尿液结合性胆红素的纯化

目的　探索一种来源方便 ,具有延续性 ,利于胆红素测定校准物或质控物制备 ,且方法简便、实用、产率和纯度可靠的结合性胆红素 (Bc)纯化方法。方法　应用 CHCl3 ∶ CH3 CH2 OH(1∶ 1,V/V)萃取与 CNBractivated Sepharose4B- HSA亲和层析相结合的方法对胆道梗阻性高胆红素尿液进行分离纯化 ,并以 Doum as J- G法和 HPL C分析产品。结果　由 CHCl3 ∶ CH3 CH2 OH(1∶ 1,V/V)萃取所得的粗纯品经 J- G法检测产率为 90 .2±5 .4% (n=3) ,胆红素含量占总干重的 6 7.9± 6 .4% (n=3)。亲和层析的洗脱峰间的分离度为 1.5、1.737。层析后再萃取所得精纯品经 HPL C分析 ,产率为 46 .9% ,纯度 94.7% ,其中 BDG76 .2 % ,BMG18.5 %。结论　采用有机物萃取和 CNBr activated Sepharose4B- HSA亲和层析相结合的方法纯化结合性胆红素 (BMG& BDG)具有很好的可靠性和可行性     

以结合性胆红素制备胆红素测定校准物的研究

OBJECTIVES: To improve the accuracy and precision of the determination of bilirubin, especially direct bilirubin (DB), and the standardization of that as well. METHODS: Purified conjugated bilirubin (Bc) and ditaurobilirubin(DTB) and their diazo products were subjected to absorption spectrum analysis. The diazo reaction characters of their calibration solutions were compared by the method of Doumas J-G(TB & DB). RESULTS: Bc, DTB and their azopigments were found to have the similar absorption spectra with the same lambda max. Their TB standard curves almost superposed together all over. Although the slopes of their DB standard curves were not markedly different ((YBc = 0.00366X + 0.00933, rBc2 = 0.9977, P < 0.01; YDTB = 0.00391X + 0.00023, rDTB2 = 0.9987, P < 0.01; Pb1-b2 > 0.05, n1 = n2 = 5), the DB value measured for Bc differed from that for DTB(n = 5, P < 0.05). In addition, the calibrators made from Bc based different matrices, such as HSA, BSA and human serum, were significantly different in DB/Bc, but no difference was seen among the concentrations. Furthermore, the DB values determined for DTB or Bc increased linearly with the corresponding concentrations, respectively, with no difference between the slopes (YBc = 0.8300XBc + 1.9463, rBc2 = 0.9977, P < 0.01; YDTB = 0.8853XDTB-0.0251, rDTB2 = 0.9986, P < 0.01; n1 = n2 = 5, Pb1-b2 > 0.05). CONCLUSIONS: The results demonstrate that the diazo reaction characters of Bc are identified with those of DTB. However, under the condition of DB, Bc reacts differently from DTB. This study also indicates that as a calibrator of DB based human serum, Bc has the similar constant effect of HCl as serum samples do, so it is a more reliable calibrator to eliminate the matrix effects.     

快速纯化胆红素的实验研究

通过正交实验分析纯化胆红素的最佳实验条件,并确立制备胆红素的工艺过程.此工艺具有简便、快速、低成本和高收率的优点.     

