Xiao Yan, Gao Xiugong, Gannot Gallya, Emmert-Buck Michael R, Srivastava Sudhir, Wagner Paul D, Amos Michael D, Barker Peter E
NIST-NCI EDRN Cancer Biomarker Reference Laboratory, National Institute of Standards and Technology, Gaithersburg, MD, USA.
Int J Cancer. 2008 May 15;122(10):2178-86. doi: 10.1002/ijc.23320.
In an effort to improve affinity biomarker validation in fixed patient tissue specimens, we have developed a novel quantum dot-based bioimaging system that utilizes chicken IgY antibody for high sensitivity and specificity relative quantitation of cancer proteins. Monospecific, polyclonal IgYs were generated against human HER2 and telomerase, and analytically validated for specificity by western blot and immunohistochemistry on tumor and normal cells and for relative affinity by layered peptide array (LPA). IgYs bound desired targets in cell lines and fixed tissues and showed greater affinity than commercial mammalian antibodies for both HER2 and telomerase proteins. In tissue microarray experiments, HER2 quantitation with IgY antibody and quantum dot imaging correlated well with chromogenic in situ hybridization (CISH), whereas telomerase quantitation suggested a trend toward correlation with prostate cancer Gleason Grade and differentiation. Although patient numbers were small, these findings demonstrate the feasibility of relative quantitation of cancer biomarkers with IgY and quantum dot fluorophores, and show promise for rigorous clinical validation in large patient cohorts.
为了改进固定患者组织样本中的亲和生物标志物验证，我们开发了一种基于量子点的新型生物成像系统，该系统利用鸡IgY抗体对癌症蛋白进行高灵敏度和特异性的相对定量。针对人HER2和端粒酶产生了单特异性多克隆IgY，并通过蛋白质印迹法和免疫组织化学法在肿瘤细胞和正常细胞上对其特异性进行了分析验证，并通过层状肽阵列（LPA）对其相对亲和力进行了验证。IgY在细胞系和固定组织中与所需靶标结合，并且对HER2和端粒酶蛋白均显示出比商业哺乳动物抗体更高的亲和力。在组织微阵列实验中，用IgY抗体和量子点成像进行的HER2定量与显色原位杂交（CISH）相关性良好，而端粒酶定量显示出与前列腺癌Gleason分级和分化相关的趋势。尽管患者数量较少，但这些发现证明了用IgY和量子点荧光团对癌症生物标志物进行相对定量的可行性，并显示出在大型患者队列中进行严格临床验证的前景。
