健康育成黑熊血液生理生化指标的测定

测定了30头(雌雄各半)健康育成黑熊的血液生理生化指标,结果显示:(1)雌性黑熊和雄性黑熊的淋巴细胞数、红细胞平均体积、平均血红蛋白含量存在极显著差异(P＜0.01);雌性黑熊的淋巴细胞数为2.99×109个/L、红细胞平均体积为62.27fL、平均血红蛋白量为23.29 pg,雄性黑熊的为1.84×109个/L、65.51 fL、24.34 pg,其余10项血液生理指标无显著差异.(2)雌性黑熊和雄性黑熊的球蛋白、白蛋白与球蛋白比值存在极显著差异(P＜0.01),雌性黑熊的球蛋白为32.91 g/L、白蛋白与球蛋白比值为1.32,雄性黑熊的为37.07 g/L、1.15,雌性黑熊和雄性黑熊的总蛋白、碱性磷酸酶存在显著差异(P＜0.05),雌性黑熊的总蛋白为75.99 g/L、碱性磷酸酶为106.86 U/L,雄性黑熊的为79.27g/L、54.33 U/L,其余9项血液生化指标无显著差异.这些结果为黑熊的保护与疾病防治积累了资料.     

黄鳝两种抗病毒基因的克隆与组织表达分析

[目的]干扰素调节因子(IRFs)-5,IRF-7在抗病毒免疫反应中发挥重要作用,研究黄鳝(Monopterus albus)干扰素调节因子的结构及表达有助于阐明黄鳝抗病毒的机理。[方法]通过PCR法扩增黄鳝IRF-5和IRF-7两个基因c DNA序列,分析其氨基酸序列,并利用荧光定量PCR技术研究黄鳝不同组织这两个基因的表达。[结果]已扩增到黄鳝IRF5 c DNA序列887bp,编码295aa;IRF7 c DNA序列1 181bp,编码378aa。荧光定量显示:IRF-5在脑中表达量很高,IRF5/β-actin高达120.3×10-2,而在主要免疫器官头肾、尾肾、肠和脾脏中这个比值仅分别为:0.17×10-2、0.11×10-2、0.69×10-2和0.60×10-2。IRF-7在皮肤中表达量很高,IRF7/β-actin比值达68.3×10-2。[结论]IRF-5可能在黄鳝神经系统中发挥重要作用,而IRF-7在黄鳝粘膜免疫中发挥重要作用。     

用Percoll试剂和DEAE-纤维素层析柱分离纯化鳝锥虫

用51%的Percoll分离液和DEAE-纤维素层析柱,从自然感染的黄鳝(Monopterus albusZuiew)中分离鳝锥虫(Trypanosoma monopteriChen et Hsieh,1964)。对含虫血液样品分别以2.1×103r/min、2.5×103r/min、2.9×103r/min、3.3×103r/min、3.6×103r/min离心5、10、15、20min,观察红细胞的去除和鳝锥虫存留百分数,结果表明:2.9×103r/min离心15min分离效果最佳。离心分离的混悬液含有锥虫、白细胞、血栓细胞和极少的红细胞,悬混液再经DEAE-纤维素过柱,得到纯净的鳝锥虫。过柱后光学显微镜观察,锥虫形态正常,运动活泼,无血细胞。本方法具有耗时短、回收率高的特点,是把梯度分离和离子交换结合在一起的一种快速有效方法,为鳝锥虫的进一步研究奠定了基础。另外,此方法还可以作为快速、准确检测自然状态下黄鳝是否感染锥虫的手段,为鳝锥虫的流行病学研究提供可靠的数据。     

