特发性高草酸尿症大鼠空肠蛋白质组二维电泳可行性研究

目的：在正式进行双向凝胶电泳实验前需要进行二维电泳的可行性评价,为双向凝胶电泳实验的顺利进行奠定实验基础。方法：取1只特发性高草酸尿大鼠（IH）和正常对照大鼠的空肠组织300mg,匀浆提取组织总蛋白,以双向电泳技术分离蛋白质,银染后扫描获得图谱,并以ImageMaster^TM 2D Platinum software（Version5．0）软件进行分析。结果：获得了清晰、稳定的凝胶蛋白图谱,正常大鼠凝胶图谱可检测到2541个蛋白点,IH大鼠凝胶图谱可检测到2654个蛋白点。结论：从获得的二维电泳图谱来看,2个蛋白质样品的蛋白质点的迁移基本稳定,从而保证后续正式的双向凝胶电泳实验有较好的重复稳定性和可行性。     

The diagnostic dilemma of congenital unstable hemoglobinopathies

Unstable hemoglobin variants represent a rare etiology of congenital hemolytic anemia. Without a high index of suspicion, plus proper laboratory testing and interpretation, the correct diagnosis can be elusive. We report on five children who were initially thought to have other congenital disorders such as hereditary spherocytosis or thalassemia, before β‐globin gene sequencing led to the definitive diagnosis. Recognizing the variable clinical presentation and laboratory data reported will aid clinicians in diagnosis of unstable hemoglobins variants in children with atypical forms of hemolytic anemia, particularly those with low pulse oximetry values or whose hemoglobin electrophoresis suggest β‐thalassemia trait. Pediatr Blood Cancer. 2010;55:1393–1395. © 2010 Wiley‐Liss, Inc.     

A simplified method of determining synovial fluid chondroitin sulfate chain length

OBJECTIVE: To determine whether dimethylmethylene blue (DMMB) analysis, when combined with agarose gel filtration chromatography (Superose 6), can be performed instead of fluorophore-assisted carbohydrate electrophoresis (FACE) to determine chondroitin sulfate (CS) chain length in synovial fluid (SF). METHODS: SF was obtained from (1) normal horses after 8 weeks of rest, (2) the same horses after 9 months of treadmill training, and (3) horses with osteochondral (OC) injury from racing. SF CS concentrations and chain lengths were determined by gel chromatography and DMMB analysis and compared with previous results determined by FACE analysis on the same samples. RESULTS: DMMB analysis showed that SF CS peak chain length in the OC injury group increased significantly (18.7 kDa) when compared to rested and exercised normal horses (15.6 kDa). The assay had a positive predictive value of 71% and a negative predictive value of 75% for discriminating between normal and injured joints. CONCLUSIONS: We report a simple and inexpensive DMMB analysis of SF CS chain length, which, when coupled with Superose 6 chromatography, discriminates between normal and post-injury joints. Similar to our previous FACE analysis results [Brown MP, Trumble TN, Plaas AHK, Sandy JD, Romano M, Hernandez J, et-al. Exercise and injury increase chondroitin sulfate chain length and decrease hyaluronan chain length in synovial fluid. Osteoarthritis Cartilage 2007;15], our DMMB results show an increase in the chain length of the CS in the SF of injured joints.     

Simultaneous determination of streptomycin and oxytetracycline in agricultural antimicrobials by CZE after an experimental design

A capillary zone electrophoresis (CZE) method was developed and validated for the simultaneous determination of both streptomycin (STP) and oxytetracycline (OTC) in bactericidal products to be used in agriculture. Using fused-silica capillaries, the influence of the electrolyte composition, pH and concentration, as well as temperature and applied voltage were investigated using a central composite design to optimize the method. The optimized electrophoretic conditions were as follows: 0.10 M sodium phosphate, pH 2.5, 7.0 kV and 20.0 degrees C. The method was validated for STP and OTC determination in agricultural formulations through the following performance criteria: linearity and linear range, sensitivity, selectivity, intra-day and inter-day precision, detectability, accuracy and ruggedness. This optimized CZE-method for the identification and quantification of STP and OTC is a potential alternative method to the HPLC methods described by the US Pharmacopeia, with the advantage that the same method could be used for the simultaneous determination of these different antibiotics.     

Determination of kava lactones and flavonoid glycoside in Scorzonera austriaca by capillary zone electrophoresis

A capillary zone electrophoretic method has been developed for the quantitative analysis of three active comppounds, 12-hydroxy-desmethoxyyangonin (HD), 12-beta-d-glucopyranoside-desmethoxyyangonin (GD) and luteolin 3'-(6-E-p-coumaroyl-beta-d-glucopyranoside) (LG) in Scorzonera austriaca with UV detection at 254 nm. The applied voltage was 25 kV and the capillary temperature was kept constant at 25 degrees C. The effect of buffer pH, the concentration of electrolyte and organic modifier on migration were studied systematically. Optimum separation was achieved with 20 mM borate buffer at pH 10.00 containing 10% (v/v) methanol. Daphnetin was used as internal standard for quantification. Regression equations revealed good linear relationship between the ratios of the peak area of each compound and its the ratios of concentration. All the correlation coefficients were higher than 0.9990. The relative standard deviations of migration time and the peak area were <1.46% and 5.13% (inter-day), and <1.65% and 5.16% (intra-day), respectively. The contents of the three compounds in S. austriaca were successfully determined with satisfactory repeatability and recovery.     

