| The global incidence of breast cancer has been rising since the late 1970s. In 2013, approximately 232,000 new cases of metastatic breast cancer (MBC) and 39,620 deaths from breast cancer were expected among U.S. women. China is not a high-incidence country, whereas breast cancer incidence is booming these years for the large population base. The growth rate of incidence of breast cancer in China is even 1-2% higher than the high-incidence countries, so the prevention and control situation is not optimistic in our country. According to the breast cancer data of 2009 published by National Cancer Center and the Department of Health Bureau of Disease Prevention and Control published in 2012:the incidence ranked first among female malignant tumor; the national prevalence was 42.55/106, cities were 51.91/106, and the countryside was 23.12/106.Mammary gland is not one of the important organs to maintain life activities. Breast cancer in situ is not life-threatening, and early stage breast cancer is even called a "curable cancer". However, in the process of the development of tumor, breast cancer cells change gradually, loosing connection between the cells, and become easy to shed. The shedding tumor cells that get into the circulation flow and get free to plant in different organizations are known as circulating tumor cells (CTCs), which are related to tumor metastases. In early stage metastasis, it is difficult to identify the metastatic sites. Therefore, CTCs can be regarded as the "liquid" biopsy of the tumor, providing convenient access to primary tumor cells, and more important, lethal metastases.ObjectiveThe aim of this study is to investigate the specificity and capture efficiency of MUC1 DNA aptamer in the detection of MCF7 cells in culture medium and artificial blood, to get the experiment parameters including the concentration of aptamer and incubation time, to confirm the affinity of MUC1 DNA aptamer to MCF7 cells, and to evaluate the feasibility using MUC1 aptamer modified chitosan/hydroxyapatite nanofilm substrates chip for the capture and detection of MBC patients’ CTCs.MethodsIn this study, we used MUC1 DNA aptamer to modify chitosan/hydroxyapatite nanofilm substrate chip after we explored the best concentration of modification. We used MUC1 DNA aptamer modified chitosan/hydroxyapatite nanofilm substrates chip to capture MCF7 cells from artificial blood, and identified captured CTCs with fluorescence microscopy (PE/FITC/DAPI-labeled).Results1. The results showed that when the concentration of MUC1 DNA aptamer was more than 5 μM, the modified chitosan/hydroxyapatite nanofilm substrates chip reached the maximal recovery yields. In addition, the maximal capture efficiency was achieved at incubation times of 45minutes or longer.2. The results showed that capture efficiency of MCF7 cell line was more than 60% using MUC1 DNA aptamer modified chitosan/hydroxyapatite nanofilm substrates chip, and the MCF7 cells were captured from artificial blood with a yield of 58.68-63%.3. The results demonstrated that MUC1 DNA aptamer had high specificity and affinity to MCF7 cells, so that made it able to replace anti-EpCAM (the traditional antibody for CTCs capture). Furthermore, MUC1 DNA aptamer modified chitosan/hydroxyapatite nanofilm substrates chip seemed potential in detection of breast cancer CTCs.ConclusionWe have demonstrated that MUC1 DNA aptamer is capable to replace traditional antibodies for breast cancer CTCs capture. We provided a new method of the detection of breast cancer CTCs and the early stage diagnosis of MBC using modified chitosan/hydroxyapatite nanofilm substrates chip. |
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