Quantitative detection of the tumor-associated antigen large external antigen in colorectal cancer tissues and cells using quantum dot probe

Shuo Wang, Wanming Li, Dezheng Yuan, Jindan Song, Jin Fang Department of Cell Biology, Key Laboratory of Cell Biology, Ministry of Public Health, and Key Laboratory of Medical Cell Biology, Ministry of Education, China Medical University, Shenyang, People’s Republic of China Abstract: The large external antigen (LEA) is a cell surface glycoprotein that has been proven to be highly expressed in colorectal cancer (CRC) as a tumor-associated antigen. To evaluate and validate the relationship between LEA expression and clinical characteristics of CRC with high efficiency, LEA expression levels were detected in 85 tissue blocks from CRC patients by quantum dot-based immunohistochemistry (QD-IHC) combined with imaging quantitative analysis using quantum dots with a 605 nm emission wavelength (QD605) conjugated to an ND-1 monoclonal antibody against LEA as a probe. Conventional IHC was performed in parallel for comparison. Both QD-IHC and conventional IHC showed that LEA was specifically expressed in CRC, but not in non-CRC tissues, and high LEA expression was significantly associated with a more advanced T-stage (P<0.05), indicating that LEA is likely to serve as a CRC prognostic marker. Compared with conventional IHC, receiver operating characteristic analysis revealed that QD-IHC possessed higher sensitivity, resulting in an increased positive detection rate of CRC, from 70.1% to 89.6%. In addition, a simpler operation, objective analysis of results, and excellent repeatability make QD-IHC an attractive alternative to conventional IHC in clinical practice. Furthermore, to explore whether the QD probes can be utilized to quantitatively detect living cells or single cells, quantum dot-based immunocytochemistry (QD-ICC) combined with imaging quantitative analysis was developed to evaluate LEA expression in several CRC cell lines. It was demonstrated that QD-ICC could also predict the correlation between LEA expression and the T-stage characteristics of the cell lines, which was confirmed by flow cytometry. The results of this study indicate that QD-ICC has the potential to noninvasively detect rare circulating tumor cells in clinical samples in real clinical applications. Keywords: quantum dots, large external antigen, quantum dot-based immunohistochemistry, quantitative analysis, colorectal cancer, quantum dot-based immunocytochemistr