title: Increased exosome production from tumour cell cultures using the Integra CELLine Culture System.
abstract: Exosomes are nanometer-sized vesicles, secreted from most cell types, with documented immune-modulatory functions. Exosomes can be purified from cultured cells but to do so effectively, requires maintenance of cells at high density in order to obtain sufficient accumulation of exosomes in the culture medium, prior to purification. Whilst high density cultures can be achieved with cells in suspension, this remains difficult with adherent cells, resulting in low quantity of exosomes for subsequent study. We have used the Integra CELLine culture system, originally designed for hybridoma cultures, to achieve a significant increase in obtainable exosomes from adherent and non-adherent tumour cells. Traditional cultures of mesothelioma cells (cultured in 75 cm(2) flasks) gave an average yield of 0.78 microg+/-0.14 microg exosome/ml of conditioned medium. The CELLine Adhere 1000 (CLAD1000) flask, housing the same cell line, increased exosome yield approximately 12 fold to 10.06 microg+/-0.97 microg/ml. The morphology, phenotype and immune function of these exosomes were compared, and found to be identical in all respects. Similarly an 8 fold increase in exosome production was obtained from NKL cells (a suspension cell line) using a CELLine 1000 (CL1000) flask. The CELLine system also incurred ~5.5 fold less cost and reduced labour for cell maintenance. This simple culture system is a cost effective, useful method for significantly increasing the quantity of exosomes available from cultured cells, without detrimental effects. This tool should prove advantageous in future studies of exosome-immune modulation in cancer and other settings.
authors.affiliation: Department of Oncology & Palliative Medicine, School of Medicine, Cardiff University, Velindre Cancer Centre, Whitchurch, Cardiff, UK. paul.mitchell@velindre-tr.wales.nhs.uk
authors.forename: J Paul
authors.initials: JP
authors.lastname: Mitchell
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Court
authors.forename: Malcolm David
authors.initials: MD
authors.lastname: Mason
authors.forename: Zsuzsanna
authors.initials: Z
authors.lastname: Tabi
authors.forename: Aled
authors.initials: A
authors.lastname: Clayton
mesh_terms: D002448:Cell Adhesion; D018929:Cell Culture Techniques; D002460:Cell Line; D049109:Cell Proliferation; D017077:Culture Media, Conditioned / Q000378:metabolism; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D054066:Leukemia, Large Granular Lymphocytic / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D008654:Mesothelioma / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D046529:Microscopy, Electron, Transmission; D010641:Phenotype; D010997:Pleural Neoplasms / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D022142:Secretory Vesicles / Q000276:immunology / Q000378:metabolism* / Q000648:ultrastructure; D013997:Time Factors; D014407:Tumor Cells, Cultured
chemical_list: D017077:Culture Media, Conditioned