动态浊度法测定反渗水中细菌内毒素含量

按《中国药典》(2010年版二部)收载的细菌内毒素检查方法及指导原则,用动态浊度法测定反渗水中细菌内毒素。将反渗水稀释32倍的无干扰因素影响,结果显示细菌内毒素回收率在90%以上。动态浊度法在反渗水稀释32倍时可进行反渗水细菌内毒素含量的定量检测。     

动态浊度法在复方甘油细菌内毒素检测中的应用

目的:探讨使用MB-80微生物动态检测系统应用动态浊度法进行复方甘油细菌内毒素检测的可行性.方法:确立动态浊度法检测复方甘油细菌内毒素的方法,然后将18份复方甘油样本使用动态浊度法和凝胶法进行检测,比较结果是否一致.结果:动态浊度法和凝胶法实验结果一致.结论:使用MB-80微生物动态检测系统应用动态浊度法检测复方甘油细菌内毒素是可行的.     

氯化钠注射液内毒素检测方法的探讨

目的采用凝胶定性法和动态浊度定量法对氯化钠注射液细菌内毒素检测比较,以选择内毒素检测的适宜方法。方法在两种试验环境下,分别应用凝胶法和动态浊度法对3个厂家批次的氯化钠注射液进行细菌内毒素检测,对检测结果进行对比分析。结果凝胶法在净化环境下通过无菌操作所得的检测结果未出现假阳性,而普通试验环境下则发现了两份假阳性(60份供试样品,占3.33%);动态浊度法在两种试验环境下所得结果一致,内毒素实测值均<0.00735EU/ml,未出现内毒素实测值超标现象。结论凝胶法检测内毒素试验易受试验环境的影响,普通试验环境下易出现假阳性结果;动态浊度法检测内毒素则不受试验环境影响,在普通实验室即可开展,且操作简单,应成为氯化钠注射液内毒素检测方法的首选。     

细菌内毒素暴露对儿童特应性致敏的影响

[目的]了解居室内毒素含量对儿童特应性致敏的影响。[方法]向安庆、淮南地区2986名8或12岁在学儿童父母发放哮喘标准调查表,动态浊度法检测儿童床铺尘内毒素含量,RAST-FEIA（放射性变应原荧光酶免疫吸附实验）检测其血清中特异性IgE（sIgE）含量。[结果]最终具有完整资料为904人,居室内内毒素含量与特应性致敏呈负相关（OR=0.65,P﹤0.05）。[结论]居室内毒素的高水平暴露可能是预防儿童特应性致敏产生的重要保护因素之一。     

定量检测茵栀黄注射液细菌内毒素方法的建立及应用

目的建立茵栀黄注射液中细菌内毒素的定量检测方法,与家兔法热原检测结果相比较,以确定动态显色基质法用于茵栀黄注射液中细菌内毒素定量检测的可行性。方法采用光度法中的动态显色基质法对茵栀黄注射液进行干扰试验,以明确供试品对动态显色法鲎试剂是否有干扰作用,确立不干扰浓度,并与家兔法热原检测结果相比较。结果使用动态显色法鲎试剂对茵栀黄注射液稀释至37.5倍时无干扰作用,回收率为50%～200%;家兔法检查3批茵栀黄注射液热原结果均符合规定,动态显色法与家兔法检查结果一致。结论动态显色基质法用于茵栀黄注射液中细菌内毒素含量的定量检测是可行的,相较于家兔法热原检查,其结果更稳定可靠,操作方便快捷,灵敏度高。     

乙型脑炎减毒活疫苗细菌内毒素凝胶法的检测

目的建立乙型脑炎减毒活疫苗细菌内毒素检测方法和质量标准,检测样品中的内毒素含量。方法中国药典2010年版三部细菌内毒素检测法(凝胶限度试验、凝胶半定量试验)。结果选用λ为0.5 EU/ml的鲎试剂,采用凝胶限度法对乙型脑炎减毒活疫苗进行细菌内毒素检查,样品经稀释至80倍或以上稀释度后对内毒素检查均无干扰,确定该疫苗的内毒素限度值为40 EU/ml[即20 EU/剂(0.5 ml)]。凝胶限度试验结果显示,疫苗每1 ml中含内毒素的量均小于40EU,即每1次人用剂量(0.5 ml)中含内毒素的量均小于20 EU。凝胶半定量测定结果显示,1人份规格疫苗中含内毒素的量为2.50～10.00 EU/ml(平均5.36 EU/ml);5人份规格疫苗中含内毒素的量为1.50～6.00 EU/ml(平均3.02 EU/ml)。结论凝胶法两种试验均可用于乙型脑炎减毒活疫苗细菌内毒素检测,与凝胶半定量试验相比,凝胶限度试验更为简便、且易于操作;采用凝胶限度法本研究建立了乙型脑炎减毒活疫苗细菌内毒素检测方法和质量标准;凝胶半定量法测定结果用准确数据显示该疫苗的内毒素含量均在人体安全限度以内。     

