Cancer Genetics and Epigenetics 2024, Vol.12, No.5, 254-269 http://medscipublisher.com/index.php/cge 259 4.2 Methods for CTC isolation 4.2.1 Density gradient centrifugation Density gradient centrifugation is a widely used method for the isolation of circulating tumor cells (CTCs) based on their density differences compared to other blood components(Fig.2). This technique involves layering a blood sample over a density gradient medium and centrifuging it to separate cells based on their buoyant densities. A study demonstrated the efficacy of this method by combining it with microfiltration, achieving high purity and recovery rates of CTCs from whole blood samples (Park et al., 2012). The selective sedimentation of CTCs within a density gradient medium effectively removes unwanted hematologic cells, yielding a much purer sample compared to other filter-based methods (Park et al., 2012). 4.2.2 Immunomagnetic separation Immunomagnetic separation leverages the specific binding of antibodies to surface markers on CTCs, allowing for their isolation using magnetic beads. This method can be highly specific and efficient, especially when combined with other techniques. For instance, a study proposed a two-step process combining immunomagnetic bead-based cell isolation with optically induced dielectrophoresis (ODEP) to enhance the purity of isolated CTCs (Chu et al., 2020). Another study developed an immunomagnetic platform that uses dual aptamers targeting different CTC phenotypes, achieving high capture efficiency and enabling detailed analysis of CTC subpopulations in colorectal cancer patients (Li et al., 2022). 4.2.3 Size-based filtration Size-based filtration exploits the size differences between CTCs and other blood cells to isolate CTCs. This method is label-free, simple, and fast, making it a popular choice for CTC isolation. A comprehensive review highlighted the advantages of size-based enrichment, noting that it preserves the viability and unmodified state of CTCs for subsequent analyses (Hao et al., 2018). However, the variability in CTC sizes can pose challenges, as some CTCs may overlap in size with leukocytes, potentially reducing the method's reliability (Park et al., 2012). Despite these challenges, size-based filtration remains a valuable tool for CTC isolation due to its simplicity and efficiency (Park et al., 2012; Hao et al., 2018). 4.2.4 Combined approaches Combining multiple isolation techniques can enhance the efficiency and purity of CTC isolation. For example, a study described a novel method that integrates density gradient centrifugation with microfiltration, achieving high sensitivity and selectivity in CTC isolation (Park et al., 2012). Another approach combined immunomagnetic separation with ODEP cell manipulation, significantly improving the purity of isolated CTCs (Chu et al., 2020). These combined methods leverage the strengths of individual techniques, providing a more robust and reliable means of isolating CTCs for clinical and research applications. 5 CTCs as Biomarkers for Colon Cancer 5.1 Diagnostic value of CTCs in early detection Circulating tumor cells (CTCs) have emerged as a promising biomarker for the non-invasive diagnosis and management of colorectal cancer (CRC) (Table 1). These cells, shed from primary tumors into the bloodstream, offer a minimally invasive means to detect and monitor cancer progression. The clinical utility of CTCs spans diagnostic, prognostic, and predictive applications, making them a multifunctional biomarker in cancer medicine (Yap et al., 2014). The diagnostic potential of CTCs in early-stage colorectal cancer is significant. Studies have shown that CTCs can be detected even in early stages of the disease, providing a valuable tool for early diagnosis. For instance, the detection of CTCs in peripheral blood has been associated with early-stage hepatocellular carcinoma, suggesting a similar potential in colorectal cancer (Qi et al.,2018). Furthermore, the analysis of circulating tumor DNA (ctDNA) and CTCs has shown promise in improving early detection strategies for CRC, complementing traditional screening methods (Marcuello et al., 2019).
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