全反式维甲酸抑制猪脂肪间充质干细胞的成脂分化

分离培养猪脂肪间充质干细胞（adipose mesenchymal stem cells, AMSCs）,流式细胞仪鉴定其表面标记.利用MTT比色检测不同浓度的全反式维甲酸（all trans retinoic acid, ATRA）对猪AMSCs增殖的影响;光学显微镜下观察猪AMSCs向脂肪细胞分化的形态学变化;油红O染色提取法分析不同浓度ATRA对猪AMSCs成脂分化的影响;RT PCR检测脂肪细胞分化标志基因LPL和aP2 mRNA的变化.MTT比色结果显示,生理浓度（1×10^－9～1×10^－8 mol/L）和药理浓度（1×10^－7～1×10^－5 mol/L）ATRA对猪AMSCs增殖均没有影响.油红O染色提取法结果表明,除1×10^－7　mol/L ATRA对猪AMSCs成脂分化没有影响外,生理浓度（1×10^－9～1×10^－8 mol/L）和其它药理浓度（1×10^－6～1×10^－5 mol/L）ATRA均显著抑制猪AMSCs成脂分化（P<0.05）.RT-PCR检测显示,ATRA显著抑制脂肪细胞分化标志基因LPL和aP2 mRNA表达（P<0.05）.     

Sysmex XN-1000血液分析仪检测血小板3种方法的比较

目的:评价Sysmex XN-1000血液分析仪检测血小板(PLT)3种方法的准确性。方法:收集广州总医院2017年1、2月住院病人血液标本共166例,分别用手工法和XN-1000血液分析仪的电阻抗法(PLT-I)、光学法(PLT-O)、荧光染色法(PLT-F)检测同一标本的血小板计数,以手工法为参考标准,比较XN-1000血液分析仪3种方法的准确性。结果:当PLT计数范围为100×10~9~300×10~9/L和300×10~9/L时,3种仪器法与手工法结果比较,差异无统计学意义(P0.05);当PLT100×10~9/L时,仪器法中的PLT-O和PLT-F法与手工法结果比较,差异无统计学意义(P0.05),而PLT-I法与手工法结果比较,差异有统计学意义(P0.05),其与手工法的相关性比较依次为PLT-FPLT-OPLT-I。结论:当PLT100×10~9/L时,仪器检测血小板计数3种方法的准确性均较好;当PLT100×10~9/L时,PLT-F和PLT-O法的检测准确性优于PLT-I法。     

龙洞山溪鲵的血细胞组成及血红蛋白含量检测

血液学指标不仅可以反映动物生理和病理变化,也能体现动物对环境的适应。本文利用血液学方法检测了27尾龙洞山溪鲵(Batrachuperus londongensis)个体的血细胞组成和血红蛋白含量,龙洞山溪鲵血红蛋白平均含量为4.16×10~(-2) g/ml;红细胞卵圆形,细胞核椭圆形位于中央或亚中央,红细胞平均含量为6.04×10~4个/mm3;白细胞数量较少,多为圆形或近圆形,平均含量为2.90×10~3个/mm3;白细胞中淋巴细胞最多,其次为单核细胞、嗜碱性细胞和中性细胞,嗜酸性细胞最少。血液参数中仅中性细胞和嗜碱性细胞的百分比在雌雄之间存在显著差异;头体长、体重和各血液参数的相关性均不显著。同时,将研究结果与其他小鲵科动物的数据进行了比较。本文报道的龙洞山溪鲵血细胞组成和血红蛋白含量的基础数据为其后续的健康监测和诊断以及适应进化的研究提供了基础资料。     

棕色田鼠血液生理生化指标的测定

报道了棕色田鼠指名亚种的正常血象值及几项血液生化值。结果表明,各项备注学和血液生化学参数值在成年个体间无性别差异（附红细胞数、血红蛋白浓度、红细胞压积外,但此三项指标尚未达到极显著差异水平）。与其它仓鼠科动物的血液生理生化参数值相比较,棕色田鼠除红细胞数、血红蛋白浓度及中性粒细胞略低外,其它各项参数虽有差异,但均无统计学意义。     

