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1.
A number of techniques are available for the extraction of lipids from a variety of tissues; however, conventional methods are characteristically labor intensive, typically involve large volumes of toxic solvents, and usually require at least 1 g of tissue. With the availability of accelerated solvent extraction (ASE) technology, the opportunity exists to modify classical lipid extraction techniques such that automated high-pressure, high-temperature extractions may be performed with the use of far smaller volumes of costly and harmsul solvents. Moreover, the high extraction efficiency attainable by ASE suggests that significantly less tissue would be required than is routinely used. This paper describes the adaptation of previously developed lipid extraction solvent systems for use with ASE toward the purpose of extracting total lipids from 100 mg of fish tissue. The efficacy of three solvent systems for lipid extraction from representative fish tissues, including a standard reference material, was explored using gravimetry and FA analysis by GC. A TG was used as a surrogate to monitor overall method performance. The findings herein demonstrate that microscale ASE represents an effective and efficient alternative to traditional lipid extraction techniques based on quantity and composition of extracted lipid, surrogate recovery, and precision.  相似文献   

2.
目的应用磁凝集法梅毒检测试剂检测梅毒特异性抗体和反应素。方法应用磁凝集法梅毒检测试剂(Tre-ponema pallidum magnetic particle agglutination,TPMPA)检测梅毒阳性血清,分析梅毒阳性血清符合率、梅毒血清抗体滴度、假阳性检出率及交叉反应。结果用TPMPA-A、TPMPA-B试剂检测26份梅毒阳性血清样品,结果均为阳性,与省血液中心和省疾控中心的检测结果的符合率为100%;应用TPMPA-B试剂检测梅毒抗体滴度结果均较用梅毒甲苯胺红不加热血清试验诊断试剂(TRUST)检测结果高2个滴度;用TPMPA-A、TPMPA-B试剂检测正常血清,结果均为阴性,与省血液中心检测结果相符;用TPMPA-A、TPMPA-B试剂检测5份风湿病患者血清和13份慢性肝病患者血清,均未出现交叉反应。结论磁凝集法梅毒检测试剂具有良好的特异性和敏感性,操作简便、省时,可初步用于检测梅毒特异性抗体和反应素。  相似文献   

3.
This report describes the results of a collaborative study organized by a joint working group of the IFCC and WHO and involving nine manufacturers of TSH immunometric assay kits. The study was designed to determine whether a calibrator with a common matrix gives better between-laboratory agreement for calibration of serum samples than the various kit calibrators, and to assess various materials for their suitability for use as common matrices. Kit calibrators, or calibrators consisting of the IRP for TSH made up in two common matrices: (a) serum from patients with untreated thyrotoxicosis or (b) serum taken from subjects treated with suppressive doses of triiodothyronine, gave similar results for the between-laboratory variation of estimates of TSH concentration for a range of serum samples. Dose-response curves for the two calibrators in ‘common’ matrices were similar to one another and to those for the kit calibrator. However, the occurrence of non-specific serum effects is shown by the comparison of results for these calibrators with results for calibrators made up in a third common matrix: serum treated with wheat germ lectin. Dose response curves for this calibrator were dissimilar to those for the other calibrators and between-laboratory variation for estimates in terms of this latter calibrator showed a substantial increase. Moreover, although the between-laboratory variances for estimates of the TSH concentration in terms of each of these calibrators (except those made up in serum treated with the wheat germ lectin) were similar for any one sample from five hyperthyroid patients, the variances were not consistent between samples, even for samples with similar mean TSH concentrations. These results suggest that a major factor in the between-laboratory variation, especially in the region near ‘zero dose’, is sample-related, and is caused by particular samples interacting differently with different assay systems.In general, it would appear that for the well-controlled ‘ultrasensitive’ TSH immunometric assay kits, included in this study, between-laboratory agreement of estimates of the TSH concentration in serum samples is not likely to be substantially improved by use of a common matrix for the standards.  相似文献   