肝癌发生不同病理阶段血清蛋白质组的比较

目的 比较肝癌发生不同病理阶段血清蛋白质组的表达变化情况,从中寻找特异性标志物.方法 用双向凝胶电泳分离正常人,慢性肝炎、肝硬化和肝癌患者的血清蛋白质,结合质谱技术对差异点进行鉴定; Western blot检测结合珠蛋白β链以验证蛋白质水平的表达.计量资料数据以均数±标准差(x±s)表示,采用方差分析和LSD检验进行两两比较.结果 4组血清的双向电泳图谱经软件匹配分析,并与基质辅助激光解析电离飞行时间质谱相结合,成功鉴定出结合珠蛋白,SAA1、SP40等30个差异蛋白质点,K cluster聚类分析不同的变化模式.在差异蛋白质点中,7个蛋白质点均为结合珠蛋白,呈现由正常→肝炎→肝硬化持续下调、到肝癌又上调的模式.Western blot证实结合珠蛋白β链的表达与双向电泳表达相一致.结论 在肝癌发生的动态过程中各疾病阶段的血清蛋白质差异表达有4种明显变化模式,可能为肝癌的早期诊断和预后提供依据,与肝癌发生发展相关的结合珠蛋白很可能是肝硬化发展到肝癌的一个潜在早期诊断标志物.     

低剂量T-2毒素与脱氧雪腐镰刀菌烯醇单独及联合染毒对小鼠末梢血有核细胞DNA损伤的观察

目的 观察低剂量T-2毒素和脱氧雪腐镰刀菌烯醇(DON)对小鼠末梢血有核细胞DNA损伤的特点及其程度,探索亚临床剂量下的毒性作用.方法 3周龄雌性Balb/c小鼠80只,体质量(14.0±1.5)g.将小鼠按体质量随机分为对照组、T-2毒素组、DON组、T-2+DON联合组,每组20只.经腹腔注射感染毒素,染毒剂量:T-2毒素5 μg·kg-1·d-1,DON 20 μg·kg-1·d-1;对照组给予等量生理盐水.染毒至16周和21周,分别取小鼠尾血,用单细胞凝胶电泳法(SCGE)观察小鼠末梢血有核细胞拖尾率、尾DNA含量、尾长、尾动量变化.结果 ①T-2毒素组,尾DNA含量、尾长、尾动量[16周:(27.71±15.85)%、(13.67±5.56)μm、4.26±3.83;21周:(28.38±15.57)%、(13.83±5.47)μm、4.37±3.82]均增加,与对照组[16周:(11.87±4.61)%、(10.59±6.70)μm、1.34±0.98;21周:(11.31±3.94)%、(10.83±7.05)μm、1.29±1.01]比较,差异有统计学意义(P均<0.05);②DON组拖尾率、尾DNA含量、尾动量[16周:5.62%、(28.13±13.31)%、3.39±2.35;21周:7.71%、(29.17±15.12)%、5.70±4.17]均增加,其中仅拖尾率与对照组(16周:4.34%;21周:4.38%)比较,差异有统计学意义(P均<0.05);③T-2+DON联合组拖尾率、尾DNA含量、尾长、尾动量[16周:6.21%、(30.14±15.48)%、(16.93±6.58)μm、5.54±4.22;21周:8.17%、(30.85±15.76)%、(17.21±6.45)μm、5.70±4.17]均增加,与对照组比较差异有统计学意义(P均<0.05);与T-2毒素或DON组比较,尾DNA含量和尾长均增加(P均<0.05).结论 低剂量T-2毒素和DON均可对小鼠未梢血有核细胞DNA产生明确损伤;T-2和DON联合染毒较单独染毒的损伤作用更明显.     

苦参对阴道毛滴虫蛋白质差异表达的影响

目的寻找阴道毛滴虫的特异蛋白质并探讨苦参对其差异蛋白质表达的影响。方法用固相pH梯度双向凝胶电泳分离阴道毛滴虫总蛋白质,凝胶用银染显色,PDQuest7.4.0软件分析。结果同组实验重复3次以上,其中正常对照组找到蛋白质点平均（435±17）个,匹配率为92.5%;苦参组找到蛋白质点平均（336±26）个,匹配率为85.6%。正常组和苦参组共有96个表达差异点,其中苦参组和正常组比较,只在正常组中表达的点有46个,只在苦参组中表达的点有27个,表达增加〉2倍的点有9个,表达降低〉0.5倍的点有14个。结论建立了阴道毛滴虫正常组和苦参组的双向凝胶电泳图谱,识别了96个差异表达的蛋白质。苦参杀灭阴道毛滴虫的作用可能与上述蛋白质的改变有关。     

The prevalence of microsatellite instability in head and neck squamous cell carcinoma

Purpose  The present study aimed to use the most definitive available techniques to resolve controversy in the literature as to the prevalence of microsatellite instability (MSI) in head and neck squamous cell carcinoma (HNSCC). Methods  Eighty patients with advanced HNSCC were enrolled in the study that examined 20 microsatellite markers with automatic fragment analysis. These markers included ones derived from the NCI reference panel and ones previously reported to detect MSI in HNSCC (HNSCC panel). Results  Only one of 80 tumors could be considered positive for MSI. For this case, both panels showed MSI-High (8/10 positive markers for the NCI reference panel and 6/10 positive markers for the HNSCC panel). Qualitatively, the observed MSI could be classified as Type B MSI. Conclusions  The present results indicate that MSI has a low prevalence in HNSCC.