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中国癌症防治杂志

中国癌症防治杂志 ›› 2013, Vol. 5 ›› Issue (2): 117-121.doi: 10.3969/j.issn.1674-5671.2013.02.08

• 基础研究 • 上一篇    下一篇

液相芯片技术定量检测AFP方法的研究

  

  1. 广西医科大学公共卫生学院环境卫生教研室；广西医科大学医学科学实验中心
  • 出版日期:2013-06-25 发布日期:2013-06-27
  • 通讯作者: 何 敏 E-mail:m_h_m868@sina.com
  • 基金资助:

    广西医科大学医学科学实验中心开放基金资助项目（KFJJ 2010-45）

Quantitation of alpha fetoprotein （AFP） in human serum using liquid chip technology

  • Online:2013-06-25 Published:2013-06-27

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摘要： 目的 建立液相芯片技术检测血清中AFP的反应体系，并对该法进行评价。方法 采用双抗体夹心液相芯片技术，分别以不同偶联量的抗-AFP单抗与荧光微球偶联，以最适偶联量的荧光微球与不同浓度的生物素-羊抗鼠IgG反应，确定检测方法的线性范围、最低检测限、精密度等指标；应用液相芯片技术检测134例肝癌患者和47例健康对照者血清中AFP的浓度，并将该法与电化学发光免疫分析法（ECLIA）进行相关性分析。结果 抗-AFP单抗的偶联最适量为8 μg，生物素-羊抗鼠IgG的最适浓度为4 μg/ml，检测AFP的线性范围为1.31～168 ng/ml，最低检测限为0.424 ng/ml，批内精密度为4.33%～7.66%，批间精密度为9.82%～13.85%。用液相芯片技术定量检测血清中AFP的浓度，各组差异有统计学意义（P<0.05）。液相芯片技术检测血清中AFP的灵敏度为85.82%，特异度为95.74%，该法与ECLIA法检测的相关系数r = 0.845（P<0.05）。结论 液相芯片技术检测血清中AFP与ECLIA法检测AFP有较好的相关性，并具有高通量、线性范围宽、灵敏度高、重复性好、节省样品和时间等优点，该法的建立将为进一步开发AFP联合检测多种肝癌标志物奠定基础。

关键词: 液相芯片, 肝癌标志物, AFP, 方法

Abstract: Objective To develop liquid chip-based method for assaying alpha fetoprotein（AFP）in human serum and to evaluate method performance． Methods Different amounts of anti-AFP monoclonal antibody were coupled to fluorescent beads using liquid chip technology in a sandwich immunoassay format. Fluorescent microspheres prepared with the optimal amount of anti-AFP antibody were reacted with different concentrations of biotin-goat anti-mouse IgG，and several performance indicators were measured，including linear range，detection limit，and accuracy.The method was used to measure serum AFP concentration in 134 patients with hepatocellular carcinoma（HCC） and 47 healthy controls，and the results were compared with those obtained by electrochemoluminescence immunoassay（ECLIA）. Results Optimal coupling was obtained with 8 μg of anti-AFP monoclonal antibody and 4 μg/ml of biotin-goat anti-mouse IgG.The linear response range for the method to detect serum AFP was 1.31~168 ng/ml and the detection limit was 0.424 ng/ml. Intra- and inter-assay coefficients of variation ranged，respectively，from 4.33% to 7.66% and from 9.82% to 13.85%.The method detected significantly diffe-rent serum AFP levels between patients with HCC and healthy controls（P<0.05）.Method sensitivity was 85.82% and specificity was 95.74%.Measurements obtained by the liquid chip method showed a correlation coefficient（r） of 0.845（P<0.05） with measurements obtained by ECLIA. Conclusion The liquid chip and ECLIA methods showed good correlation in detecting AFP.Since the liquid chip method offers a wider detection range and greater sensitivity than ECLIA，in addition to showing good repeatability，it may allow faster detection with fewer samples.This approach may also prove useful for detecting AFP together with other multiplexing markers in HCC.

Key words: Liquid chip, Tumor marker, AFP, Method