黄鳝的繁殖生态学研究

以洞庭湖稻区黄鳝为调查研究对象 ,通过野外调查和室内实验系统研究了黄鳝的繁殖生态。对黄鳝的性腺发育周年变化、繁殖行为、自然产卵过程、产卵条件及繁殖洞穴的构造进行了观察和记录 ,产卵室是黄鳝繁殖洞穴特有的构造 ,繁殖洞穴泥土p H平均值为 6 .73± 1.0 12。亲鳝有护卵习性 ,通过对 30尾守洞亲鳝的性腺观察和切片验证 ,守洞鳝多为雄鳝 (占 6 1.3% ) ,少数为兼性偏雄性 (占 38.7% ) ,守洞鳝 1:0 0～ 2 :0 0全在外面活动 ,守洞鳝体长 L (cm)与体重 W (g)回归方程为 :W=1.2 5×10 - 2 L1 .4 2 (r=0 .76 )。人工模拟条件下 ,泡沫组受精卵的平均孵化率 (85 .2 % )极显著地高于对照组 (2 5 .4 % ) (t=8.18,t0 .0 5=2 .4 5 ) ,仔鱼平均成活率 (6 4 .0 % )显著高于对照组 (14 .0 % ) (t=3.73) ,同时对繁殖季节黄鳝为孵卵而所吐的泡沫的作用进行了分析。另外 ,证实了黄鳝产卵的最适放养密度为 2～ 3尾 / m2。为黄鳝的全人工繁殖和半人工繁殖提供了合理化的建议     

海南鳽部分血液生理生化指标

应用血球分析仪和全自动生化分析仪测定了海南鳽的7项血液生理指标和33项血清生化指标.结果显示:海南鳽像兀鹫一样具有大型的红细胞,与已报道的白鹭、池鹭、夜鹭、彩鹳、石鸡等几种鸟类相比,其平均红细胞体积(MCV)和平均红细胞血红蛋白含量(MCH)较高,红细胞计数(RBC)、红细胞压积(HCT)和血红蛋白浓度(HGB)均较低;尿素(Urea)含量较高.有关红细胞的生理指标揭示了海南鳽的血液携氧能力较低.生化指标与其他鸟类的差异可能是由物种不同所致.     

黄鳝ISSR-PCR反应体系的建立及条件优化

以黄鳝基因组DNA为模板,采用正交试验设计方法,对各反应因子、引物退火温度和循环参数进行优化.建立了黄鳝的最适ISSR-PCR反应体系,25 μl反应体系中含2.5 mmol/ L Mg2+,250 μmol/ L dNTPs,0.25 μmol/ L 引物,1.0 U Taq DNA聚合酶,30 ng DNA模板.最佳反应程序:94℃预变性5 min;94℃变性40 s,48～57℃复性40 s(随引物而确定),72℃延伸1.5 min,循环次数40;72℃延伸10 min.利用所建立的ISSR反应体系,获得了清晰、重复性好、多态性高的DNA谱带.     

Production of the eel grass <Emphasis Type="Italic">Zostera marina</Emphasis> Linnaeus, 1753 in the White Sea

The results of year-round studies of the photosynthesis and respiration rates of the eel grass Zostera marina L. in the White Sea are presented. The annual production of eel grass is estimated to be 9.86 × 10¹¹ kcal. It is concluded that the total annual production of the eel grass constitutes about 3% of the annual phytoplankton production (3 × 10¹³ kcal; according to the data of Bobrov et al., 1995) in the White Sea.     

