title: In-situ and label-free optical monitoring of the adhesion and spreading of primary monocytes isolated from human blood: dependence on serum concentration levels.
abstract: Adhesion and spreading of primary monocytes isolated from human blood were monitored utilizing optical waveguide lightmode spectroscopy (OWLS); a highly sensitive label-free biosensor technique using evanescent optical waves generated at a biocompatible surface. Appropriate development on a custom built setup enabled the OWLS cuvette to be operated as a 1.5 ml mini-incubator, controlling both temperature and CO2 levels. The incubator-equipped OWLS is readily applicable for delicate and long-term studies on sensitive primary cells, demonstrated here through monitoring the serum dependence of the adhesion and spreading of human monocytes. Moreover, the custom-built setup enables the simultaneous monitoring of the position and overall width of the OWLS resonant peaks. This unique feature makes it possible to distinguish the refractive index variations induced by the adsorption of secreted material from refractive index changes provoked by cellular spreading. A definite attachment and spreading activity was observed on the substratum (glassy silica-titania), when the serum level of the culturing medium was 0.0-0.01%. Increasing serum concentration resulted in a steep fall in monocyte surface adhesion and spreading. 1.0% serum level practically abolished all spreading activity measured by OWLS, and the number of attached cells was significantly decreased, too. Serum addition to fully spread cells provoked a reduction in the cell-substratum contact area, clearly detectable by the biosensor. Cell spreading was inhibited by pre-coating the sensor surface with considerable amounts of serum proteins. These findings suggest that monocyte spreading is inhibited by the adsorption of serum biomolecules to the substratum, rather than by soluble factors present in the serum. All of these results were obtained completely noninvasively with real time monitoring; demonstrating the capabilities of OWLS to sensitively monitor the adhesion properties of immune cells isolated from human blood. The current study is, therefore, a significant step towards the application of label-free optical biosensors in medical diagnostics.
authors.affiliation: Department of Biological Physics, Eötvös Loránd University, Pázmány P. stny. 1/A, H-1117 Budapest, Hungary; Nanobiosensorics Group, Hungarian Academy of Sciences, Research Centre for Natural Sciences, Institute for Technical Physics and Materials Science, Konkoly-Thege út 29-33, H-1120 Budapest, Hungary.
authors.forename: Norbert
authors.initials: N
authors.lastname: Orgovan
authors.affiliation: Nanobiosensorics Group, Hungarian Academy of Sciences, Research Centre for Natural Sciences, Institute for Technical Physics and Materials Science, Konkoly-Thege út 29-33, H-1120 Budapest, Hungary; Doctoral School of Molecular- and Nanotechnologies, University of Pannonia, Veszprém, Hungary.
authors.forename: Rita
authors.initials: R
authors.lastname: Salánki
authors.affiliation: MTA-ELTE Immunology Research Group, ELTE, Pázmány P. stny. 1/C, H-1117 Budapest, Hungary.
authors.forename: Noémi
authors.initials: N
authors.lastname: Sándor
authors.affiliation: Department of Immunology, Eötvös Loránd University, Pázmány P. stny. 1/C, H-1117 Budapest, Hungary.
authors.forename: Zsuzsa
authors.initials: Z
authors.lastname: Bajtay
authors.affiliation: MTA-ELTE Immunology Research Group, ELTE, Pázmány P. stny. 1/C, H-1117 Budapest, Hungary; Department of Immunology, Eötvös Loránd University, Pázmány P. stny. 1/C, H-1117 Budapest, Hungary.
authors.forename: Anna
authors.initials: A
authors.lastname: Erdei
authors.affiliation: Department of Biological Physics, Eötvös Loránd University, Pázmány P. stny. 1/A, H-1117 Budapest, Hungary.
authors.forename: Bálint
authors.initials: B
authors.lastname: Szabó
authors.affiliation: Nanobiosensorics Group, Hungarian Academy of Sciences, Research Centre for Natural Sciences, Institute for Technical Physics and Materials Science, Konkoly-Thege út 29-33, H-1120 Budapest, Hungary. Electronic address: horvathr@mfa.kfki.hu.
authors.forename: Robert
authors.initials: R
authors.lastname: Horvath
mesh_terms: D015374:Biosensing Techniques / Q000295:instrumentation*; D002448:Cell Adhesion; D002478:Cells, Cultured; D004867:Equipment Design; D006801:Humans; D008858:Microscopy, Phase-Contrast / Q000295:instrumentation; D009000:Monocytes / Q000166:cytology*; D044967:Serum / Q000737:chemistry
keywords: Cellular adhesion; Label-free optical biosensor; Mini-incubator; Optical waveguide lightmode spectroscopy; Primary monocyte; Serum dependence