以结合性胆红素制备胆红素测定校准物的研究

目的　以结合性胆红素制备胆红素测定校准物 ,改善胆红素尤其是直接胆红素测定的准确度和精密度。方法　对结合性胆红素 (Bc)和二牛磺酸胆红素 (DTB)及其重氮产物进行吸收光谱分析 ,并比较其 DoumasJ- G法总胆红素、直接胆红素反应特征。结果　结合性胆红素和 DTB及其重氮产物具有相似的吸收光谱 ,最大吸收峰也在相同位置 ,两者的总胆红素标准曲线几乎完全重合。它们的直接胆红素测定值具有显著性差异 (P<0 .0 5 ,n=5 ) ,但直接胆红素标准曲线斜率无显著性差异 (Y结合性胆红素 =0 .0 0 36 6 X+0 .0 0 933,r结合性胆红素 2 =0 .9977,P<0 .0 1;YDTB=0 .0 0 391X+0 .0 0 0 2 3,r DTB2 =0 .9987,P<0 .0 1;Pb 1 - b2 >0 .0 5 ,n1 =n2 =5 )。结合性胆红素在不同基质(牛血清白蛋白、人血清白蛋白、人血清 )中的直接胆红素 /结合性胆红素值有显著性差异 (P<0 .0 1,n=5 ) ,而不同浓度间无明显差异 (P>0 .0 5 ,n=5 ) ,其不同浓度下结合性胆红素、DTB与相应的直接胆红素测定值均呈线性趋势 ,但这两者斜率同样无显著性差异 (Y结合性胆红素 =0 .830 0 X结合性胆红素 +1.946 3,r结合性胆红素 2 =0 .9977,P<0 .0 1;YDTB=0 .885 3XDTB- 0 .0 2 5 1,r DTB2 =0 .9986 ,P<0 .0 1;n1 =n2 =5 ,Pb1 - b2 >0 .0 5 )。结论　结合     

273份蛋白阴性胆红素阳性尿液的管型分析

目的：观察干化学检验蛋白阴性胆红素阳性尿液沉渣中管型的阳性率和类型。方法选取干化学检测蛋白阴性胆红素阳性尿液样本273份,检验尿沉渣中管型的阳性率和类型。结果胆红素（＋）管型阳性率为2.4％,胆红素（＋＋）管型阳性率为6．8％,胆红素（＋＋＋）管型阳性率为14．2％。其中胆红素管型占80．9％,透明管型14．3％,颗粒管型4．8％。结论尿干化学检测蛋白阴性胆红素（＋＋）及以上管型阳性率明显升高,以胆红素管型为主。干化学蛋白阴性胆红素阳性尿液应联合显微镜检测,提高管型检测的准确性。     

良性梗阻性黄疸病人结合胆红素和非结合胆红素的预后价值

目的 :通过对 37例良性梗阻性黄疸病人围减黄手术期血清总胆红素 ( TBil)、结合胆红素 ( Bc)和非结合胆红素 ( Bu)的测定 ,了解其变化规律 ,确定其临床意义。方法 :分别于手术前 1d,手术后 1,3,7,10 ,14 ,18,2 1d采集血标本 ,使用美国干式生化分析仪测定其中 TBil、Bc、Bu的含量。结果 :排泄良好组在减黄术后 TBil、Bc、Bu前几天迅速下降 ,之后平稳消退 ,Bc/TBil不断下降 ,7d后绝大多数病例小于 0 .2 ;排泄不良组中不少病例在术后 TBil、Bc、Bu不但不消退 ,反而有一定的升高 ,原因不明 ,之后缓慢减低 ,在减黄过程中无明显消退阶段 ,Bc/TBil始终大于 0 .2。结论 :Bc/TBil是预测减黄效果的良好指标     

新生儿未结合胆红素大于总胆红素原因分析

临床检验常出现患者的胆红素结果不配套,直接胆红素(DB)结果反而比总胆红素(TBIL)结果高,使检验人员难以解释。特别是干化学法问世以来,尤其是用于新生儿黄疸的检测中,此种现象尤为突出。结合胆红素(DC)常常为零,未结合胆红素或未结合胆红素加结合胆红素的结果大于总胆红素结果。使传统湿化学法测定的总胆红素和直接胆红素,两者相减,等于间接胆红素的方法受到置疑。新生儿黄疸是新生儿最常见的症状,有生理性和病理性之分。准确有效地测定新生儿胆红素(NBIL)对儿科医生正确进行鉴别诊断,尽早采取合理治疗措施,预防核黄疸发生有着十分重…     

