Multiparametric analysis of single circulating cell by intravital flow cytometry
Abstract
Circulating tumor cells (CTCs) are cancer cells that originate from primary tumor and release into systemic or lymphatic system. Some of these tumor cells will lodge into distant organs and initial new cancer growth. And thus, CTCs constitute a major cause for mortality and provides a marker for staging progression, evaluating therapy, and monitoring recurrence. Current methods on CTC detection are blood tests. A blood test involves blood extraction, isolation/labeling of CTCs, and then quantitative analysis. Because CTCs are rare in initial stage of metastasis, the detection sensitivity of blood tests is limited by extraction volume, usually 10-20 mL. Owing to this reason, blood tests cannot reliably detect CTCs in early stage. Additionally, a blood test is not preferred to be performed periodically due to anemia of cancer patients and time/labor consuming. To this end, we developed a method-intravital flow cytometry (IFC) to quantitate CTCs in vivo as well as to address the above technical issues of blood tests. This method involves intravenous injection of a tumor-specific fluorescent conjugate (folate conjugates or DUPA conjugates) that binds to folate receptors or PSMA on CTCs followed by multiphoton imaging of superficial vasculature to quantitate CTCs. Studies in a murine metastatic model demonstrated that CTCs can be detected and quantitated two weeks before micrometastases can be identified by conventional techniques. Based on the evidence that folate or DUPA conjugates can label CTCs from peripheral blood with a specificity of >99%, we adapted IFC to develop a fast and simple method for CTC quantitation using conventional flow cytometry. Using the optimized method, human CTCs can be selectively labeled and quantitated when present at ∼2 CTCs/ml, opening opportunities for earlier assessment of metastatic disease. Finally, we have also explored a method for quantitation of cell adhesion in vivo based on statistical modeling using spatial-tempo cross-correlation algorithm.
Degree
Ph.D.
Advisors
Low, Purdue University.
Subject Area
Analytical chemistry|Biophysics
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