@misc{10481/72537,
year = {2021},
month = {12},
url = {http://hdl.handle.net/10481/72537},
abstract = {The isolation of circulating tumour cells (CTCs) in colorectal cancer (CRC) mostly relies
on the expression of epithelial markers such as EpCAM, and phenotypic characterisation is usually
performed under fluorescence microscopy with only one or two additional markers. This limits
the ability to detect different CTC subpopulations based on multiple markers. The aim of this
work was to develop a novel protocol combining two platforms (IsoFluxTM and ImageStream®X) to
improve CTC evaluation. Cancer cell lines and peripheral blood from healthy donors were used to
evaluate the efficiency of each platform independently and in combination. Peripheral blood was
extracted from 16 early CRC patients (before loco-regional surgery) to demonstrate the suitability of
the protocol for CTC assessment. Additionally, peripheral blood was extracted from nine patients
one month after surgery to validate the utility of our protocol for identifying CTC subpopulation
changes over time. Results: Our protocol had a mean recovery efficiency of 69.5% and a limit of
detection of at least four cells per millilitre. We developed an analysis method to reduce noise from
magnetic beads used for CTC isolation. CTCs were isolated from CRC patients with a median of
37 CTCs (IQ 13.0–85.5) at baseline. CTCs from CRC patients were significantly (p < 0.0001) larger than cytokeratin (CK)-negative cells, and patients were stratified into two groups based on BRAFV600E
and PD-L1 expression on CK-positive cells. The changes observed over time included not only the
number of CTCs but also their distribution into four different subpopulations defined according to
BRAFV600E and PD-L1 positivity. We developed a novel protocol for semi-automatic CTC isolation
and phenotypic characterisation by combining two platforms. Assessment of CTCs from early CRC
patients using our protocol allowed the identification of two clusters of patients with changing
phenotypes over time.},
organization = {"Garantia Juvenil" fellowship	8040},
organization = {Ministry of Economy, Competitiveness, Enterprises and Universities	DOC_01682},
publisher = {MDPI},
keywords = {Circulating tumour cells},
keywords = {IsoFlux},
keywords = {ImageStream},
keywords = {CRC},
keywords = {CTC heterogeneity},
title = {Deep Phenotypic Characterisation of CTCs by Combination of Microfluidic Isolation (IsoFlux) and Imaging Flow Cytometry (ImageStream)},
doi = {10.3390/cancers13246386},
author = {Ruiz Rodríguez, Antonio J. and Molina Vallejo, María Pilar and Aznar Peralta, Inés and González Puga, María Cristina and Cañas García, Inés and González Flores, Encarnación and Lorente Acosta, José Antonio and Serrano Fernández, María José and Garrido Navas, María del Carmen},
}