遗传性非结合性高胆红素血症研究进展

遗传性非结合性高胆红素血症是由尿苷二磷酸葡萄糖醛酸转移酶（uridine diphosphate glucuronosyl transferase 1A1,UGT1A1）基因突变所致的以非结合胆红素升高为主的高胆红素血症,临床以不同程度的黄疸为主要表现,没有溶血性疾病指征或肝功能异常指标。根据 UGT1A1 酶缺乏程度及血清非结合胆红素水平,分为 Gilbert 综合征、Crigler–Najjar 综合征Ⅰ型及Ⅱ型。本文将对非结合性高胆红素血症 UGT1A1 基因研究进展、临床诊断、治疗和预后等方面进行综述     

Purification and antimicrobial activity of human neutrophil defensins]

Neutrophils are one of the weapons of host defenses against microbial infection. Their ability to kill the invading microorganisms depends on two principle mechanisms. One depends on production of reactive oxygen intermediates (ROI) by stimulated neutrophils, and the other depends on the delivery of antimicrobial contents of the neutrophils' cytoplasmic granules, oxygen-independent. The defensins have the highest concentration in the neutrophils, and the broadest antimicrobial spectrum, being capable of killing gram-positive and gram-negative bacteria, fungi and some envelope viruses. We purified human defensins from the neutrophils' granules by gel permeation chromatography and SDS-preparative acrylamide gel electrophoresis. The molecular weight of human defensins is between 3,000-4,000 daltons. After testing, C. neoformans was susceptible to these defensins. Under condition of 37 degrees C, pH 7.4 and low ionic strength, antifungal activity by human defensins was related to its concentration and incubating time. All of these illustrate that nonoxidative killing mechanism of neutrophils, especially the function of defensins is very important in host defenses.     

血清胆红素对脂蛋白的影响

观察２３７例受试者空腹血清胆红素对血脂的影响，并比较不同血清总胆红素水平组间血脂的变化。结果显示，血清总胆红素及直接胆红素水平与总胆固醇和低密度脂蛋白胆固醇呈负相关，提示胆红素影响血脂代谢，低工素血症可能是冠心病一种析的危险因子。     

高效液相色谱法测定尿中苯妥英代谢物浓度

A high-performance liquid chromatographic method for the determination of p-hydroxyphenytoin (p-HDPH) in human urine was reported. Following acid hydrolysis of urine sample at 90 degrees C for 1 h, an internal standard, nitrophenol, was added. The sample was extracted with n-hexane-ethyl acetate (50:50), and organic layer was evaporated. The residue was dissolved in methanol and chromatographed on an Ultrasphere-ODS column, using a mobile phase of phosphate buffer (0.03 mol/L, pH 6.0): methanol (65:35) at a flow rate of 0.8 ml/min. The eluent was monitored at 240 nm. The standard curve was linear within the range 5.0-200 micrograms/ml (r = 0.9998). Analytical recovery rates were 102.8 +/- 7.3% (p-HDPH 9.70 micrograms/ml, n = 5) and 104.9 +/- 6.4% (p-HDPH 54.70 micrograms/ml, n = 5). The cumulative recovery of p-HDPH in 0-12 h volunteers' urine samples accounted for 20% of the oral dose of 100 mg phenytoin sodium.     

尿胆红素检测对妊娠肝内胆汁淤积症的预测价值

目的探讨尿胆红素检测对妊娠肝内胆汁淤积症的预测价值。方法 2008年1～6月对成都市妇产科医院门诊133例临床怀疑为ICP的孕妇进行尿胆红素检测和血胆汁酸检测。以血胆汁酸结果为金标准,对两种检测方法的一致性进行评价。结果尿胆红素阳性者其血胆汁酸水平亦高,两者有一致性。结论尿胆红素检测可随机进行,有很高的特异性,可用于ICP的预测。     

