Automated assay of plasma (1*3) -beta-D-glucan for early diagnosis of deep mycosis

血浆 (1*3)-β-D-葡聚糖自动检测用于早期诊断深部真菌病

• 批准号: 06672300
• 负责人: OBAYASHI Taminori
• 金额: $ 1.28万
• 依托单位: Jichi Medical School (JMS)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06672300/
• 关键词: deep mycosis; (1*3) -beta-D-glucan; triple helix; single strand; automation; horseshoe crab; (1*3) -beta-D-glucan binding protein; Factor G; カブトガニ血球抽出液; 抗(1-3)-β-D-グルカン抗体; ELISA; 三重鎖螺旋構造; カイネティック・モード

Automatic assay of plasma (1*3) -beta-D-glucan with limulus reagent has been materialized with a new pretreating mixture which obviates the necessity of centrifugation. The mixture consists of KOH,KCL,and polybrene. KOH is to remove limulus-test interfering factors and enhance the sensitivity of the test by dissembling the triple helix of the glucan into single strands, which are a more potent activator of factor G.KCL is to keep the dissembled glucan in single strands, and polybrene is to prevent the formation of nonspecific turbidity. The adoption of kinetic mode has broadened the dynamic range up to 500 pg/ml. With a cutoff of 20 pg/ml, the sensitivity and the specificity of the test for invasive deep mycoses were 90% and 100%, respectively.Recently a new protein was isolated from horseshoe crab hemolymph that specifically binds to (1*3) -beta-D-glucan, thereby inhibiting the activation of factor G by the glucan. Using this new protein and anti- (1*3) -beta-D-glucan antibody, we developed an ELISA for (1*3) -beta-D-glucan determination. Plasma glucan concentrations in invasive deep mycoses showed a good correlation between the two methods, another evidence that the glucan is increased in blood in this conditon.

采用鲎试剂自动测定血浆（1*3）-β-D-葡聚糖已通过一种新的预处理混合物实现，从而消除了离心的必要性。该混合物由 KOH、KCL 和聚凝胺组成。 KOH是通过将葡聚糖的三螺旋分解成单链来去除鲎试验的干扰因素，提高测试的灵敏度，这是更有效的G因子激活剂。KCL是使分解后的葡聚糖保持单链，聚凝胺是防止非特异性浊度的形成。动力学模式的采用将动态范围扩大至 500 pg/ml。以 20 pg/ml 为临界值，该检测对侵袭性深部真菌病的敏感性和特异性分别为 90% 和 100%。 最近从鲎血淋巴中分离出一种新蛋白质，该蛋白质特异性结合 (1*3) -β-D-葡聚糖，从而抑制葡聚糖对 G 因子的激活。使用这种新蛋白质和抗 (1*3) -β-D-葡聚糖抗体，我们开发了用于 (1*3) -β-D-葡聚糖测定的 ELISA。侵袭性深部真菌病中的血浆葡聚糖浓度显示两种方法之间具有良好的相关性，这是在这种情况下血液中葡聚糖增加的另一个证据。

项目成果

Obayashi,T: "Plasma (1→3)-β-D-glucan determination in the diagnosis of invasive deep mycosis and fungal febrile episodes" Lancet. 345. 17-20 (1995)

Obayashi, T：“血浆 (1→3)-β-D-葡聚糖测定在诊断侵袭性深部真菌病和真菌发热发作中的应用”《柳叶刀》345. 17-20 (1995)。

Tamura, H: "Automated kinetic assay for endotoxin and (1*3) -beta-D-glucan in human blood." Clin Chim Acta. 226. 109-112 (1994)

Tamura, H：“人血液中内毒素和 (1*3)-β-D-葡聚糖的自动动力学测定。”

Tamura, H: "Plasma (1*3) -beta-D-glucan assay and immunohistochemical staining of (1*3) -beta-D-glucan in fungal cell walls." J Clin Lab Anal. 11 (in press).

Tamura, H：“真菌细胞壁中 (1*3) -β-D-葡聚糖的血浆 (1*3) -β-D-葡聚糖测定和免疫组织化学染色。”

Tamura,H: "Plasma (1→3)-β-D-glucan assay and immunohistochemical Staining of (1→3)-β-D-glucan in fungal cell walls" Journal of Clinical Laboratory Analysis. 11(印刷中). (1997)

Tamura, H：“真菌细胞壁中 (1→3)-β-D-葡聚糖测定和 (1→3)-β-D-葡聚糖的免疫组织化学染色”《临床实验室分析杂志》11（出版中）。 (1997)

Tamura,H: "Plasma (1→3)-β-D-glucan assay and immuns histochemical staining of (1→3)-β-D-glucan in fungal cell walls" J Clin Lab Aual. 11(印刷中). (1197)

Tamura, H：“真菌细胞壁中 (1→3)-β-D-葡聚糖的血浆 (1→3)-β-D-葡聚糖测定和免疫组织化学染色”J Clin Lab Aual 11（出版中）。 (1197)