥＬＩＳＡ方法的灵敏度可达０．８ｎｇ／ｍｌ,组内变异系数为 ５．９ ％,组间变异系数为７．６％,回收率达９０％以上。初步应用表明,鲤和团头鲂垂体抽提液、草鱼血清、鲤血清及鲫血清在该测定系统中有剂量依存的反应曲线,而大口鲶、黄颡鱼、中华鲟及黄鳝鱼垂体抽提液及大口鲶、胡子鲶和罗非鱼血清在该测定系统中没有交叉反应。     

AN INDIRECT METHOD TO ASSAY FOR MITOTIC CENTERS IN SAND DOLLAR (DENDRASTER EXCENTRICUS) EGGS

It is possible consistently to induce sea urchin and sand dollar eggs to cleave directly from one cell into four cells. This is done by exposing the fertilized eggs to benzimidazole for 20 to 30 min beginning about early metaphase. The mitotic apparatus regresses, the cells do not cleave, and shortly after they are returned to normal sea water an early-prophase-appearing nucleus is present in each cell. Each cell then organizes a tetrapolar tetrahedral mitotic apparatus de novo, instead of transforming a bipolar mitotic apparatus into a tetrapolar figure, and cleaves one-to-four. In another type of experiment, it appears that sand dollar eggs exposed to mercaptoethanol during the first period of mitotic center duplication have only half as many centers by first cleavage metaphase as the normal controls. This is consistent with an earlier report by Mazia et al (1960). Using this same experimental technique, it was demonstrated that benzimidazole, on the contrary, does not interfere with mitotic center duplication in sand dollar eggs. A labeling experiment demonstrated that benzimidazole does not interfere markedly with the normal pattern of incorporation of C¹⁴-thymidine into the DNA of sea urchin eggs. The data reported here suggest that judicious treatment of sand dollar eggs (and probably sea urchin eggs, too) with benzimidazole can induce the eggs to cleave into as many cells as there were mitotic centers sometime earlier, for example at early metaphase of the first cleavage division. This provides a very useful tool for studies on the process of mitotic center duplication.     

AN ELECTRICAL TESTING METHOD FOR THE DETECTION OF LISTERIA SPECIES IN CONFECTIONERY PRODUCTS

An electrical testing method for the detection of Listeria spp. in confectionery products and associated raw materials was developed in which samples can be negatively screened within 48h. The method involves a 24h resuscitation in Listeria enrichment broth followed by a 24h test in a Bactometer M128 using a broth (Claremont broth) developed from Oxford agar. The comparative study involved analysis of 511 samples (chocolate, dairy, cereal, nut and fruit products) tested by the Interim Australian Standard method (AS) and the Bactometer method (BM). The sensitivity and specificity of the BM for samples §1 Listeria/g was 100% and 99.8%, respectively, when compared to the AS method. When samples containing < 1 Listeria/g sample were added to the analysis, the sensitivity and specificity marginally dropped (98.3% and 99.6%). The electrical capacitance method is rapid and easy to perform, with a negative result being available within 48h making it a viable tool in a positive release QA program in food manufacturing factories.     

一次PCR扩增单纯疱疹和巨细胞病毒基因的检测方法

宫内感染是影响优生优育质量的一个重要因素,由病毒引起的宫内感染主要有以下四类病原：ＨＳＶ、ＣＭＶ、ＴＯＸＯ和风疹病毒（ＲＶ）,其中ＨＳＶ和ＣＭＶ的感染率较高。孕期这两种病原的感染会影响胎儿的正常发育,严重的可导致流产甚至胎儿畸形。目前已有针对这两种病...     

AN OPTICAL DETECTION SYSTEM FOR THE STUDY OF FINE SCALE VERTICAL DISPLACEMENT OF MICROALGAE IN AN ARTIFICIAL WATER COLUMN

A proper knowledge of the vertical organization of the phytoplankton is of fundamental importance for our understanding of the functions of pelagic ecosystems. Essential in this context is the existence of vertical gradients in environmental parameters. However, little is known about how the fine vertical structures of phytoplankton species are formed and maintained. In situ study of phytoplankton is biased by the fact that submersing instruments can disturb or even destroy the fine vertical gradients in species composition and/or cell numbers. We have designed and constructed an optical instrumentation system by which fine-scale vertical displacements of microalgae can be studied in an artificial water column without influencing fine physical, chemical, and biological structures of the water column. This enables us to find out more about the fine-scale behavioral responses of microalgae to vertical gradients in environmental parameters. We describe the main system, present some test results, and conclude that our optical system is able to reveal fine-scale vertical displacements of microalgae in an artificial water column and that the system can detect differences in cell densities down to 100 cells·mL⁻¹.     

RAPID DETECTION OF MICROBIAL CONTAMINATION IN TRICLOSAN AND HIGH FLUORIDE DENTIFRICES USING AN ATP BIOLUMINESCENCE ASSAY

Abstract The Celsis ATP Bioluminescence method was optimized and validated to detect the presence of microbial contamination in High Fluoride and Triclosan dentifrice formulations. Several enrichment broths were evaluated by using a 24–27 h incubation period. The ATP concentrations of the enrichment broths were found to a range from 0.012 to 0.040 nM. None of the tested enrichment broths were found to exhibit any sample inhibition/enhancement effects on the ATP Bioluminescence reaction. Dentifrice suspensions were inoculated with bacteria, yeast, and mold. All test microorganisms (ca. 1–15 CFU/g) were detected within a 24–27 h incubation period by using TAT Broth Base enrichment broths containing different concentrations of the following ingredients: Tween 20, Neopeptone, Dextrose, Triton X-100, Thiosulfate, Sodium Dibasic Phosphate, and Glycine. Negative ATP response after 24–27 h of incubation at 35C indicates the absence of contamination from these products.     

鼻咽癌患者血清中抗Epstein-Barr病毒早期和晚期膜抗原抗体的检测

对Epstein-Barr(EB)病毒抗原的研究,发现有淋巴细胞确定的膜抗原(Lydma)、早期抗原(EA)、壳抗原(VCA)、核抗原(EBNA)、早期膜抗原(EMA)和晚期膜抗原(LMA)。除了Lydma抗原外,鼻咽癌患者对上述抗原均产生相应的IgG和IgA抗体。因而研究这些抗体,对阐明EB病毒与鼻咽癌的关系及鼻咽癌的早期诊断都十分有价值。     

糖尿病患者人巨细胞病毒感染的抗体检测分析

目的 :了解人巨细胞病毒 (HCMV)感染与糖尿病的关系。方法 :运用捕获 EL ISA、间接 EL ISA对 516例糖尿病患者及 14 8例健康正常人 HCMV-Ig M、HCMV-Ig G进行检测 ,并对比分析 HCMV感染状态 ,根据糖尿病患者的初发年龄和病程的不同进行了分组探讨。结果 :糖尿病患者 HCMV-Ig M、HCMV-Ig G抗体阳性率明显高于对照组 (P<0 .0 1) ,始发病年龄越小及病程越短的糖尿病病人 HCMV-Ig M抗体阳性率明显升高 ;年龄较大及病程较长的糖尿病人 HCMV-Ig G检出率较高。结论 :年龄小或病程短的糖尿病人以原发性 HCMV感染为主 ,HCMV感染在糖尿病的发病机制中起一定的作用