Detecting cell-in-cell structures in human tumor samples by E-cadherin/CD68/CD45 triple staining

Although Cell-in-cell structures (CICs) had been documented in human tumors for decades, it is unclear what types of CICs were formed largely due to low resolution of traditional way such as H&E staining. In this work, we employed immunofluorescent method to stain a panel of human tumor samples simultaneously with antibodies against E-cadherin for Epithelium, CD68 for Macrophage and CD45 for Leukocytes, which we termed as "EML method" based on the cells detected. Detail analysis revealed four types of CICs, with tumor cells or macrophage engulfing tumor cells or leukocytes respectively. Interestingly, tumor cells seem to be dominant over macrophage (93% vs 7%) as the engulfer cells in all CICs detected, whereas the overall amount of internalized tumor cells is comparable to that of internalized CD45(+) leukocytes (57% vs 43%). The CICs profiles vary from tumor to tumor, which may indicate different malignant stages and/or inflammatory conditions. Given the potential impacts different types of CICs might have on tumor growth, we therefore recommend EML analysis of tumor samples to clarify the correlation of CICs subtypes with clinical prognosis in future researches.