Enzyme activity in dried blood spot as a diagnostic tool for adenosine deaminase 2 deficiency
Enzyme activity in dried blood spot as a diagnostic tool for adenosine deaminase 2 deficiency
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DOI：
10.1016/j.ab.2021.114292
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发表时间：
2021-06-28
影响因子：
2.9
通讯作者：
Iwaki-Egawa, Sachiko
中科院分区：
文献类型：
--
作者：
Ito, Moeko;Nihira, Hiroshi;Iwaki-Egawa, Sachiko
背景：腺苷脱氨酶 2 (DADA2) 缺乏症是一种由腺苷脱氨酶 2 (ADA2) 基因突变引起的自身炎症性疾病。功能性 ADA2 活性的丧失会导致血管炎综合征、免疫缺陷和造血障碍。需要早期诊断才能有效治疗。方法：我们开发了一种基于干血斑 (DBS) 的 ADA2 活性比色测定法。在样品制备过程中使用肝素亲和纯化以更有效地改进测定。检查了 DBS 储存期间 ADA2 的稳定性以及 DADA2 患者和健康对照的 ADA2 活性。结果：从健康对照的 DBS 中提取出活性 ADA2。 DBS 中的 ADA2 活性无论冷冻或冷藏保存，均可保持稳定至少 90 天。在健康对照和患者之间观察到 ADA2 活性存在显着差异。在患者的 DBS 中未检测到 ADA2 活性。结论：我们的新 DBS ADA2 活性测定实验简单，适应性强，除了酶标仪外不需要特殊设备。通过肝素亲和纯化实现了低背景。该方法清楚地区分健康对照和患者。可以在 DBS 中可靠地测量 ADA2 活性，从而提供早期诊断 DADA2 的机会。
Background: Deficiency of adenosine deaminase 2 (DADA2) is an autoinflammatory disease caused by mutations in the adenosine deaminase 2 (ADA2) gene. Loss of functional ADA2 activity results in vasculitis syndrome, immunodeficiency, and hematopoietic disorders. Early diagnosis is required for effective treatment. Methods: We developed a dried blood spot (DBS)-based ADA2 activity colorimetric assay. Heparin-affinity purification was used during sample preparation to improve the assay more efficiently. The stability of ADA2 during DBS storage and ADA2 activity of DADA2 patients and healthy controls were examined. Results: Active ADA2 was extracted from the DBS of healthy controls. ADA2 activity in DBS, stored either frozen or refrigerated, remained stable for at least 90 days. A significant difference in ADA2 activity was observed between healthy controls and patients. No ADA2 activity was detected in DBS from patients. Conclusions: Our new DBS ADA2 activity assay is experimentally simple, highly adaptable, and requires no special equipment except for a microplate reader. A low background was achieved with heparin-affinity purification. The method differentiates clearly between healthy controls and patients. ADA2 activity can be reliably measured in DBS, providing an opportunity to diagnose DADA2 at an early stage.