Semi-automatic PD-L1 Characterization and Enumeration of Circulating Tumor Cells from Non-small Cell Lung Cancer Patients by Immunofluorescence
| Autor: | Jessica Garcia, Léa Payen, Jean-Phillippe Aurel, Frédérique Le Breton, Kathryn W Li, Sébastien Couraud, David Barthelemy, Florence Geiguer, Brigitte Manship, Claire Rodriguez-Lafrasse, Julie Ballandier |
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| Přispěvatelé: | Hospices Civils de Lyon (HCL), Centre de Recherche en Cancérologie de Lyon (UNICANCER/CRCL), Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM), Biolidics Limited, Institut de Physique des 2 Infinis de Lyon (IP2I Lyon), Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS) |
| Rok vydání: | 2019 |
| Předmět: |
Epithelial-Mesenchymal Transition
Lung Neoplasms General Chemical Engineering Cell Microfluidics Fluorescent Antibody Technique [SDV.CAN]Life Sciences [q-bio]/Cancer Vimentin Cell Separation Immunofluorescence B7-H1 Antigen General Biochemistry Genetics and Molecular Biology Circulating tumor cell Carcinoma Non-Small-Cell Lung Cell Line Tumor medicine Biomarkers Tumor Humans medicine.diagnostic_test biology General Immunology and Microbiology Chemistry Immunomagnetic Separation General Neuroscience Mesenchymal stem cell medicine.disease Neoplastic Cells Circulating Primary tumor 3. Good health Lymphatic system medicine.anatomical_structure Cancer cell Cancer research biology.protein |
| Zdroj: | Journal of visualized experiments : JoVE Journal of visualized experiments : JoVE, JoVE, 2019, pp.e59873. ⟨10.3791/59873⟩ |
| ISSN: | 1940-087X |
| Popis: | International audience; Circulating tumor cells (CTCs) derived from the primary tumor are shed into the bloodstream or lymphatic system. These rare cells (1-10 cells per mL of blood) warrant a poor prognosis and are correlated with shorter overall survival in several cancers (e.g., breast, prostate and colorectal). Currently, the anti-EpCAM-coated magnetic bead-based CTC capturing system is the gold standard test approved by the U.S. Food and Drug Administration (FDA) for enumerating CTCs in the bloodstream. This test is based on the use of magnetic beads coated with anti-EpCAM markers, which specifically target epithelial cancer cells. Many studies have illustrated that EpCAM is not the optimal marker for CTC detection. Indeed, CTCs are a heterogeneous subpopulation of cancer cells and are able to undergo an epithelial-to-mesenchymal transition (EMT) associated with metastatic proliferation and invasion. These CTCs are able to reduce the expression of cell surface epithelial marker EpCAM, while increasing mesenchymal markers such as vimentin. To address this technical hurdle, other isolation methods based on physical properties of CTCs have been developed. Microfluidic technologies enable a label-free approach to CTC enrichment from whole blood samples. The spiral microfluidic technology uses the inertial and Dean drag forces with continuous flow in curved channels generated within a spiral microfluidic chip. The cells are separated based on the differences in size and plasticity between normal blood cells and tumoral cells. This protocol details the different steps to characterize the programmed death-ligand 1 (PD-L1) expression of CTCs, combining a spiral microfluidic device with customizable immunofluorescence (IF) marker set. |
| Databáze: | OpenAIRE |
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