Bioactive surfaces for antibody-antigen complex detection by Atomic Force Microscopy

6th Vacuum and Surface Sciences Conference of Asia and Australia

Ierardi, Vincenzo^1,2 ; Ferrera, Francesca^3 ; Millo, Enrico^3,4 ; Damonte, Gianluca^3,4 ; Filaci, Gilberto^3,5 ; Valbusa, Ugo^1,2

Nanobiotechnologies, National Institute of Cancer Research (IST), Largo R. Benzi, Genova 16132, Italy^1

Nanomed Lab, Physics Department, University of Genova, Via Dodecaneso, 33, Genova, 16146, Italy^2

Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV, 7, Genova, 16146, Italy^3

Department of Experimental Medicine, University of Genova, Viale Benedetto XV, 1, Genova, 16132, Italy^4

Department of Internal Medicine, University of Genova, Viale Benedetto XV, 6, Genova, 16146, Italy^5

关键词: Antibody-antigen;    Bioactive surfaces;    Biological samples;    Functionalizations;    Label-free detection;    Roughness surfaces;    Solid substrates;    Surface activation;   

Others  :  https://iopscience.iop.org/article/10.1088/1742-6596/439/1/012001/pdf
DOI  :  10.1088/1742-6596/439/1/012001

来源: IOP

【 摘 要 】

Recently there has been a great develop of new antibodies immobilization procedures, that keep antibodies to retain their orientation and functionality after the binding to a solid support. This allows the formation of immune-complexes useful for the detection of biomarkers from biological samples. We have developed a new method of functionalization for solid substrates that involves an initial surface activation, then a functionalization by means of 3-aminopropyltriethoxysilane-followed by another functionalization step with a layer of very small peptides, which have a high affinity to the Antibody Fc portion, acting as antibody linkers. These antibody binding peptides can immobilize the antibodies with a proper configuration that allows an unambiguous detection of antibody-antigen complexes by means of atomic force microscopy (AFM). The AFM can act as a powerful label free detection technique, which allows to detect, in principle, single molecule interactions, with the only limitation to use substrate with low-roughness surfaces; in this case, the roughness can be interpreted as background noise in the AFM analysis. Moreover, our functionalization method can be used to obtain bioactive surfaces on a wide range of solid supports, making them capable to suitably immobilize the antibodies for the antigenic binding.