Molecular detection of various rickettsiae in mites (acari: trombiculidae) in southern Jeolla Province, Korea

This study revealed the presence of various rickettsial agents in mites from wild rodents collected in Southern Jeolla Province, Korea, by nested polymerase chain reaction (PCR) and sequence analysis of a partial citrate synthase and rickettsia outer membrane protein B genes. Rickettsial agents closely related to the Rickettsia species TwKM02, R. australis, and the Rickettsia species Cf15 were successfully identified in this study, for the first time in Korea, and R. japonica, R. akari, R. conorii, R. felis, and R. typhi were also detected, as previously described. The data presented in this paper extend knowledge on the geographic distribution of SFG rickettsiae in eastern Asia and it may necessary to consider the role of mites in rickettsial transmission.     

Molecular detection of bacterial indicators in cosmetic/pharmaceuticals and raw materials

PCR assays were compared with standard microbiological methods for rapid detection of the United States Pharmacopoeia (USP) bacterial indicators in artificially contaminated samples of raw materials and cosmetic/pharmaceutical products. DNA primers containing the specific sequences of the uidA gene of the β-glucuronidase enzyme for Escherichia coli, the membrane lipoprotein gene oprL for Pseudomonas aeruginosa, and the 16S ribosomal gene for Staphylococcus aureus were used for detection in the PCR reaction. Contaminated samples were incubated for 24 h at 35°C. After incubation in broth media with and without 4% Tween 20, samples were streaked on selective growth media. After 5–6 days, all microbial indicators were morphologically and biochemically identified using standard methods while detection and identification by the PCR-based assays was completed within 27–30 h. Rapid PCR detection of E. coli, S. aureus, and P. aeruginosa will allow a faster quality evaluation and release of raw materials and cosmetic/pharmaceutical products sensitive to microbial contamination. Received 21 June 1998/ Accepted in revised form 11 January 1999     

Fluorescent metal nanoshell and CK19 detection on single cell image

In this article, we report the synthesis strategy and optical properties of a novel type of fluorescence metal nanoshell when it was used as imaging agent for fluorescence cell imaging. The metal nanoshells were made with 40 nm silica cores and 10 nm silver shells. Unlike typical fluorescence metal nanoshells which contain the organic dyes in the cores, novel metal nanoshells were composed of Cy5-labelled monoclonal anti-CK19 antibodies (mAbs) on the external surfaces of shells. Optical measurements to the single nanoparticles showed that in comparison with the metal free labelled mAbs, the mAb-Ag complexes displayed significantly enhanced emission intensity and dramatically shortened lifetime due to near-field interactions of fluorophores with metal. These metal nanoshells were found to be able to immunoreact with target cytokeratin 19 (CK19) molecules on the surfaces of LNCAP and HeLa cells. Fluorescence cell images were recorded on a time-resolved confocal microscope. The emissions from the metal nanoprobes could be clearly isolated from the cellular autofluorescence backgrounds on the cell images as either individuals or small clusters due to their stronger emission intensities and shorter lifetimes. These emission signals could also be precisely counted on single cell images. The count number may provide an approach for quantifying the target molecules in the cells.     

大鼠离体脑片癫痫放电特征及EC—海马环路的作用

目的和方法：采用４００～５００μｍ大鼠水平脑切片强直电刺激海马Ｓｃｈａｅｆｅｒ侧枝（６０Ｈｚ、２ｓ）全细胞、细胞外同步记录ＣＡ１神经元胞体电活动和相应树突区场电位,探讨其在癫痫发生中的作用。结果：①５３片脑片上记录到细胞内、外同步发生的原发性后放,持续２０ｓ以上,放电形式和持续时间常在第６个刺激串后趋于稳定。ＣＡ１神经元的原发性后放常跟在强直电刺激引起的阵发性去极化或超极化偏移之后（ＰＤＳ、ＰＨＳ）。它可以从紧张性放电向爆发性放电转化,振幅逐渐递增并与细胞外癫痫样放电同步,产生癫痫放电极性偏移;②其中８／４０脑片细胞外可记录到继发性后放之后出现的自发性发作样癫痫放电,长达数分钟,与全细胞记录的ＥＰＳＰ同步。切断ＥＣ与海马之间的联系可以易化海马癫痫电活动（３／５）。结论：ＥＣ输入到海马的神经通路可能在封闭的ＥＣ海马环路中起着重要的门控作用     