ESTIMATION OF EFFECTIVE EOSINOPOIESIS AND BONE MARROW EOSINOPHIL RESERVE CAPACITY IN NORMAL MAN

The eosinophil reserve capacity of the post-mitotic granulocyte compartment in the bone marrow and the effective eosinopoiesis in three haematologically normal men have been quantified by means of kinetic parameters of [³H]thymidine flash-labelled peripheral blood eosinophils. From (a) the time of the appearance in the blood of labelled eosinophils after the tracer injection, (b) the inflow characteristics of the labelled eosinophils in the blood and (c) the magnitude of the eosinophil granulocyte pool in the venous blood, the effective eosinopoiesis (i.e. the eosinophil turnover) was calculated to range between 0.014 and 0.031 × 10⁹ cells/kg body weight per day (mean 0.22 × 10⁹ cell/kg per day). The post-mitotic eosinophil reserve capacity of the bone marrow ranged from 0.09 to 0.20 × 10⁹ cells/kg body weight (mean 0.14 × 10⁹ cells/kg). The large reserve pool and the high turnover rate may contribute to sudden rises of the peripheral blood oesinophil counts in some cases of eosinophilia.     

雌激素对A549细胞系EMT过程的影响

目的:探讨雌激素对A549细胞系EMT标志物表达量的影响。方法:用不同浓度的雌激素刺激A549细胞系,并用q-RT-PCR和Western-blot实验检测各组细胞中EMT标志物表达量的变化,用Transwell实验检测不同浓度雌激素对细胞迁移能力的影响,计算各组间有无统计学差异。结果:当雌激素浓度为1×10-9 mol/L、1×10-8 mol/L、1×10-7 mol/L时,Vimentin的m RNA表达量分别为:2.14±0.55、4.72±0.63、2.21±0.47,显著高于空白对照组,组间有统计学差异,E-cadherin的m RNA表达量分别为:0.64±0.15、0.46±0.11、0.59±0.13,显著低于空白对照组,组间有统计学差异,蛋白表达量也有同样趋势;细胞迁移数分别为58.63±7.33、80.12±9.32、61.89±8.22,组间有统计学差异。当雌激素浓度为1×10-8 mol/L时,Vimentin的表达量最高,E-cadherin的表达量最低,细胞迁移数最高。结论:适宜浓度雌激素可以促进Vimentin的表达,抑制E-cadherin的表达,提高细胞迁移能力,当雌激素浓度为1×10-8 mol/L时,促进Vimentin表达、抑制E-cadherin表达和促进细胞迁移的作用最显著。由此认为,雌激素对A549细胞系发生EMT过程有促进作用。     

Concentration of human thymidine kinase 1 discover invisible malignant human tumours

Early discover of risk progression of invisible carcinomas is important for a prerequisite successful treatment. Here, we investigated whether concentration of human thymidine kinase 1 (HTK1) discover invisible malignant human tumours. The HTK1 concentration of tumour cellular based on HTK1 IgY-polyclonal-antibody (HTK1-IgY-pAb) was determined by using a novel automatic chemiluminescence analyser with sandwich biotin-streptavidin (SBSA) platform. Minimum number of cells able to be detected by this technology used cells with low and high concentration of HTK1. The limit visibility by tumour imaging is approximately 1 mm in diameter, corresponding to approximately 10⁹ cells with a cell diameter of 1 µm. Based on a HTK1 standard curve and a molecular weight of HTK1 of 96 kD, the HTK1protein (HTK1p) concentration per cell was calculated to be 0.021 pg. Assuming 200 pg in total protein/cell, approximately 50 × 10⁶ growing malignant cells in the body were calculated to releases HTK1 into 5-liter blood. A HTK1 values of 3.914, 0.435 and 0.009 pmol/L corresponds to 10 × 10⁵, 2 × 10⁵ and 1 × 10⁵ growing malignant cells, respectively. The lowest detectable sensitivity of HTK1 is 0.009 pmol/L in 1 × 10⁵ growing malignant cells and 0.01 pmol/L in blood serum, detectable in health screening. Comparing the novel automatic chemiluminescence analyser with the original ECL dot-blot assay using serum HTK1p (health screening, n = 265) showed high correlation (r = 0.8743, P < .000). In conclusion, the novel automatic chemiluminescence analyser with SBSA platform is a reliable method with high accuracy to determine carcinoma invisible.     

