Kang Keren, Wu Peidian, Li Wenmei, Tang Shixing, Wang Jihua, Luo Xiaochun, Xie Mingquan
a School of Bioscience and Bioengineering, South China University of Technology, Guangzhou , Guangdong Province , China.
b National Engineering Laboratory of Point-of-Care Testing, Guangzhou Wondfo Biotech Co. Ltd , Guangzhou, Guangdong Province , China.
Scand J Clin Lab Invest. 2015;75(8):693-8. doi: 10.3109/00365513.2015.1054870.
To develop a rapid, sensitive and specific assay for quantification of serum heart-type fatty acid binding protein (H-FABP) based on immunofluorescence of specific monoclonal antibodies.
We generated novel H-FABP-directed monoclonal antibodies by cloning of spleen cells of mice immunized with H-FABP. Epitopes were mapped and antigen affinity was assessed by surface plasmon resonance (SPR). The H-FABP specific monoclonal antibodies were coupled to fluorescent beads and sprayed onto a nitrocellulose membrane facilitating quantification of H-FABP by immunofluorescence. Reagent cross-reactivity, interference resistance, accuracy and sensitivity were examined. A total of 103 clinical samples were used to compare the sensitivity and specificity of the new assay to a commercially available Randox kit.
This new assay could be finished within 15 min, with sensitivity reaching 1 ng/ml. In a trial of 103 clinical serum samples, the new testing kit results were highly correlated with those from the Randox kit (R(2) = 0.9707). Using the Randox kit as the reference kit, the sensitivity of the new assay was 98.25%, and specificity was 100%.
An immunofluorescence-based H-FABP assay employing novel monoclonal antibodies could rapidly, specifically and sensitively detect H-FABP in serum samples, providing an effective method for rapid clinical assessment of H-FABP index in the clinic.
基于特异性单克隆抗体的免疫荧光技术，开发一种快速、灵敏且特异的血清心脏型脂肪酸结合蛋白（H-FABP）定量检测方法。
我们通过克隆用H-FABP免疫的小鼠脾细胞，制备了新型的针对H-FABP的单克隆抗体。通过表面等离子体共振（SPR）技术绘制表位图谱并评估抗原亲和力。将H-FABP特异性单克隆抗体偶联到荧光微球上，并喷涂在硝酸纤维素膜上，以便通过免疫荧光对H-FABP进行定量分析。检测试剂的交叉反应性、抗干扰性、准确性和灵敏度。共使用103份临床样本，比较新检测方法与市售朗道（Randox）试剂盒的灵敏度和特异性。
这种新检测方法可在15分钟内完成，灵敏度达到1 ng/ml。在对103份临床血清样本的检测中，新检测试剂盒的结果与朗道试剂盒的结果高度相关（R² = 0.9707）。以朗道试剂盒作为参考试剂盒，新检测方法的灵敏度为98.25%，特异性为100%。
采用新型单克隆抗体的基于免疫荧光的H-FABP检测方法能够快速、特异且灵敏地检测血清样本中的H-FABP，为临床快速评估H-FABP指标提供了一种有效的方法。
