title: Single cell adhesion assay using computer controlled micropipette.
abstract: Cell adhesion is a fundamental phenomenon vital for all multicellular organisms. Recognition of and adhesion to specific macromolecules is a crucial task of leukocytes to initiate the immune response. To gain statistically reliable information of cell adhesion, large numbers of cells should be measured. However, direct measurement of the adhesion force of single cells is still challenging and today's techniques typically have an extremely low throughput (5-10 cells per day). Here, we introduce a computer controlled micropipette mounted onto a normal inverted microscope for probing single cell interactions with specific macromolecules. We calculated the estimated hydrodynamic lifting force acting on target cells by the numerical simulation of the flow at the micropipette tip. The adhesion force of surface attached cells could be accurately probed by repeating the pick-up process with increasing vacuum applied in the pipette positioned above the cell under investigation. Using the introduced methodology hundreds of cells adhered to specific macromolecules were measured one by one in a relatively short period of time (∼30 min). We blocked nonspecific cell adhesion by the protein non-adhesive PLL-g-PEG polymer. We found that human primary monocytes are less adherent to fibrinogen than their in vitro differentiated descendants: macrophages and dendritic cells, the latter producing the highest average adhesion force. Validation of the here introduced method was achieved by the hydrostatic step-pressure micropipette manipulation technique. Additionally the result was reinforced in standard microfluidic shear stress channels. Nevertheless, automated micropipette gave higher sensitivity and less side-effect than the shear stress channel. Using our technique, the probed single cells can be easily picked up and further investigated by other techniques; a definite advantage of the computer controlled micropipette. Our experiments revealed the existence of a sub-population of strongly fibrinogen adherent cells appearing in macrophages and highly represented in dendritic cells, but not observed in monocytes.
authors.affiliation: Doctoral School of Molecular- and Nanotechnologies, University of Pannonia, Veszprém, Hungary; Nanobiosensorics Group, Research Centre for Natural Sciences, Institute for Technical Physics and Materials Science, Budapest, Hungary; Department of Biological Physics, Eötvös University, Budapest, Hungary.
authors.forename: Rita
authors.initials: R
authors.lastname: Salánki
authors.affiliation: Department of Hydrodynamic Systems, Budapest University of Technology and Economics, Budapest, Hungary.
authors.forename: Csaba
authors.initials: C
authors.lastname: Hős
authors.affiliation: Nanobiosensorics Group, Research Centre for Natural Sciences, Institute for Technical Physics and Materials Science, Budapest, Hungary; Department of Biological Physics, Eötvös University, Budapest, Hungary.
authors.forename: Norbert
authors.initials: N
authors.lastname: Orgovan
authors.affiliation: Doctoral School of Molecular- and Nanotechnologies, University of Pannonia, Veszprém, Hungary; Nanobiosensorics Group, Research Centre for Natural Sciences, Institute for Technical Physics and Materials Science, Budapest, Hungary.
authors.forename: Beatrix
authors.initials: B
authors.lastname: Péter
authors.affiliation: MTA-ELTE Immunology Research Group, Eötvös University, Budapest, Hungary.
authors.forename: Noémi
authors.initials: N
authors.lastname: Sándor
authors.affiliation: Department of Immunology, Eötvös University, Budapest, Hungary.
authors.forename: Zsuzsa
authors.initials: Z
authors.lastname: Bajtay
authors.affiliation: MTA-ELTE Immunology Research Group, Eötvös University, Budapest, Hungary; Department of Immunology, Eötvös University, Budapest, Hungary.
authors.forename: Anna
authors.initials: A
authors.lastname: Erdei
authors.affiliation: Nanobiosensorics Group, Research Centre for Natural Sciences, Institute for Technical Physics and Materials Science, Budapest, Hungary.
authors.forename: Robert
authors.initials: R
authors.lastname: Horvath
authors.affiliation: Nanobiosensorics Group, Research Centre for Natural Sciences, Institute for Technical Physics and Materials Science, Budapest, Hungary; Department of Biological Physics, Eötvös University, Budapest, Hungary; CellSorter Company for Innovations, Budapest, Hungary.
authors.forename: Bálint
authors.initials: B
authors.lastname: Szabó
mesh_terms: D001331:Automation; D002448:Cell Adhesion / Q000187:drug effects; D003201:Computers; D003713:Dendritic Cells / Q000166:cytology; D005340:Fibrinogen / Q000494:pharmacology; D006801:Humans; D057446:Hydrodynamics; D008264:Macrophages / Q000166:cytology; D044085:Microfluidics; D009000:Monocytes / Q000166:cytology* / Q000187:drug effects; D009716:Numerical Analysis, Computer-Assisted; D011092:Polyethylene Glycols / Q000494:pharmacology; D011107:Polylysine / Q000031:analogs & derivatives / Q000494:pharmacology; D033081:Shear Strength; D059010:Single-Cell Analysis / Q000295:instrumentation* / Q000379:methods*
chemical_list: C443714:polylysine-graft-(poly(ethylene glycol)); D011107:Polylysine; D011092:Polyethylene Glycols; D005340:Fibrinogen