4.
目的建立狂犬病病毒抗体竞争ELISA检测方法,用于不同株狂犬病疫苗免疫后抗体水平的检测。方法用市售的不同狂犬病疫苗株抗原免疫NIH小鼠,制备免疫血清,用不同的疫苗株抗原包被酶标板,分别测定小鼠血清抗体效价,并通过不同株抗原抗体的交叉中和实验,分析抗原的差异性;在此基础上,将不同毒株抗原按不同比例混合包被酶标板,分别测定阳性血清样本,并与快速免疫荧光抑制试验(RFFIT)检测结果比较,确定包被抗原模式,建立狂犬病病毒抗体竞争ELISA检测方法,绘制标准曲线,确定最佳定量范围及灵敏度,确定Cut-off值;并对其特异性、精密性、准确性及稳定性进行验证;用建立的方法与其他市售狂犬病病毒抗体检测试剂分别检测血清样品,分析检测结果的一致性及相关性。结果狂犬病病毒抗原AG∶CTN-1=4∶1为最适包被抗原模式,试剂经优化后,检测抗体效价范围在535.00~33.44 mIU/ml之间,具有良好的线性关系(r>0.99),最低检出限为8.36 mIU/ml,Cut-off值为0.735 mIU。该方法检测人血清白蛋白、破伤风阳性血清、乙肝表面抗原阳性血清、白喉阳性质控血清均未发生反应;试验内变异系数在6.68%~7.84%之间;回收率在97.25%~104.50%之间;试剂置37℃3 d,与4℃保存试剂测定结果差异无统计学意义(P>0.05)。该方法检测血清样品与市售狂犬病病毒抗体检测试剂比较,均具有良好的一致性;与RFFIT测定结果比较,二者差异无统计学意义(P>0.05),回归方程为Y=-0.475+3.246 X,相关系数为0.801。结论已建立了狂犬病病毒抗体竞争ELISA检测方法,可用于大规模狂犬病病毒抗体的筛查。  相似文献   

5.
This study is aimed at improving a protocol for measuring fatty acids in cattle hair with respect to sensitivity, repeatability, and speed to increase its applicability as a biomarker. For the investigation, 14 hair samples from German Holstein cows are used. Alternative methods for grinding the hair (mortar vs mill), lipid extraction (modified Folch vs kit extraction), and solvent evaporation before injection on a gas chromatograph (evaporated vs unevaporated extracts) are tested. Hair ground with a mill compared to that with a mortar has smaller particles and a higher concentration of total lipids after extraction (p < 0.02). The kit used for lipid extraction is faster, and the amount of extracted total lipids and individual fatty acids, especially C12:0, is increased (p = 0.001). The analysis of unevaporated methyl ester extracts using gas chromatography (GC) analysis yields 5.8 and 1.3 higher amounts of C10:0 and C12:0, respectively, than those of evaporated extracts (p < 0.001). According to the results, the protocol for determining fatty acids in cattle hair can be improved by grinding the hair with a mill, extraction of lipids with a kit, and direct loading of methyl ester extracts in a gas chromatograph. Practical Applications: The fatty acid profile of hair reflects the metabolic status of an animal for the previous 1–3 weeks, as these fatty acids are not influenced by diurnal and short‐term fluctuations. An improved protocol is developed that increases the throughput of fatty acid analysis and improves its applicability for practical use. For breeding and animal welfare, the analysis of cattle hair is possible for more efficient evaluation of the hair fatty acid profile as a robust biomarker in a larger animal population.  相似文献   

6.
目的研究用化学发光免疫分析法(CLIA)检测人血清孕酮(PROG)。方法采用竞争抑制法,用高亲和力的多克隆抗体包被,碱性磷酸酶标记孕酮,金刚烷增敏化学发光体系作为酶底物来检测PROG。并考察试剂的灵敏度、精密性、准确性、特异性和测定范围。将试剂盒置4℃存放14个月,检测质控血清的PROG含量,考察试剂的稳定性。并与进口试剂进行比较。结果CLIA法检测PROG灵敏度达0.1ng/ml;特异度99.5%;测定范围在0.1~200ng/ml之间;用PROG浓度分别为高、中、低的质控血清测定精密性,批内和批间变异系数均小于10%,回收率在91.7%~108.4%之间;与雌二醇、雌三醇和雌酮的交叉反应率低于0.1%,与睾酮和可的松无交叉反应;试剂在4℃存放14个月,质控血清的测定值均在规定范围内;与进口全自动化学发光试剂BeckmanAccessTM相比有较好的相关性。二者测定结果的符合率为99.2%。结论化学发光法灵敏度高,特异性好,稳定性强,检测范围宽,有很好的准确性和精密性,是现有放射免疫法的理想替代方法。  相似文献   

