Atomic Force Microscopy and Anodic Porous Allumina of Nucleic Acid Programmable Protein Arrays

Abstract

The methodological aspects are here presented for the NAPPA (Nucleic Acid Programmable Protein Arrays) characterization by atomic force microscopy and anodic porous alumina. Anodic Porous Alumina represents also an advanced on chip laboratory for gene expression contained in an engineered plasmid vector. The results obtained with CdK2, CDKN1A, p53 and Jun test genes expressed on NAPPA and the future developments are discussed in terms of our pertinent and recent Patents and of their possibility to overcome some limitations of present fluorescence detection in probing protein-protein interaction in both basic sciences and clinical studies.

Nanotechnologies in Protein Microarrays

“…Lectin-coated QDs (Poly)saccharide Protein glycosylation FRET between QDs-AuNPs [53] GNP (Poly)saccharide Lectin Fluorescence of FSNPs [54] NiNTA-DsRed FT Proteins Immobilization concept Fluorescence [55] Cys-DNA + recombinant fusion proteins Proteins Immobilization concept Fluorescence [56] DDI oligonucleotide-tagged proteins Proteins EGF Location of subcellular area of cells [57] Lipidic membrane + ganglioside receptors Proteins Cholera toxin b SPRi [72] SWNT + RA-specific (CCC) peptide Peptides Rheumatoid factor QCM [73] A-enolase peptides Peptides Sera-specific PDAC Abs QCM_D [74] Aptamer-AgNPs Nucleic acids/aptamers PDGF-BB SPR/fluorescence of modified AgNPs [58] ssDNA aptamer monolayer Nucleic acids/aptamers Thrombin Nanoparticles-enhanced SPRi [59] DNA bar code Nucleic acids/aptamers PSA NPs absorbance [75] DNA template Nucleic acids/aptamers BRIP1, Jun, ATF2 QCM_F + QCM_D [76] DNA template on anodic porous alumina Nucleic acids/aptamers CdK2, CDKN1A, p53, Jun AFM [77] Fluorescent SWNT + His-tagged proteins…”

“…AFM is applicable to a wide range of molecular species, many of which are difficult to label by popular tags, including proteins and other molecules [108]. AFM can validate protein expression on the array surface, whereas these measurements were applied to demonstrate gene expression using mica-coated slides containing 16 spots, having printed one 300 microns NAPPA spot containing around 400 genes [77]. Moreover, AFM-based scratching techniques offer unique approach for the fabrication of nanoscaled arrays [109], which can serve as a platform for implementation of other nanotechnological recognition probes, as aptamers or s ynthetic peptides.…”

Introduction to the special issue on 'Nanostructures by Valve Metal Anodization'

“…On excitation by a mercury lamp at ~530 nm wavelength, fluorescence emission was observed at ~593 nm in real time by a sensitive CCD photo camera coupled with the optical microscope. From the fluorescence maps frequency histograms were obtained, before and after a rinsing treatment that showed possible reuse of the substrates, for potential application in biomedical assays [21]. In particular, the authors observed by AFM the occurrence of APA pores in excess of 1 µm, even if anodization was carried out under standard conditions (0.5 M oxalic acid, 5-20 °C, 50 V).…”

Protein Confinement in the Anodic Porous Alumina Microarray

“…In recent years, APA has attracted significant interest for sensor [1][2], photonic devices [3] and protein crystals growth [4,5] due to the specific characteristics of its ordered multi-cavities array [6]. The fast development of the proteomic field and the complexity proteinprotein and proteins-small molecules interaction require new advanced protein arrays in micro and nanoscale dimension.…”