title: Creating adhesive and soluble gradients for imaging cell migration with fluorescence microscopy.
abstract: Cells can sense and migrate towards higher concentrations of adhesive cues such as the glycoproteins of the extracellular matrix and soluble cues such as growth factors. Here, we outline a method to create opposing gradients of adhesive and soluble cues in a microfluidic chamber, which is compatible with live cell imaging. A copolymer of poly-L-lysine and polyethylene glycol (PLL-PEG) is employed to passivate glass coverslips and prevent non-specific adsorption of biomolecules and cells. Next, microcontact printing or dip pen lithography are used to create tracks of streptavidin on the passivated surfaces to serve as anchoring points for the biotinylated peptide arginine-glycine-aspartic acid (RGD) as the adhesive cue. A microfluidic device is placed onto the modified surface and used to create the gradient of adhesive cues (100% RGD to 0% RGD) on the streptavidin tracks. Finally, the same microfluidic device is used to create a gradient of a chemoattractant such as fetal bovine serum (FBS), as the soluble cue in the opposite direction of the gradient of adhesive cues.
authors.affiliation: Centre for Vascular Research and Australian Centre for Nanomedicine, The University of New South Wales.
authors.forename: Siti Hawa
authors.initials: SH
authors.lastname: Ngalim
authors.forename: Astrid
authors.initials: A
authors.lastname: Magenau
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhu
authors.forename: Lotte
authors.initials: L
authors.lastname: Tønnesen
authors.forename: Zoe
authors.initials: Z
authors.lastname: Fairjones
authors.forename: J Justin
authors.initials: JJ
authors.lastname: Gooding
authors.forename: Till
authors.initials: T
authors.lastname: Böcking
authors.forename: Katharina
authors.initials: K
authors.lastname: Gaus
mesh_terms: D000269:Adhesives / Q000737:chemistry / Q000494:pharmacology; D000818:Animals; D001710:Biotin / Q000737:chemistry; D002417:Cattle; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology*; D002630:Chemotactic Factors / Q000737:chemistry* / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D055767:Immobilized Proteins / Q000737:chemistry / Q000494:pharmacology; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000379:methods*; D009842:Oligopeptides / Q000737:chemistry / Q000494:pharmacology; D012995:Solubility; D019809:Streptavidin / Q000737:chemistry
chemical_list: D000269:Adhesives; D002630:Chemotactic Factors; D055767:Immobilized Proteins; D009842:Oligopeptides; D001710:Biotin; C047981:arginyl-glycyl-aspartic acid; D019809:Streptavidin