7.
目的评价人巨细胞病毒(Human cytomegalovirus,HCMV)pp65 IgG-亲和力指数(Avidity index,AI)快速诊断试剂的临床应用价值。方法以自制的截短pp65重组蛋白作为诊断试剂,对本室血清库中保存的标本,采用ELISA间接法检测其特异性IgG抗体,分析该试剂的灵敏度、特异性和准确度。检测临床收集的89份患者血清标本(其中5例被明确诊断为HCMV活动性感染)的pp65 IgG抗体,并与意大利DIESSE公司IgG检测试剂盒结果进行比较。同时,对pp65 IgG阳性标本,运用尿素变性ELISA间接法检测其AI值,与意大利DIESSE公司IgM检测试剂盒结果进行比较,评价AI检测在HCMV感染诊断中的价值。结果用pp65快速诊断试剂检测阳性、阴性血清特异性IgG抗体,均与原确认结果相同,灵敏度、准确度和特异性均达100%。两种诊断试剂IgG抗体检测结果,差异无显著意义。pp65诊断试剂对pp65 IgG阳性标本的AI值进行检测,AI<60%标本共41份,阳性率为46.07%,较参比试剂IgM检测阳性率(23.59%)高;5份明确诊断HCMV活动性感染的患者血清标本中,pp65诊断试剂与参比试剂检测IgG抗体,各有4份阳性,一致率为100%,pp65诊断试剂检测AI值为阳性的标本有3份,其中,参比试剂检测IgM为阳性的仅有1份。临床明确诊断为HCMV肺炎的患者血清标本,诊断试剂检测AI值为可疑阳性,但参比试剂检测HCMV IgM抗体为阴性。结论pp65 IgG-AI快速诊断试剂可初步诊断HCMV感染,简单快速,重复性好,具有良好的临床应用前景。  相似文献   

8.
Osteoarthritis of the knee (OAK) is a chronic degenerative disease and progresses with an imbalance of cytokines and macrophages in the joint. Studies regarding the use of platelet-rich plasma (PRP) as a point-of-care treatment for OAK have reported on its effect on tissue repair and suppression of inflammation but few have reported on its effect on macrophages and macrophage polarization. Based on our clinical experience with two types of PRP kits Cellaid Serum Collection Set P type kit (leukocyte-poor-PRP) and an Autologous Protein Solution kit (APS leukocyte-rich-PRP), we investigated the concentrations of humoral factors in PRPs prepared from the two kits and the effect of humoral factors on macrophage phenotypes. We found that the concentrations of cell components and humoral factors differed between PRPs purified using the two kits; APS had a higher concentration of M1 and M2 macrophage related factors. The addition of PRP supernatants to the culture media of monocyte-derived macrophages and M1 polarized macrophages revealed that PRPs suppressed M1 macrophage polarization and promoted M2 macrophage polarization. This research is the first to report the effect of PRPs purified using commercial kits on macrophage polarization.  相似文献   

9.
目的评价HIV抗原/抗体联合检测快速法(VIDAS HIV Duo Quick)和超敏法(VIDAS HIV Duo Ultra)试剂的特异性及敏感性。方法用这2种试剂和HIV抗体参比试剂,分别检测287份HIV感染或疑似感染者样品、1240份非HIV感染者血清/血浆样品,对与参比试剂检测结果不一致样品,进行抗体的确认及HIV p24抗原和RNA的检测,并比较其特异性和敏感性。用这2种试剂和HIV-1p24抗原参比试剂,分别检测3株病毒培养上清的系列稀释样品,并比较其检测p24抗原敏感性的差异。结果共检测1277份HIV抗体阴性样品和250份HIV抗体阳性样品,与参比试剂相比,快速法试剂的特异性为99.45%,敏感性为100%,而超敏法试剂的特异性为99.14%,敏感性为99.20%。两种试剂检测病毒株培养上清的敏感性均不低于HIV-1p24抗原参比试剂,且能检出HIV感染窗口期样品。结论这2种试剂在增加HIV p24抗原检测的情况下,对HIV抗体检测的特异性和敏感性均没有明显降低,并且能检测出HIV感染窗口期样品。  相似文献   