细菌内毒素定量检测在炎性疾病中的诊断意义

目的:探讨细菌内毒素定量检测在炎性疾病中的诊断意义.方法采用动态浊度法.结果:实验组细菌内毒素定量较对照组及CRP组和WBC组显著增高(P＞0.01),其次CRP组较对照组及WBC组增高(P＞0.05),最后WBC组较对照组轻微增高(P＞0.05).结论:细菌内毒素水平检测是诊断和监测细菌性疾病感染的一个重要参数,通过内毒素水平的定量快速检测可以鉴别诊断细菌性和非细菌性感染性和炎症,且细菌内毒素定量检测在监控严重感染时是比CRP、白细胞计数更好的工具.     

血站采血用品细菌内毒素检测的临床意义

目的：通过检测一次性采血器材和注射用液体制剂的细菌内毒素含量，杜绝不合格产品进入血站，为医疗用血机构提供安全、合格的血液制品。方法：对12份检测标本用动态浊度法检测。结果：12份检测标本有2份内毒素含量超标，合格率为83．33％。结论：早期、有效检测一次性采血器材和注射用液体制剂的细菌内毒素是预防内毒素血症发生的重要措施。     

浸提温度对细菌内毒素影响的初步研究

目的:为热原实验和细菌内毒素实验确定合适的浸提温度。方法:采用《中国药典》2010年版(二部)附录检测细菌内毒素的动态浊度法,在不同浸提温度下测定细菌内毒素溶液中细菌内毒素的含量。结果:溶液中的细菌内毒素含量在121°C1h和80°C 1h的浸提条件下会大大减少,在37°C 1h的浸提条件下不会发生明显改变。结论:热原实验和细菌内毒素实验最适合的浸提温度是37°C。     

细菌内毒素的热压稳定性研究

目的　对比研究环境内毒素与精制内毒素的热稳定性 ,考察热压灭菌前后细菌内毒素水平的相关性 ,为静脉输液相关医疗器具及药品生产过程的质量控制和医院感染工作提供定量参考依据。方法　分别制备环境内毒素与精制内毒素溶液 ,经两次热压灭菌处理 ,EDS99内毒素测定系统动态浊度法定量测定不同处理后样品的细菌内毒素水平 ,对测定结果进行统计分析和相关回归分析。结果　经单次热压处理后 ,环境内毒素和精制内毒素的对数衰减分别为 (0 .73± 0 .2 3)、(0 .97± 0 .5 8) ,经第二次热压处理后 ,两者分别继续衰减 (0 .6 8± 0 .14)、(0 .86± 0 .5 2 )。结论　环境内毒素与精制内毒素的热稳定性差异无显著性 (P>0 .0 5 ) ,热压处理后的细菌内毒素水平与起始水平有显著的相关性 [r>r0 .0 1 (4 ) ],推广应用细菌内毒素定量检查法具有重要的实践意义。     

参麦注射液细菌内毒素检查方法的探讨

目的：考察参麦注射液细菌内毒素检查方法的可行性,加强对参麦注射液的质量监控。方法：采用细菌内毒素检查法,考察参麦注射液对鲎试剂的干扰试验,并检测供试品细菌内毒素的限度。结果：对样品进行1∶4稀释,用0.125 EU/mL鲎试剂无干扰,与家兔热源检查法结果一致。结论：与家兔法检查本品热原相比,细菌内毒素法可行性高,此方法避免了家兔热源检查法操作的复杂性,且灵敏度高、方便快捷。     

Endotoxin liberation by strains of N. meningitidis isolated from patients and healthy carriers

The main objective of this study was to assess whether the capacity of Neisseria meningitidis to release endotoxin depends upon the type of strain or upon bacterial mass. Endotoxin release was studied in 32 strains isolated from patients with meningococcal infections and in 49 from asymptomatic carriers, using a quantitative test (limulus test with a chromogenic substrate). The results show that the strains from patients release significantly higher amounts of endotoxin than strains from carriers regardless of serogroup and isolation site. No correlation was found between stage of bacterial growth and the amount of endotoxin liberated. These findings suggest that endotoxin liberation is a characteristic of certain strains of N. meningitidis and is not determined simply by bacterial mass.     