Proteome analysis of circulating exosomes in health and breast cancer

Microvesicles were isolated from blood plasma and total blood of healthy females and breast cancer patients by filtration and ultracentrifugation. According to flow cytometry, different subpopulations of exosomes were represented in blood of healthy donors and cancer patients at different levels with median fluorescence intensity (MFI) values in both groups arranged in the following order: CD24/СD9 > СD9/СD81 > CD9/CD63 = CD24/CD63. Concentration of exosomes in blood plasma of healthy females estimated by nanoparticle tracking analysis (NTA) did not exceed (3.71 ± 1.15) × 10⁷ particles/mL of blood and did not differ from that in plasma of breast cancer patients, which averaged (3.99 ± 1.03) × 10⁷ particles/mL of blood. Concentration of total exosomes in blood (including exosomes from plasma and blood cell surface-bound exosomes) did not depend on the presence/absence of a tumor; the values were (7.66 ± 0.7) × 10⁷ particles/mL of healthy blood and (9.4 ± 1.24) × 10⁷ particles/mL of blood from cancer patients. Comparative analysis of exosomes using 2-D electrophoresis with subsequent analysis of 2-D proteomic maps revealed proteins missing in blood or differentially expressed in healthy females and breast cancer women. The data presented provide the possibility for identification of exosomal proteomic markers and isolation of tumor-specific exosomes, which contributes to the development of breast cancer diagnostics.     

A new spectrofluorimetric method for the determination of some tetracyclines based on their interfering effect on resonance fluorescence energy transfer

A rapid, simple, inexpensive and highly sensitive spectrofluorimetric method was developed for the determination of trace amounts of some tetracyclines (TCs), namely tetracycline hydrochloride (TCH), oxytetracycline hydrochloride (OTCH) and minocycline hydrochloride (MCH). Binding rhodamine B (RhB) to gold nanoparticles (Au NPs) resulted in quenching of the fluorescence of RhB by a resonance energy transfer (FRET) mechanism, with Au NPs as the energy acceptors. The presence of TCs caused the release of RhB molecules and recovered their fluorescence, and this was used as a basis for the quantitative determination of TCs. The reaction was monitored spectrofluorimetrically by measuring the increase in fluorescence of RhB at 572 nm starting 5 min after mixing the reagents in Tris buffer solution (pH 6.5). The effect of various experimental factors such as buffer type, pH, concentrations of the involved reagents and reaction time were studied to optimize the reaction conditions. Under optimum conditions, the calibration graphs were linear within the ranges 2.08 × 10^?9–1.04 × 10^?6 mol/L, 2.01 × 10^?9–1.00 × 10^?6 mol/L and 2.02 × 10^?9–1.01 × 10^?6 mol/L and detection limits (LODs) of 0.61 × 10^?9, 0.32 × 10^?9 and 0.66 × 10^?9 mol/L were calculated for TCH, OTCH and MCH, respectively, with corresponding percent relative standard deviations (%RSDs) of 1.18, 1.21 and 1.54 (n = 5). The method was successfully applied to the determination of TCs in drinking water, human urine, bovine milk and breast milk samples. Copyright © 2014 John Wiley & Sons, Ltd.     

利用GFP表达系统检测雌激素类化合物的研究

根据雌激素类化合物的转录调节原理,构建受雌激素应答元件（ERE）调控的3×ERE EGFP-N1重组报告基因载体,转染雌激素受体阳性的MCF-7乳腺癌细胞,分离出稳定转染的细胞克隆ERE-GFP-MCF7,将不同浓度的雌二醇（E2）及其他化合物加入稳定转染的细胞后检测GFP表达水平的变化。雌二醇（E2）以剂量依赖的方式诱导稳定转染细胞ERE-GFP-MCF7表达GFP,最大效应浓度为1×10－10mol/L ,EC50为1.5×10－11 mol/L;已知的植物雌激素大豆甙元和白藜芦醇同样以剂量依赖的方式诱导GFP的表达,EC50分别为2.4×10－7 mol/L和6.2×10－6 mol/L,而葡萄籽多酚没有明显的雌激素样活性。雌激素拮抗剂他莫西芬以剂量依赖的方式抑制雌二醇诱导GFP的表达,最大抑制浓度为1×10－7 mol/L。利用此细胞模型检测不同化合物诱导的GFP表达强度可快速检测雌激素受体的配体。     