10.
几家HCV抗体诊断试剂盒检测系列血清的比较   总被引:3,自引:0,他引:3  
应用Abbott、UBI及国内几家的HCV第二代抗体诊断试剂盒A、B、c及D检测25名HCV感染者的210份感染后不同时期的系列血清,首次采用ELISA及RIBA分片段检测抗体,并与敏感的PCR法检测血清中HCV RNA结果相比较。虽然各试剂盒的总体检测符合率无显著性差异,但个别国产试剂盒的早期检测能力尚有待提高。用任何一家抗体检测试剂盒均会漏检小部分标本,抗体检测阴性的血清在100000倍稀释后仍可检出HCV RNA,因此,有必要发展包括更多片段包被的第三代HCV抗体诊断试剂盒。  相似文献   

11.
目的应用IU标准对HBsAg检测试剂进行比较分析。方法筛选1份HBsAg阳性血浆,血清学检测证实为HBsAg adr亚型,参照WHO HBsAg标准品稀释方法进行稀释,以IU为单位的HBsAg国际标准品为标准,应用双平行线终点稀释方法对该样品进行定量标定。以标定的IU为单位样品,对国内外13家HBsAg检测试剂进行检测。结果待标定的样品经平行稀释后,测定的浓度值与理论浓度值误差均在11%以内,应用IU标准对国内外HBsAg检测试剂进行检测,国外试剂的最低检出限可达0.02~0.06IU/ml,国产试剂除1家可达0.14IU/ml外,其余试剂均在0.29~0.91IU/ml之间。结论应用IU标准检测HBsAg检测试剂,国外试剂的灵敏度明显高于国内试剂。  相似文献   

12.
目的评价结核分枝杆菌特异性IFNγ体外释放试验(interferon γ release assay,IGRA)全血检测试剂盒的质量。方法采用10个结核分枝杆菌特异性IGRA全血检测试剂盒检测参考样品,以参考样品检测值和样品浓度进行线性回归,获得线性回归方程及相关系数(R^2)。按各试剂盒说明书判定参考样品,以阳性的最低参考样品浓度作为相应试剂盒的最低检测浓度。结果 10个结核分枝杆菌特异性IGRA全血检测试剂盒的R2均高于0. 9,最低检测浓度分布在0. 4~2 IU/mL之间。结论结核分枝杆菌特异性IGRA全血检测试剂盒的质量稳定,线性分析和最低检测浓度检测可用于该试剂盒IFNγ测定部分的质量控制。  相似文献   

13.
Complex lipids including sphingolipid and plasmalogens were expected to be used as functional supplement, although their physiological activities have not been fully demonstrated. Although these complex lipids exist voluminously in brain and nervous tissues, hardly any animal resources of these lipids have been used since the outbreak of bovine spongiform encephalopathy. Thus, the chemical composition and concentration method of complex lipids from the skin of mature laying hens, a huge amount of which is wasted every year, has been investigated. Total lipid yield (32 g/100 g) prepared from chicken skin contained 2% complex lipids. Total lipids predominantly consisted of triacylglycerol (TAG), with phosphatidylcholine, sphingomyelin (SM) and phosphatidylethanolamine (PE) generally predominant as complex lipids. PE was primarily plasmalogens (62 mol%), of which arachidonic acid (47.6 mol%) and docosahexaenoic acid (11.2 mol%) were the predominant fatty acids. The component sphingoid base of sphingomyelin was almost totally 4-trans sphingenine (sphingosine). The complex lipids were able to be separated from an ethanol extract of minced skin in good yield by solvent fractionation with a hexane/ethanol system. Moreover, highly purified SM (>95 wt%) was prepared by a combination of solvent fractionation and alkaline/acidic hydrolysis from the ethanol extract. Thus, it was shown that culled chicken skin could be a potential resource of the antioxidant phospholipid plasmalogens and human-type sphingolipid.  相似文献   

