BackgroundCirculating tumor cells (CTCs) are solid tumor epithelial cells that shed into the blood, which are highly correlated with tumor metastasis, drug resistance, prognosis, and so on. Most circulating tumor cells express epithelial cell adhesion molecules (EPCam) on the surface, so some methods use EPCAM to capture CTCs directly. However, a large number of studies in recent years have proved that the expression of epCAM is quite heterogeneous, and the expression of epCAM in cells from different tissues**, and even cells within the same tumor tissue, is very different before and after, which makes the method of capturing CTCs relying on EPCAM has great uncertainty or even complete non-captureability. Subtraction Enrichment (SE) technology is completely independent of the expression of tumor cell surface markers (e.g., EPCAM), and uses immunomagnetic particles conjugated to a special monoclonal antibody panel against human leukocyte surface antigen and a special centrifugation medium for isolating non-bloodborne cells to quickly and efficiently remove red blood cells, leukocytes and plasma proteins from the blood of tumor patients, thereby effectively enriching circulating rare cells in peripheral blood specimens of patients, including** Circulating tumor cells, circulating vascular endothelial cells, stem cells, etc. from solid tumors of all different tissues. The obtained tumor cells are neither encapsulated by magnetic beads nor bound to any antibodies against tumor cell surface markers (e.g., eEPCAM), thus avoiding activation of intracellular signaling pathways. Different from other conventional methods, the removal of red blood cells in this experiment avoids the use of hypotonic lysis method, so as to minimize the damage to the enriched tumor cells, so that the obtained target cells maintain a good natural state and cell morphology, which can be applied to a series of subsequent morphological and functional studies. Such as immunofluorescence staining-chromosome fluorescence in situ hybridization (IFISH), protein expression, etc., and genetic analysis of single tumor cells obtained with the help of microdissection technology.
Clinical significance of CTC-iFISH assay in tumor body fluid biopsyImmunofluorescence staining - chromosomal fluorescence in situ hybridization (IFISH).On the basis of distinguishing between bloodborne and non-bloodborne cells, immunofluorescence staining and chromosomal fluorescence in situ hybridization technology were successfully integrated into the simultaneous implementation of the immunofluorescence staining-chromosomal fluorescence in situ hybridization technology platform (TM-ifish), which can effectively distinguish circulating vascular endothelial cells (CECs) and circulating tumor cells (CTCs) while distinguishing bloodborne cells, thus greatly increasing the number of blood cells Specificity of CTC assays. CTC cells detected by the IFISH method can be used for subsequent single-cell or multicellular studies of a range of genes, proteins, or other functions of interest.
CTC-iFISH tumor liquid biopsy has different clinical significance at different stages:(1) Early diagnosis, auxiliary diagnosis, and baseline prognosis;(2) Neoadjuvant chemotherapy;(3) Dynamic changes in CTC before and after surgery and **;(4) Detect the ** condition of MRD, etc.
Practical application of CTC-iFISH detection in Meijie Medicine
In the years of CTC-IFISH practice, Mindjet Medical Pathology Center has accumulated rich experience in clinical testing and scientific research. So far, a variety of tumor marker proteins can be detected, including: pan-cancer epcam, CK, vimentin, etc.;Tumor-specific proteins: PSMA, CA199, CA125, etc.;Stem cell proteins: CD44, CD133, etc.;**Target proteins: HER2, PD-L1, etc. Equipped with CytoIntelligen's CTC phase enrichment and iFISH detection kits, Maijie Medical can conduct targeted data and module analysis, and comprehensively help the clinical testing and scientific research work of many customers. 
Figure 1: In situ detection of abnormal chromosome number and expression of HER2 and PD-L1 tumor markers on the same CTC cell.
Figure 2: Example of MindJet CTC-iFISH staining** with a total of 20 circulating tumor cells and 0 circulating tumor cell clumps detected.
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