Assessment of three variations of the 1,9-dimethylmethylene blue assay for measurement of sulfated glycosaminoglycan concentrations in equine synovial fluid
Assessment of three variations of the 1,9-dimethylmethylene blue assay for measurement of sulfated glycosaminoglycan concentrations in equine synovial fluid
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DOI：
10.2460/ajvr.2003.64.900
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发表时间：
2003-07-01
影响因子：
1
通讯作者：
Lumsden, JH
中科院分区：
文献类型：
--
作者：
Oke, SL;Hurtig, MB;Lumsden, JH
目的：确定3种不同的1,9-二甲基亚甲基蓝（DMMB）检测方法在测定马滑液（SF）中硫酸糖胺聚糖（sGAG）浓度时是否具有可比性。样本群-从13匹马的患病关节中采集了25个SF样本，从4匹马的非患病（对照）关节中采集了13个SF样本。通过直接分光光度法（即Farndale法）、微孔板法和间接DMMB法测定从正常和病变关节收集的SF样品以及用核酸酶、木瓜蛋白酶和透明质酸酶消化的样品中的硫酸糖胺聚糖浓度。结果：所有3种检测方法与sGAGs标准溶液反应相似，并与核酸酶、木瓜蛋白酶和透明质酸酶消化SF样品。核酸不是重要的干扰物质，木瓜蛋白酶和透明质酸酶不能交替使用来消化SF。3种方法均具有满意的精密度（SD < 10%），但每种DMMB方法对马SF中sGAG的测定结果存在显著差异。结论及临床意义：在DMMB测定前，SF样品应先用木瓜蛋白酶或透明质酸酶消化。当使用不同的DMMB测定法时，研究人员目前无法比较临床信息，因为每种DMMB测定法在SF中产生的sGAG浓度有很大的不同。在这里检查的3种测定法中，我们建议使用直接分光光度法DMMB测定法。
Objective-To determine whether 3 variations of the 1,9-dimethylmethylene blue (DMMB) assay yield comparable results when measuring sulfated glycosaminoglycan (sGAG) concentrations in equine synovial fluid (SF).Sample Population-25 samples of SF collected from affected joints of 13 horses and 13 samples of SF collected from nonaffected (control) joints of 4 horses.Procedure Sulfated glycosaminoglycan concentrations were measured by the direct spectrophotometric (ie, Farndale), microplate, and indirect DMMB assays in samples of SF collected from normal and affected joints and in samples digested with nucleases, papain, and hyaluronidase.Results-All 3 assays reacted similarly to standard solutions of sGAGs and digestion of SF samples with nucleases, papain, and hyaluronidase. Nucleic acids were not important interfering substances, and papain and hyaluronidase could not be used interchangeably to digest SF All 3 assays proved to have satisfactory precision (SD < 10%), but each DMMB assay resulted in significantly different measures of sGAG in equine SF.Conclusions and Clinical Relevance-Samples of SF should be digested with papain or hyaluronidase prior to measurement via DMMB assay. Researchers currently are unable to compare clinical information when variations of the DMMB assay are used, because each DMMB assay yields substantially different sGAG concentrations in SF Of the 3 assays examined here, we recommend use of the direct spectrophotometric DMMB assay.