14.
Fried foods are frequently served by fast food establishments but the evaluation of the oil used is quite laborious, expensive, and requires a well-structured laboratory with sophisticated equipment. Moreover, p-anisidine, used as the reagent in the traditional test for monitoring the alkenal concentration of frying oils, is carcinogenic. The DiaMed F.A.T.S. kits for the determination of alkenal (AlkalSafe™ STD) and malonaldehyde (AldelSafe™ STD), equivalent to the p-anisidine and TBA tests, respectively, are safe, fast and accurate, using compact equipment and generating fewer residues than the official methods. The results obtained using these kits were compared with those obtained using the official methods for determining alkenal (AOCS Cd 18b-90) and malonaldehyde (AOCS Cd 19b-90), in 20 samples collected from an institutional restaurant. Based on the least squares regression analysis, the AlkalSafe kit results were highly correlated with the p-anisidine values (r = 0.74), but there was a lack of correlation between the results of the AldeSafe™ kit and the TBA test. Both kits were significantly more sensitive than the official methods, as revealed by the results of the Tukey test. Although the TBA values for the samples investigated were minimal, suggesting the inadequacy of the test for monitoring frying oils, the greater sensitivity of the kit makes it a relatively feasible option.  相似文献   

15.
Extraction of lipids from biological tissues is a crucial step in lipid analysis. The selection of appropriate solvent is the most critical factor in the efficient extraction of lipids. A mixture of polar (to disrupt the protein-lipid complexes) and nonpolar (to dissolve the neutral lipids) solvents are precisely selected to extract lipids efficiently. In addition, the disintegration of complex and rigid cell-wall of plants, fungi, and microalgal cells by various mechanical, chemical, and enzymatic treatments facilitate the solvent penetration and extraction of lipids. This review discusses the chloroform/methanol-based classical lipid extraction methods and modern modifications of these methods in terms of using healthy and environmentally safe solvents and rapid single-step extraction. At the same time, some adaptations were made to recover the specific lipids. In addition, the high throughput lipid extraction methodologies used for liquid chromatography-mass spectrometry (LC-MS)-based plant and animal lipidomics were discussed. The advantages and disadvantages of various pretreatments and extraction methods were also illustrated. Moreover, the emerging green solvents-based lipid extraction method, including supercritical CO2 extraction (SCE), is also discussed.  相似文献   

16.
An optimized procedure for extraction of total and non-polar lipids from microalgae is proposed. The effects of solvent, pretreatment (lyophilization, inactivation of lipases, and addition of antioxidants) and cell-disruption (liquid nitrogen, sonication, and bead beating) on total lipid content, lipid class, and fatty acid composition were examined. Chloroform–methanol 1:1 was shown to be the best solvent mixture for extraction of total lipids from microalgae. When performing this extraction, lyophilized algae can be used, no pretreatment with isopropanol to inactivate the lipases is needed and addition of antioxidants is not necessary. Furthermore, cell-disruption is not essential, although in that case two extractions must be performed in series to ensure that, irrespective of the microalgal species, all lipids are extracted. Determination of non-polar lipid content should be performed by separation of the total lipid extract on an SPE column. Extraction using petroleum ether is only appropriate when a bead beater is used for pretreatment.  相似文献   

17.
目的建立抗戊型肝炎病毒(Anti-HEV)IgG抗体的定量线性标准品,并进行初步应用。方法利用抗-HEV IgG和抗-HEV IgM ELISA检测试剂筛选出1份抗-HEV IgG阳性血清L9,经基因1型和4型的HEV ORF2C-端抗原及239抗原进行Western blot确认后,用WHO定量标准品,由3个实验室协作标定,利用量反应平行线法计算其抗-HEV IgG的含量。考察已标定的L9血清的稳定性,并用所标定的1.5倍系列稀释的血清对国内外6家抗-HEV IgG试剂的灵敏度进行检测。选择一灵敏度较高的试剂,在其线性范围内取L9的5个稀释度作为抗-HEV IgG抗体定量线性标准,对高、中、低浓度的3份临床血清重复检测5次,考察其重复性;对实验感染猴的系列血清中抗-HEV IgG含量进行定量检测,考核该定量线性标准品的应用效果;并对每次定量试验中的线性方程进行分析,确定相关系数r值和斜率k值的范围。结果经国内外试剂检测筛选出的阳性血清L9与基因1型和4型的HEV ORF2 C-端抗原及239抗原均有阳性反应。经协作标定,L9血清抗-HEV IgG含量为16.9U/ml。L9血清在-20℃下保存6、12、18个月,2~8℃保存24、48、96h后,定量结果均在95%置信区间内,且抗-HEV IgG含量均未明显下降。6家抗-HEVIgG检测试剂灵敏度差异较大,范围为0.03~5.00U/ml。确定L9血清从0.42U/ml开始的5个1.5倍系列稀释度,作为某一试剂抗-HEVIgG抗体定量线性标准品。利用该线性定量标准检测高、中、低浓度的3份临床血清,定量结果重复性较好;对实验感染猴系列血清进行定量检测,结果可有效地反映抗体水平变化趋势;94%的定量检测试验,r≥0.98,1.15≥k≥0.95。结论已建立了抗-HEVIgG抗体定量线性标准品,可用于疫苗免疫原性评价和流行病学调查。  相似文献   

