title: High-resolution imaging for the detection and characterisation of circulating tumour cells from patients with oesophageal, hepatocellular, thyroid and ovarian cancers.
abstract: Interest has increased in the potential role of circulating tumour cells in cancer management. Most cell-based studies have been designed to determine the number of circulating tumour cells in a given volume of blood. Ability to understand the biology of the cancer cells would increase the clinical potential. The purpose of this study was to develop and validate a novel, widely applicable method for detection and characterisation of circulating tumour cells. Cells were imaged with an ImageStream(X) imaging flow cytometer which allows detection of expression of multiple biomarkers on each cell and produces high-resolution images. Depletion of haematopoietic cells was by red cell lysis, leukocyte common antigen CD45 depletion and differential centrifugation. Expression of epithelial cell adhesion molecule, cytokeratins, tumour-type-specific biomarkers and CD45 was detected by immunofluorescence. Nuclei were identified with DAPI or DRAQ5 and brightfield images of cells were collected. The method is notable for the dearth of cell damage, recoveries greater than 50%, speed and absence of reliance on the expression of a single biomarker by the tumour cells. The high-quality images obtained ensure confidence in the specificity of the method. Validation of the methodology on samples from patients with oesophageal, hepatocellular, thyroid and ovarian cancers confirms its utility and specificity. Importantly, this adaptable method is applicable to all tumour types including those of nonepithelial origin. The ability to measure simultaneously the expression of multiple biomarkers will facilitate analysis of the cancer cell biology of individual circulating tumour cells.
authors.affiliation: Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, United Kingdom.|Newcastle upon Tyne Hospitals NHS Foundation Trust, Northern Oesophago-Gastric Cancer Unit, Newcastle upon Tyne, United Kingdom.
authors.forename: Barry M
authors.initials: BM
authors.lastname: Dent
authors.affiliation: Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, United Kingdom.
authors.forename: Laura F
authors.initials: LF
authors.lastname: Ogle
authors.affiliation: Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, United Kingdom.|Queen Elizabeth Hospital, Northern Gynaecological Oncology Centre, Gateshead, United Kingdom.
authors.forename: Rachel L
authors.initials: RL
authors.lastname: O'Donnell
authors.affiliation: Newcastle upon Tyne Hospitals NHS Foundation Trust, Northern Oesophago-Gastric Cancer Unit, Newcastle upon Tyne, United Kingdom.
authors.forename: Nicholas
authors.initials: N
authors.lastname: Hayes
authors.affiliation: Newcastle upon Tyne Hospitals NHS Foundation Trust, Northern Centre for Cancer Care, Newcastle upon Tyne, United Kingdom.
authors.forename: Ujjal
authors.initials: U
authors.lastname: Malik
authors.affiliation: Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, United Kingdom.
authors.forename: Nicola J
authors.initials: NJ
authors.lastname: Curtin
authors.affiliation: Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, United Kingdom.
authors.forename: Alan V
authors.initials: AV
authors.lastname: Boddy
authors.affiliation: Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, United Kingdom.|Newcastle upon Tyne Hospitals NHS Foundation Trust, Northern Centre for Cancer Care, Newcastle upon Tyne, United Kingdom.
authors.forename: E Ruth
authors.initials: ER
authors.lastname: Plummer
authors.affiliation: Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, United Kingdom.|Queen Elizabeth Hospital, Northern Gynaecological Oncology Centre, Gateshead, United Kingdom.
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Edmondson
authors.affiliation: Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, United Kingdom.|Newcastle upon Tyne Hospitals NHS Foundation Trust, The Liver Unit, Newcastle upon Tyne, United Kingdom.
authors.forename: Helen L
authors.initials: HL
authors.lastname: Reeves
authors.affiliation: Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, United Kingdom.|Newcastle University Institute for Ageing, Newcastle upon Tyne, United Kingdom.
authors.forename: Felicity E B
authors.initials: FE
authors.lastname: May
authors.affiliation: Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, United Kingdom.
authors.forename: David
authors.initials: D
authors.lastname: Jamieson
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000954:Antigens, Surface / Q000378:metabolism; D014408:Biomarkers, Tumor / Q000378:metabolism; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D016130:Immunophenotyping; D008113:Liver Neoplasms / Q000175:diagnosis*; D008297:Male; D008875:Middle Aged; D009362:Neoplasm Metastasis; D009367:Neoplasm Staging; D009360:Neoplastic Cells, Circulating / Q000378:metabolism* / Q000473:pathology*; D010051:Ovarian Neoplasms / Q000175:diagnosis*; D013964:Thyroid Neoplasms / Q000175:diagnosis*
chemical_list: D000954:Antigens, Surface; D014408:Biomarkers, Tumor
keywords: ImageStreamX imaging flow cytometry; circulating tumour cells; hepatocellular carcinoma; oesophageal adenocarcinoma; ovarian cancer; thyroid carcinoma