Yang Guangtian, Li Jishun, Zhang Shenglan, Ouyang Huixiang, Jiang Chunhai, Pan Hongcheng
Guangxi Key Laboratory of Electrochemical and Magneto-chemical Functional Materials, College of Chemistry and Bioengineering, Guilin University of Technology, Guilin 541004, China.
Guangxi Key Laboratory of Electrochemical and Magneto-chemical Functional Materials, College of Chemistry and Bioengineering, Guilin University of Technology, Guilin 541004, China.
J Immunol Methods. 2023 Dec;523:113574. doi: 10.1016/j.jim.2023.113574. Epub 2023 Oct 24.
Serum amyloid A (SAA) is an acute-phase protein produced in response to inflammatory proteins during infections, inflammation, trauma, surgery, cancer, and other conditions. Early and accurate detection of SAA is necessary for diagnosis and monitoring of disease progression. To meet this need, we developed a gradient lateral flow immunoassay test strip using Au nanoparticles as signal reporters. The test strip has three test (T1, T2, and T3) lines with progressively decreasing concentrations of SAA antibody, enabling the determination of high, medium, and low concentrations of SAA in serum. The test strip results were analyzed using three distinct readout methods, each with different sensitivity, accuracy, and precision for SAA concentration measurements. Qualitative judgment is based on the color of the T1 line. Semi-quantitative assessment of SAA concentration is determined by the number of colored T-lines. Specifically, color development in T1 line alone indicates a concentration range of 10-50 μg/mL, while T1 and T2 lines together indicate a range of 50-100 μg/mL, and development in all three lines (T1, T2, and T3) indicates a concentration of >100 μg/mL. Quantitative analysis was performed using either smartphone imaging or image scanning with ImageJ software. By using a five-parameter logistic function, we found a strong correlation (R = 0.998) between the ratio of signal intensities of (T1 + T2 + T3) to the control (C) line and SAA concentrations ranging from 5 to 1000 μg/mL. At lower concentrations (0-100 μg/mL), we observed a proportional relationship between the value of (T1 + T2 + T3)/C and SAA concentration. The limit of detection for SAA was 9.33 ng/mL (or 6.53 μg/mL of SAA in undiluted serum samples) for the smartphone method and 3.06 ng/mL (or 2.14 μg/mL of SAA in undiluted serum samples) for the scanner method. The gradient test strip was highly consistent with a commercially available SAA immunochromatographic test strip when tested with real human serum samples. Passing-Bablok regression indicated that results obtained using the smartphone app and scanner methods of the gradient test strip were comparable to those obtained using the commercial test strip. The gradient test strip is flexible and adaptable, providing solutions for qualitative, semi-quantitative, and quantitative SAA measurements.
血清淀粉样蛋白A（SAA）是一种急性期蛋白，在感染、炎症、创伤、手术、癌症及其他病症期间，机体对炎性蛋白作出反应时产生。早期准确检测SAA对于疾病诊断和病情进展监测至关重要。为满足这一需求，我们研发了一种以金纳米颗粒作为信号报告分子的梯度侧向流动免疫分析试纸条。该试纸条有三条检测线（T1、T2和T3），其上SAA抗体浓度逐渐降低，能够测定血清中高、中、低浓度的SAA。试纸条检测结果采用三种不同的读数方法进行分析，每种方法在测量SAA浓度时具有不同的灵敏度、准确性和精密度。定性判断基于T1线的颜色。SAA浓度的半定量评估由显色的T线数量决定。具体而言，仅T1线显色表明浓度范围为10 - 50μg/mL，T1线和T2线同时显色表明浓度范围为50 - 100μg/mL，三条线（T1、T2和T3）均显色表明浓度>100μg/mL。定量分析使用智能手机成像或用ImageJ软件进行图像扫描。通过使用五参数逻辑函数，我们发现（T1 + T2 + T3）与对照（C）线的信号强度比值与5至1000μg/mL的SAA浓度之间存在强相关性（R = 0.998）。在较低浓度（0 - 100μg/mL）时，我们观察到（T1 + T2 + T3）/C值与SAA浓度之间存在比例关系。智能手机方法检测SAA的检测限为9.33ng/mL（或未稀释血清样本中6.53μg/mL的SAA），扫描仪方法的检测限为3.06ng/mL（或未稀释血清样本中2.14μg/mL的SAA）。当用真实人类血清样本进行测试时，梯度试纸条与市售SAA免疫层析试纸条高度一致。Passing - Bablok回归表明，使用梯度试纸条的智能手机应用程序和扫描仪方法获得的结果与使用商业试纸条获得的结果相当。梯度试纸条灵活且适应性强，为SAA的定性、半定量和定量测量提供了解决方案。
