Dynamic physical properties of dissociated tumor cells revealed by dielectrophoretic field-flow fractionation

Abstract

Metastatic disease results from the shedding of cancer cells from a solid primary tumor, their transport through the cardiovascular system as circulating tumor cells (CTCs) and their engraftment and growth at distant sites. Little is known about the properties and fate of tumor cells as they leave their growth site and travel as single cells. We applied analytical dielectrophoretic field-flow fractionation (dFFF) to study the membrane capacitance, density and hydrodynamic properties together with the size and morphology of cultured tumor cells after they were harvested and placed into single cell suspensions. After detachment, the tumor cells exhibited biophysical properties that changed with time through a process of cytoplasmic shedding whereby membrane and cytoplasm were lost. This process appeared to be distinct from the cell death mechanisms of apoptosis, anoikis and necrosis and it may explain why multiple phenotypes are seen among CTCs isolated from patients and among the tumor cells obtained from ascitic fluid of patients. The implications of dynamic biophysical properties and cytoplasmic loss for CTC migration into small blood vessels in the circulatory system, survival and gene expression are discussed. Because the total capacitance of tumor cells remained higher than blood cells even after they had shed cytoplasm, dFFF offers a compelling, antibody-independent technology for isolating viable CTCs from blood even when they are no larger than peripheral blood mononuclear cells.

ApoStream™, a new dielectrophoretic device for antibody independent isolation and recovery of viable cancer cells from blood

“…21 The isolation of rare cancer cells from blood in particular exploits differences in dielectric properties between blood cells (lymphocytes, monocytes, and granulocytes) and cancer cells. 19,20 For the separation of cancer cells from healthy blood cells, the ApoStream device operates in a modified form to conventional DEP-FFF, in that the cancer cells are attracted by positive DEP forces towards the electrode plane, and thus away from the bulk of the blood cells that are levitated by negative DEP into the hydrodynamic flow velocity profile. This is accomplished by applying the voltage signal at a frequency in between the so-called DEP crossover frequency of cancer cells and PBMCs.…”

“…19,20 In these earlier studies, DEP-FFF was applied using a batch mode configuration that limited the number of cells processed in a given run because cells must remain spaced by several diameters to avoid dipole-dipole interactions that can perturb DEP responses. 19,21 As a result, the loading capacity using the batch mode of operation was limited to less than a million cells per run and required processing of multiple batches in order to complete a CTC analysis for a typical 7.5 ml blood sample.…”

Automated electrorotation shows electrokinetic separation of pancreatic cancer cells is robust to acquired chemotherapy resistance, serum starvation, and EMT

“…Because the time constant associated with this relaxation is approximately proportional to the product of specific membrane capacitance and cell radius, the observed decrease in crossover frequency is explainable due to the observed increase in median cell radius and in median specific membrane capacitance of the gemcitabineresistant sub-clones (Table I). Wang et al, 41 Shim et al, 8 and Gascoyne et al 42 attribute higher specific membrane capacitances to larger effective membrane areas due to ruffles and folds in the membrane, changing the apparent effective specific capacitance. Alternatively, Gagnon et al 43 FIG.…”

“…According to Wang et al, 41 Shim et al, 8 and Gascoyne et al, 42 this increase in surface area would putatively change the ruffles and folds in the cell membrane, increasing their effective surface area and specific membrane capacitance. Furthermore, Salmanzadeh et al 49 and Mulhall et al 50 observed that aggressiveness of cancer cell phenotype correlates with an increase in specific membrane capacitance for mouse ovarian surface epithelial and oral cancer cells, respectively.…”

“…2 Furthermore, circulating pancreatic cells exist in a subset of patients without cancer but with potentially cancerous pancreatic cystic lesions. 4 To obtain this information, CTCs have been successfully captured from blood samples through mechanical, 7 electromechanical 8 and immunoaffinity methods, 9-11 the last being the most common. However, challenges exist capturing subsets of CTCs that have reduced antigen expression, often associated with the epithelial to mesenchymal transition (EMT) 12 or acquired resistance to chemotherapy.…”