18.
检测HCV RNA的PCR试剂质控试行参考品的建立   总被引:3,自引:0,他引:3  
选慢性丙型肝炎病人中HCVRNA含量不等的阳性血清作为阳性标准,采用选自HCV基因5’端非编码区的不同引物来扩增病毒基因,扩增产物与分子量标准进行比较,部分产物用酶切初步分析进行确证。选国家HCV抗体诊断试剂盒临床验证参考品中的阴性样品作为阴性标准,经反复检测显示HCVRNA为阴性。选强阳性血清经5%小牛血清稀释作为灵敏度标准,检测范围为10-5~10-7。应用该参考品对北医肝病研究所的三批HCVRNA诊断试剂盒进行检测,阳性和阴性标准均符合要求,灵敏度为10-5~10-6。  相似文献   

19.
Singh EJ  Gershbein LL  O'neill HJ 《Lipids》1966,1(4):274-278
Techniques for the quantitative analysis of hair lipids using thin-layer chromatography (TLC) together with a proximate analysis of components in one sample deduced by these criteria are presented. Mono-, di- and triglycerides were separated by TLC using Silica Gel G as adsorbent. The chromatoplates were developed with 98% acetone+2% petroleum ether. Glycerides moved with the solvent front. The requisite portions were scraped off the plates and extracted with acetone and ether. Further TLC, limiting the migration of triglycerides and diglycerides was afforded by use of 95% ethanol as solvent in one direction while monoglycerides moved with the solvent front. For the separation of monoglycerides, chloroform was used as solvent in a second direction. Reference standards and several mixtures were run simultaneously and the spots identified by charring with concentrated sulfuric acid containing dichromate. Additional checking was effected by IR spectra. For determination of glyceride composition, methyl esters of the component fatty acids were prepared by transesterification and submitted to gas chromatography. Comparison of the levels of each of the constituent fatty acids showed no remarkable differences between the three classes of glycerides in one hair lipid pool. Although certain discrepancies in the amounts of a few fatty acid components might be construed for one pool of lipids from hair of white full-headed men (WF-9A) in contrast to findings with two Negro pools, no unequivocal conclusions can be drawn presently.  相似文献   

20.
目的对Gen-Probe公司的Procleix Ultrio(Ultrio)和Roche公司的Cobas TaqScreen MPX(MPX)HBV/HCV/HIV核酸筛查试剂的质量进行初步评价。方法从我国不同地区收集60份HIV-1感染者样品(包括59份HIV抗体阳性样品和1份HIV抗体阴性的HIV-1感染窗口期样品)及540份HIV抗体阴性样品,将60份HIV-1感染者样品随机分布于540份HIV阴性样品中,按照合并检测模式对600份样品进行检测,并对每种试剂的检测结果为阳性的样品汇集池(pool)分别按其说明书进一步进行拆分和/或鉴别试验。结果 MPX试剂和Ultrio试剂的单一人份样品检测(Individual donor test,IDT)结果一致的样品共586份,二者的符合率为97.67%,MPX试剂共有12份样品检测结果为假阳性,Ultrio试剂共有6份样品检测结果为假阳性,4份样品MPX试剂检测为阳性,而Ultrio试剂检测为阴性,其核酸含量较低(1份样品HIV-1RNA<40cp/ml,3份样品HBV DNA<12cp/ml),由于无法完成跟踪随访,因此不能确定其最终感染状态。结论 MPX试剂和Ultrio试剂检测结果具有较高的一致性,但对于HBV DNA、HCVRNA和HIV-1RNA含量较低的样品,二者检测结果可能存在一定的差异。  相似文献   

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