The analysis of CTC with high sensitivity and specificity is challenging, as it is required to pinpoint individual CTC in a 7.5 mL blood sample with more than 50 million leukocytes [
14]. In the current study, the two technologies CytoTrack and CellSearch are compared under in vitro conditions. CytoTrack is a novel method recently described [
9] and the CellSearch CTC Test is FDA-approved as an aid in the monitoring of patients with metastatic breast, colon, and prostatic cancer [
15]. It should be pointed out that the CytoTrack technology has as yet not been clinically validated although work is ongoing with serial measurements of CTC among patients with metastatic breast cancer [
16]. The two technologies use very different approaches. Whereas the CellSearch approach is a ‘closed’ automated, FDA cleared system intended for clinical monitoring, the CytoTrack approach is an ‘open’ manual research system which allow for studying other markers for CTC enumeration and characterization of CTC. In CytoTrack, all nucleated cells in a blood sample are DAPI-stained, immunostained with antibodies against cytokeratin and CD45 and cytosmeared over a CytoDisc. Afterwards, fluorescence scanning is performed, suspected CTC on the disc are identified, and an image gallery is created for manual validation. In CellSearch, plasma is removed and magnetic sorting of EpCAM positive cells is carried out, subsequently the cells are stained for cytokeratins 8, 18, and 19, DAPI and CD45 and placed in a magnetic cartridge where image galleries of suspected CTC events are recorded for manual validation. The different approaches could in principle give rise to significantly different abilities to detect CTC, with CytoTrack relying on cytokeratin signal to detect cells and CellSearch dependent on both EpCAM and cytokeratin expression. In the current study of in vitro detection of MCF-7, a cytokeratin- and EpCAM-positive cell line, the two CTC technologies CytoTrack and CellSearch have similar recovery of cells spiked into blood (69 vs. 71 %,
p = 0.58). It should be noted that the recoveries were estimated on the basis of FACSAria spiking counts of cells in blood samples, a count which was previously estimated to result in actual spiking numbers around 74 % of the expected FACSAria spiking cell counts [
9]. The spiking was performed with care taken to obtain single cells; however, a fraction of cell clusters contained two or more cells. Therefore, the results of both CTC number (total number of both single cells and cells in clusters), cells in clusters (only cells in clusters counted), and clusters (the number of clusters counted) were recorded. Both CTC, cells in clusters and clusters had similar recoveries by CytoTrack and CellSearch. CellSearch, however, has a lower variability (CV%) in the analysis, which may be due to the automated procedures in CellSearch versus manual procedures in the CytoTrack analysis.
Nonetheless, CytoTrack proves to have a recovery and a reproducibility matching that of CellSearch, which is promising with regard to future studies including test of CTC in clinical samples from breast, colon, and prostate cancer patients. Moreover, further markers, both surface and intracellular phenotypic and genetic, can be explored to improve the performance of the CytoTrack across different cell types. A specific challenge for CellSearch is when CTC undergo epithelial to mesenchymal transition (EMT) as the EpCAM expression is reduced or not present in mesenchymal cells which makes it difficult to detect these cells [
17,
18]. The possibility to explore markers in cells of mesenchymal origin thus seems especially interesting since if markers for EMT are present on the surface or inside CTC it is possible for CytoTrack to find the cells. This is the first in vitro comparison between CytoTrack and CellSearch, and considerable work is required for further comparison. CytoTrack is a novel technology and thus there is a need for more in vitro studies of cells from different cancer types spiked into blood and in vitro studies with EpCAM or CK weak cells to investigate the limits of the two technologies. Also, studies of basic clinical performance in various cancers as well as studies of intra- and inter-lab and inter-observer variability are required. In conclusion, our data indicate that CytoTrack and CellSearch despite fundamentally different technologies may yield similar results when using a cell line selected to match the markers used by both technologies. Our investigation provides a basis for further studies using either different cells lines and/or different CTC markers.