Department of Logistics Management, Baoshan Branch, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200444, China.
Department of Medical Intensive Care Unit, Baoshan Branch, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200444, China.
BMC Biotechnol. 2022 Sep 30;22(1):27. doi: 10.1186/s12896-022-00758-2.
Lymphocytic leukemia (LL) is a primary malignant tumor of hematopoietic tissue, which seriously affects the health of children and the elderly. The study aims to establish a new detection method for screening acute/chronic LL using time-resolved fluorescence immunoassay (TRFIA) via quantitative detection of S100 calcium binding protein A8 (S100A8) and leucine-rich alpha-2-glycoprotein 1 (LRG1) in serum.
Here a sandwich TRFIA was optimized and established: Anti-S100A8/LRG1 caputre antibodies immobilized on 96-well plates captured S100A8/LRG1, and then banded together with the anti-S100A8/LRG1 detection antibodies labeled with Europium(III) (Eu)/samarium(III) (Sm) chelates. Finally time resolved fluorometry measured the fluorescence intensity.
The sensitivity of S100A8 was 1.15 ng/mL(LogY = 3.4027 + 0.4091 × LogX, R = 0.9828, P < 0.001, dynamic range: 2.1-10,000 ng/mL), and 3.2 ng/mL for LRG1 (LogY = 3.3009 + 0.4082 × LogX, R = 0.9748, P < 0.001, dynamic range: 4.0-10,000 ng/mL). The intra-assay and inter-assay CVs were low, ranging from 5.75% to 8.23% for S100A8 and 5.30% to 9.45% for LRG1 with high specificity and affinity in serum samples. Bland-Altman plots indicated TRFIA and ELISA kits have good agreement in clinical serum samples. Additionally, the cutoff values for S100A8 and LRG1 were 1849.18 ng/mL and 588.08 ng/mL, respectively.
The present TRFIA method could be used for the quantitative detection of S100A8 and LRG1 in serum, and it has high sensitivity, accuracy and specificity. Clinically, this TRFIA method could be suitable for screening of LL via the quantitative detection of S100A8 and LRG1.
淋巴细胞白血病（LL）是一种原发性造血组织恶性肿瘤，严重影响儿童和老年人的健康。本研究旨在通过定量检测血清中 S100 钙结合蛋白 A8（S100A8）和富含亮氨酸α-2-糖蛋白 1（LRG1），建立一种新的用于筛选急性/慢性 LL 的时间分辨荧光免疫分析（TRFIA）检测方法。
本研究优化并建立了一种夹心 TRFIA 方法：将 S100A8/LRG1 捕获抗体固定在 96 孔板上，捕获 S100A8/LRG1，然后与用铕（III）（Eu）/钐（III）（Sm）螯合物标记的抗 S100A8/LRG1 检测抗体结合。最后，时间分辨荧光法测量荧光强度。
S100A8 的检测灵敏度为 1.15ng/mL（LogY=3.4027+0.4091×LogX，R=0.9828，P<0.001，动态范围：2.1-10000ng/mL），LRG1 的检测灵敏度为 3.2ng/mL（LogY=3.3009+0.4082×LogX，R=0.9748，P<0.001，动态范围：4.0-10000ng/mL）。S100A8 的批内和批间 CV 较低，范围为 5.75%-8.23%，LRG1 的批内和批间 CV 较低，范围为 5.30%-9.45%，在血清样本中具有高特异性和亲和力。Bland-Altman 图表明，TRFIA 和 ELISA 试剂盒在临床血清样本中具有良好的一致性。此外，S100A8 和 LRG1 的截断值分别为 1849.18ng/mL 和 588.08ng/mL。
本研究建立的 TRFIA 方法可用于血清中 S100A8 和 LRG1 的定量检测，具有较高的灵敏度、准确性和特异性。临床上，该 TRFIA 方法可通过定量检测 S100A8 和 LRG1 用于 LL 的筛选。