Capillary electrophoresis coupled with end‐column electrochemiluminescence for the determination of ephedrine in human urine,and a study of its interactions with three proteins

A tris(2,2‐bipyridyl)ruthenium(II) (Ru(bpy)₃²⁺)‐based electrochemiluminescence (ECL) detection coupled with capillary electrophoresis (CE) method has been established for the sensitive determination of ephedrine for the first time. Under the optimized conditions [ECL detection at 1.15 V, 25 mmol/L phosphate buffer solution (PBS), pH 8.0, as running buffer, separation voltage 12.5 kV, 5 mmol/L Ru(bpy)₃²⁺ with 60 mmol/L PBS, pH 8.5, in the detection cell] linear correlation (r = 0.9987) between ECL intensity and ephedrine concentration was obtained in the range 6.0 × 10^–8–6.0 × 10^–6 g/mL. The detection limit was 4.5 × 10^–9 g/mL (S:N = 3). The developed method was successfully applied to the analysis of ephedrine in human urine and the investigation of its interactions with three proteins, including bovine serum albumin (BSA), cytochrome C (Cyt‐C) and myoglobin (Mb). The number of binding sites and the binding constants between ephedrine and BSA, Cyt‐C and Mb were 8.52, 12.60, 10.66 and 1.55 × 10⁴ mol/L, 6.58 × 10³ mol/L and 1.59 × 10⁴ mol/L, respectively. Copyright © 2011 John Wiley & Sons, Ltd.     

Flow‐injection chemiluminescence determination of haemoglobin in the blood

In this study, a sensitive and simple flow‐injection chemiluminescence (CL) method was developed for the quantitative analysis of haemoglobin. The method is based on the ability of haemoglobin to enhance the CL signal generated by a H₂O₂–K₄Fe(CN)₆–fluorescein alkaline system enhanced by CdTe quantum dots. Under the optimized conditions, haemoglobin can be detected in concentration range 7.35 × 10^–9–2.5 × 10^–6 mol/L, with a detection limit (3σ) of 1.8 × 10^–9 mol/L and a relative standard deviation (RSD; for 5 × 10^–7 mol/L haemoglobin) of 2.06% (n = 11). The present CL method was successfully applied for the determination of haemoglobin in three kinds of blood samples taken from an infant, an adult man, an adult woman and two reference samples. Compared with previous reports, the CL method described in this work is simple and rapid, with high sensitivity. Copyright © 2012 John Wiley & Sons, Ltd.     

Potassium permanganate–acridine yellow chemiluminescence system for the determination of fluvoxamine,isoniazid and ceftriaxone

Based on the oxidation of acridine yellow by permanganate in basic medium, a new chemiluminescence system was developed for the sensitive determination of some important drugs. The remarkable inhibiting effect of fluvoxamine, ceftriaxone and isoniazid on this reaction was applied to their detection. A possible mechanism was proposed for this system based on chemiluminescence emission wavelengths and experimental observations. Under optimum conditions, calibration graphs were obtained for 1 × 10^?9 to 1 × 10^?6 mol/L of fluvoxamine; 2 × 10^?8 to 8 × 10^?6 mol/L of ceftriaxone and 5 × 10^?8 to 4 × 10^?5 mol/L of isoniazid. This proposed method was satisfactorily used in the determination of these drugs in pharmaceutical samples and human urine and serum. Copyright © 2014 John Wiley & Sons, Ltd.