The diagnostic value of combined detection of genetic markers and serum protein markers on breast cancer

Objective: The research is to explore the diagnostic value of several detection methods including separated and combined detection of the related genes and related proteins of breast cancer and combined detection of all genetic markers and serum protein markers on breast cancer. Method: The mRNA level expression of the related genes of breast cancer was detected by FQ-PCR technique and the ratio of BRCA-1, Myc, C-erbB2 and β2 micro-globulin was used to express levels of BRCA-1, Myc and C-erbB2; the related proteins of breast cancer were detected through ELISA. Then the research data was analyzed by SPSS19.0 software with t-test as comparison method, and ROC curve was used to calculate the sensitivity, specificity and accuracy of the diagnostic models. Result: No difference can be found among the six indexes in the control group and benign breast tumor group while compared with the benign breast tumor group and the control group, the breast cancer group was significantly different from them; combined detection of genes and that of proteins were both superior to their separated detection; all-marker combined detection was superior to separated detection, which is consistent with combined detection of genes and proteins. Conclusion: More detection indexes will not necessarily outcome better detection effect. Hence, appropriate detection indexes and number are needed to achieve better diagnosis effect. In order to conduct more specific method, more test samples are needed for further researches.     

B7-2胚胎性癌细胞诱导分化过程中的表型转变

胚胎性癌细胞(简称EC细胞)作为一类肿瘤(畸胎瘤)的干细胞近年受到广泛的重视,从胚胎学、肿瘤学和分子生物学等许多学科领域都应用它作为实验材料,离体诱导分化研究是其中的一个方面。B 7-2 EC细胞是我们从129品系小鼠的自发睾丸畸胎瘤中分离克隆得到的一株多能EC细胞,它在同种同基因小鼠     

磷脂酸含量测定法评价烟曲霉孢子对肺上皮细胞磷脂酶D活性的影响

目的 优化检测烟曲霉刺激A549细胞后磷脂酸（phosphotidic acid,PA）含量变化的方法,间接反应细胞内磷脂酶D（Phospholipase D,PLD）活性变化.方法 建立烟曲霉ATCC13073刺激肺上皮细胞模型;采用甲醇氯仿法提取胞内脂质;用改良的磷脂酸含量测定法测定PA标准品和细胞内PA水平变化规律.结果 PA标准品在5～ 250 μmol/L呈线性关系;经膨胀孢子刺激后,肺上皮细胞内PA含量显著升高,休眠孢子在这一过程中对肺上皮细胞内PA含量无明显作用.结论 改良的PA测量法能快速、稳定而有效地测定细胞内的PLD活性.烟曲霉膨胀孢子能显著激活肺上皮细胞内的PLD活性.     

Rapid detection of Phytophthora nicotianae by simple DNA extraction and real‐time loop‐mediated isothermal amplification assay

Phytophthora nicotianae is a phytopathogenic oomycete with a wide host range and worldwide distribution. Rapid detection and diagnosis at the early stages of disease development are important for the effective control of P. nicotianae. In this study, we designed a simple and rapid loop‐mediated isothermal amplification (LAMP)‐based detection method for P. nicotianae. We tested three DNA extraction methods and selected the Kaneka Easy DNA Extraction Kit version 2, which is rapid and robust for LAMP‐based detection. The designed primers were tested using mycelial DNA from 35 species (81 isolates) of Phytophthora, 12 species (12 isolates) of Pythium, one isolate of Phytopythium and one isolate each from seven other soil‐borne pathogens. All of the 42 P. nicotianae isolates were detected by these primers, and no other isolates gave positive results. Three isolates were tested for the sensitivity of the reaction, and the lowest amounts of template DNA that could be detected were 10 fg for two isolates and 1 fg for the third. The target was detected within 25 min in all tested samples, including DNA extracted from both inoculated and naturally infected plants. In contrast, PCR assays with P. nicotianae‐specific primers failed or showed weakened detection in several samples. Thus, we found that the rapid DNA extraction and LAMP assay methods developed in this study can be used to detect P. nicotianae with high sensitivity, specificity and stability.