Pyrogenic reactions associated with single daily dosing of intravenous gentamicin.

OBJECTIVE: To identify risk factors associated with an unexpected outbreak of pyrogenic reactions (PR) following intravenous gentamicin. DESIGN: We conducted two cohort studies. PRs were defined as chills, rigors, or shaking within 3 hours after initiating the gentamicin infusion during the preepidemic (December 1, 1997-January 15, 1998) or epidemic (May 1-June 15, 1998) periods. We tested gentamicin vials for endotoxin using the limulus amebocyte lysate assay. SETTING: Inpatient services of a large community hospital in Los Angeles, California. RESULTS: During the epidemic period, 22 (15%) of 152 patients developed documented PRs following intravenous gentamicin. PRs were more likely among patients receiving single daily dosing (SDD) than multiple daily dosing gentamicin (20/73 [27%] vs. 2/79 [3%]; relative risk, 10.8; 95% confidence interval, 2.6 44.7). Laboratory analysis of gentamicin vials found endotoxin levels that were higher among Fujisawa-brand gentamicin (implicated brand) than gentamicin used after the outbreak terminated (non-implicated brand). Although endotoxin levels in the vials did not exceed US Pharmacopeia limits (1.7 endotoxin units/mg gentamicin), the use of SDD gentamicin may place patients at greater risk of receiving doses of endotoxin above the threshold for PRs in humans. CONCLUSIONS: Reassessment of the acceptable amounts of endotoxin in gentamicin and other parenteral products should be considered when dosing intervals used in clinical practice change.     

Endotoxaemia in Vibrio El Tor cholera

Endotoxaemia and endotoxin-induced changes were sought in Nigerian patients presenting with cholera/diarrhoea. The organism was Vibrio cholerae, bio-type El Tor, serotype Hikojima. The limulus amoebocyte lysate gelation test was used qualitatively by the clot method, whilst a spectrophotometric method was used quantitatively to measure endotoxin levels. 25 acutely ill patients tested had detectable endotoxaemia by the Escherichia coli endotoxin standard. The highest endotoxin level was found in a patient with sub-conjunctival haemorrhage. Changes in platelet counts, the detection of complement breakdown product C3d in plasma, the elevation of fibrin degradation products, the finding of elevated, normal or depressed C3 levels and the absence of circulating immune complexes, suggest a pathogenic role for endotoxin in Vibrio cholerae El Tor diarrhoea.     

建立可吸收医用膜细菌内毒素检查的新方法

[目的]检验粘克^TM可吸收医用膜是否符合生物材料的热原要求,建立粘克^TM可吸收医用膜的细菌内毒素检查方法。[方法]按《中华人民共和国药典》2005年版附录及GB/16175-1996规定,同时采用热原检查法和细菌内毒素检查法对粘克^TM可吸收医用膜72h的浸提液样品进行检测。[结果]该浸提液对细菌内毒检测不产生干扰,细菌内毒素试验结果为阴性。同时受试家兔体温升高均低于0.6℃,并且3只家兔体温升高总和低于1.4℃。[结论]粘克^TM可吸收医用膜不含引起机体发热的内毒素或致热原,符合医用生物材料的使用标准。用细菌内毒素检查法替代家兔热原法对该可吸收膜进行热原检查是可行的。     

Environmental endotoxin measurement: the Kinetic Limulus Assay with Resistant-parallel-line Estimation.

A Limulus assay method was specifically designed for environmental endotoxin aerosols. Application of new statistical and sample preparation methods strengthened the validity and precision of the Limulus test. Statistically, the Kinetic Limulus Assay with Resistant-parallel-line Estimation (KLARE) differed from conventional analytic methods (as used in chromogenic assays and other kinetic methods) by routinely using a dilution series of the unknown sample as well as the standard to compute potency and an estimate of variance for each sample. Analysis of dose-response slopes for the standard and unknowns detected inhibition and enhancement effects--without multiple assay. Concentration-dependent interference and a more complex, concentration-independent interference with the Limulus assay were detected. Resistant regression and a standardized data analysis corrected for concentration-dependent interference. Sample preparation in a buffer eliminated concentration-independent interference and, thus, improved both the validity and the precision of potency measurements. The utility of a sample buffer and of parallel-line analysis, with both turbidimetric and chromogenic lysates, was demonstrated by assay of three control standard LPS and reference LPS (EC5). The limit of detection for endotoxin was less than 1 pg/ml in buffer. Samples containing greater than or equal to 10 pg/ml were measured with a coefficient of variation of approximately 6% in a single assay. Reproducibility of potency estimates for four samples over 3 days was compared on the basis of standard errors of the mean. The conventional method gave on average a CV of 65% while the resistant-parallel-line method gave, on average, a CV of 6%. Also, the conventional method failed to detect interference and, thus, included data from invalid assays. Conventional analysis of environmental aerosol samples was highly sensitive to the choice of dilution factor causing as much as 1000% variation in the result. By contrast, KLARE results changed by at most 30% with similar changes in initial dilution because KLARE was able to detect, and correct for, the influence of interferant compounds.     