人血浆中MBL-MASP复合物的纯化与分离

目的从人血浆中分离纯化甘露聚糖结合凝集素(MBL)和MBL相关丝氨酸蛋白酶(MASP)。方法用非活化Sepharose 4B两步亲和层析纯化MBL-MASP复合物．再以Sephacryl S-300柱凝胶过滤将MBL和MASP分离。在纯化MBL—MASP复合物的整个过程中，除凝胶过滤缓冲液外，均加入丝氨酸蛋白酶抑制剂苯甲磺酰氟(PMSF)和金属蛋白酶抑制剂1．10-邻二氮杂菲(1，10-phenanthroline)．并控制温度在4℃。结果获得高度纯化的MBL和酶原形式的MASP。SDS—PAGE和Western blotting表明所纯化的MBL相对分子质量为28000和32000肽链构成的功能性多聚体；配体结合测定和酵母菌凝集试验证实所纯化的MBL具有生物学活性：结论建立了一种比较简便的同时分离纯化MBL和MASP酶原的方法。     

从基因重组大肠杆菌分离纯化重组人白介秦－2

从ＥｓｃｈｅｒｉｃｈｉａｃｏｌｉＭ５２４８（Ｂｉｏ２７５ｃＩ８５７ＨＩ）中，经发酵培养，离心，超声破碎，溶解，二次凝胶过滤层析，复性处理及冷冻干燥，一次获得了３６ｍｇ较高纯度的重组人白介素－２。     

脑胶质瘤中性糖脂抗原的分离纯化及分析

应用离子交换层析技术从脑胶质瘤、正常人脑及胎儿脑组织中分离并纯化出中性糖脂（Ｈ─ＧＳＬ），经高效薄层色谱分析，纯度可靠。胶质瘤Ｎ─ＧＳＬ中出现一组特异性条带，主要分布于Ｇｌｃｃｅｒ与ＣＴＨ之间，而正常人脑则以Ｇｌｃｃｅｒ和Ｇａｌｃｃｅｒ表达为主，ＣＤＨ及ＣＴＨ等长糖链Ｎ─ＧＳＬ缺失。胎儿脑组织Ｎ─ＧＳＬ与胶质瘤Ｎ─ＧＳＬ相似，主要分布于ＣＴＨ与Ｇｌｏｂｏｓｉｄｅ之间。     

人血清载脂蛋白A—Ⅳ的纯化,鉴定及逆向转运胆固醇的研究

In order to study the function of apoA-IV, human serum apoA-IV was isolated and purified. LDS was isolated with NaBr density gradient ultracentrifugation and then incubated with intralipid according to the method of Weinberg and Scanu. Protein complex containing apoA-IV was obtained. The apo-IV was purified from protein complex by preparative SDS-PAGE, electroelution and removal of SDS. ApoA-IV thus prepared gave a single band on SDS-PAGE with a MW of 46 kD. Amino acid composition and isoelectric focusing pI (5.27 and 5.47) of apoA-IV were similar to those reported in the literature. The effects of apoA-IV and liposome (apoA-IV: DMPC) on cholesterol efflux from hum skin fibroblast were studied. There was no significant difference (P > 0.5) between apoA-IV and control but a very significant difference (P < 0.001) between liposome (apoA-IV:DMPC) and control, suggesting that liposomes (apoA-IV:DMPC) play a role in reverse cholesterol transport. The results may offer a new approach for the prevention and treatment of atherosclerotic coronary heart disease.     

液相色谱法纯化人血清C2蛋白

目的 建立一种从血清中提纯C2的方法，得到稳定的，有活性的纯品，便于今后进一步研究其生理病理功能，方法 利用冷球蛋白沉淀，优球蛋白沉淀，离子交换色谱，硫酸铵沉淀和亲和色谱几种方法，将血清中不同性质的杂蛋白除去，选择相对简单且分离效果好的条件。提高终产物中目标蛋白C2的纯度和含量。结果 成功地从1L血浆中提纯到了2．49mg C2，产率为12．46％，活性回收率为13．6％，终产物在PH中性时稳定，且无需加入蛋白稳定剂 结论 本法与以往纯化C2的方法相比，简便可行，产率及活性回收率较高，适用于实验室研究和大规模制备。