title: Multispot, label-free biodetection at a phantom plastic-water interface.
abstract: Recognizing and quantifying specific biomolecules in aqueous samples are constantly needed in research and diagnostic laboratories. As the typical detection procedures are rather lengthy and involve the use of labeled secondary antibodies or other agents to provide a signal, efforts have been made over the last 10 y to develop alternative label-free methods that enable direct detection. We propose and demonstrate an extremely simple, low-cost, label-free biodetector based on measuring the intensity of light reflected by the interface between a fluid sample and an amorphous fluoropolymer substrate having a refractive index very close to that of water and hosting various antibodies immobilized in spots. Under these index-matching conditions, the amount of light reflected by the interface allows straightforward quantification of the amount of antigen binding to each spot. Using antibodies targeting heterologous immunoglobulins and antigens commonly used as markers for diagnoses of hepatitis B and HIV, we demonstrate the limit of detection of a few picograms per square millimeter of surface-bound molecules. We also show that direct and real-time access to the amount of binding molecules allows the precise extrapolation of adhesion rates, from which the concentrations of antigens in solution can be estimated down to fractions of nanograms per milliliter.
authors.affiliation: Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, Università degli Studi di Milano, 20090 Segrate, Italy.
authors.forename: Fabio
authors.initials: F
authors.lastname: Giavazzi
authors.forename: Matteo
authors.initials: M
authors.lastname: Salina
authors.forename: Roberto
authors.initials: R
authors.lastname: Cerbino
authors.forename: Mattia
authors.initials: M
authors.lastname: Bassi
authors.forename: Davide
authors.initials: D
authors.lastname: Prosperi
authors.forename: Erica
authors.initials: E
authors.lastname: Ceccarello
authors.forename: Francesco
authors.initials: F
authors.lastname: Damin
authors.forename: Laura
authors.initials: L
authors.lastname: Sola
authors.forename: Marco
authors.initials: M
authors.lastname: Rusnati
authors.forename: Marcella
authors.initials: M
authors.lastname: Chiari
authors.forename: Bice
authors.initials: B
authors.lastname: Chini
authors.forename: Tommaso
authors.initials: T
authors.lastname: Bellini
authors.forename: Marco
authors.initials: M
authors.lastname: Buscaglia
mesh_terms: D000906:Antibodies / Q000378:metabolism; D000941:Antigens / Q000302:isolation & purification* / Q000378:metabolism; D015415:Biomarkers / Q000378:metabolism*; D002623:Chemistry Techniques, Analytical / Q000379:methods*; D015658:HIV Infections / Q000175:diagnosis; D006509:Hepatitis B / Q000175:diagnosis; D006801:Humans; D007118:Immunoassay; D008027:Light; D055620:Optical Phenomena; D010969:Plastics / Q000737:chemistry*; D040081:Protein Array Analysis; D014867:Water / Q000737:chemistry*
chemical_list: D000906:Antibodies; D000941:Antigens; D015415:Biomarkers; D010969:Plastics; D014867:Water
keywords: biomolecular detection; immunoassay; optical biosensor; protein microarray; reflective phantom interface