一次性采血袋细菌内毒素的动态浊度检测法

目的研究使用MB-80微生物动态检测系统即动态浊度法进行采血袋内毒素检测的可行性。方法建立动态浊度法检测采血袋内毒素的方法 ,然后将20份采血袋样本使用动态浊度法和凝胶法进行实验,比较结果。结果动态浊度法和凝胶法检测20份采血袋均18份合格,2份不合格,结果一致。结论使用MB-80微生物动态检测系统应用动态浊度法检测采血袋内毒素是可行的。     

Detection of lipopolysaccharide in suspected bacteriuric urine using a carbocyanine dye.

Currently practiced methods for the detection of gram negative bacteriuria require culturing and overnight incubation. Such an approach to bacteriuria detection is unacceptable for any screening program which requires rapid presumptive evidence of infection. In this study, the lipopolysaccharide-dependent formation of a unique dye absorption spectra of the cationic carbocyanine dye, 1-ethyl-2-[3-(1-ethylnaphtho[1,2d]-thiazolin-2-ylidene)-2-methylpropenyl] naphtho[1,2d]-thiazolium bromide, was used to detect bacteriuria caused by gram negative organisms in a hospitalized population. In an evaluation of 168 first morning and randomly collected suspected bacteriuric urines, the dye test detected 66% of the loop plate positive urines with false positive and false negative values of 28% and 34%, respectively. However, 37% of the false positive results occurred in urines containing less than 10(5) gram negative bacteria/ml and an additional 24% of the false positives were seen for patients currently receiving antibiotic treatment. Urine specimens were also evaluated using the limulus lysate assay for lipopolysaccharide.     

Work area measurements as predictors of personal exposure to endotoxin and cotton dust in the cotton textile industry

OBJECTIVES: To determine if work area measurements of endotoxin and/or cotton dust obtained from the vertical elutriator (VE) can be used to predict levels of personal endotoxin exposure as measured by the Institute of Occupational Medicine (IOM) inhalable dust sampler in the cotton textile industry. METHODS: Fifty-six work area cotton dust samples were collected from 14 areas and 82 personal cotton dust samples were collected from 41 workers in three textile mills (Mills A, B and C) in Shanghai, China. Cotton dust concentrations were determined gravimetrically from sample filters, of which endotoxin concentrations were determined using a kinetic chromogenic modification of the limulus amoebocyte lysate assay. Linear regression models were used to determine the association between log IOM personal endotoxin concentration and log VE area endotoxin concentration. RESULTS: Median cotton dust and endotoxin concentrations measured from VE area samples in the three mills were 0.36 mg m(-3) and 1280.76 endotoxin units per cubic meter (EU m(-3)), respectively, compared to 1.74 mg m(-3) and 2226.83 EU m(-3) from IOM personal samples. Excluding samples from weaving processes, we observed linear associations between VE area measures of endotoxin and IOM personal endotoxin concentrations; VE area concentration of endotoxin explained 83 and 89% of the total variation in IOM personal endotoxin concentration for Mills A and B, respectively (Mill A: R2 = 0.83, P < 0.0001; Mill B: R2 = 0.89, P < 0.0001). Although area measures of cotton dust was also a significant predictor of person endotoxin, the model explained less of the variance in personal endotoxin measurements. CONCLUSIONS: Specific to the conditions of the textile mills investigated in this study, work area measurements of endotoxin, but not cotton dust, may be reasonable proxies for personal levels, at least for rank-ordering exposures.     

Lysozyme Reference Standard (Control 031) of National Institute of Health Sciences

The "Lysozyme Reference Standard (Control 951031)" of the National Institute of Health Sciences was prepared. The lysozyme potency of the standard material was assayed against the Lysozyme Reference Standard (Control 951) by turbidimetric method two turbidimetric methods using the dried y-cells of Micrococcus luteus as a substrate. The potency of the standard material was in satisfactory agreement with that of Lysozyme Reference Standard (Control 951) and was defined as 1 mg [potency] per mg.