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5. Ultrasensitive Bioanalytical Imaging

Author

Mirasoli, Mara, primary, Venturoli, Simona, additional, Guardigli, Massimo, additional, Dolci, Luisa Stella, additional, Simoni, Patrizia, additional, Musiani, Monica, additional, and Roda, Aldo, additional

Published
2010
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14. Chemiluminescent low-light imaging of biospecific reactions on macro- and microsamples using a videocamera-based luminograph

Author

Roda, Aldo, Pasini, Patrizia, Musiani, Monica, Girotti, Stefano, Baraldini, Mario, Carrea, Giacomo, and Suozzi, Anna

Subjects
Imaging systems -- Evaluation, Fluorimetry -- Equipment and supplies, Chemiluminescence -- Diagnostic use, Chemistry
Abstract

The analytical performance of a low-light imaging luminograph for quantitative luminescence analysis was evaluated in terms of sensitivity, spatial resolution, accuracy, precision, and sample geometry, at the macrolevel and in combination with optical microscopy. The system allows for the detection of 400 amol of enzymes such as alkaline phosphatase and horseradish peroxidase using 1,2-dioxetanes and luminol/p-iodophenol or acridancarboxylate esters, respectively, as chemiluminescent substrates. Enzymatic activity and spatial distribution of nylon net immobilized-alkaline phosphatase was studied; the system permits the quantification of the immobilized enzyme with a spatial resolution as low as 1 [[micro]meter]. Other applications, such as the alkaline phosphatase localization in 8 [[micro]meter] intestinal mucosa cryosections, quantitative immunocytochemistry, and dot blot DNA hybridization reactions, were studied and optimized. The system was also employed for in situ hybridization assay of cytomegalovirus DNA in infected human fibroblasts. The presence of a viral genome was revealed with digoxigenin-labeled probes and alkaline phosphatase-labeled antidigoxigenin antibody, using chemiluminescent substrate for this enzyme. The luminescent signal was intense and stable, and the probe was imaged and quantified within single cells with higher intensity in the nuclei, with a spatial resolution as low as 1 [[micro]meter] and very low background. The results show that this technique is an ultrasensitive and potent analytical tool to localize and quantify biomolecules at microscopic level, and it is suitable for many bioanalytical applications.

Published
1996

27. Parvovirus B19-derived fetal complications following gestational infection

Author

BONVICINI, FRANCESCA, GALLINELLA, GIORGIO, GENTILOMI, GIOVANNA ANGELA, MANARESI, ELISABETTA, DI FURIO, FRANCESCA, GUERRA, BRUNELLA, SIMONAZZI, GIULIANA, PUCCETTI, CHIARA, MUSIANI, MONICA, ZERBINI, MARIALUISA, M. Contoli, F.Bonvicini, G.Gallinella, G.A.Gentilomi, E.Manaresi, F.Di Furio, B.Guerra, G.Simonazzi, C.Puccetti, M.Contoli, M.Musiani, and M.Zerbini.

Subjects
B19 fetal infection
Published
2011

28. Comparative evaluation of an automated assay for parvovirus B19 antibody detection

Author

BONVICINI, FRANCESCA, GALLINELLA, GIORGIO, GENTILOMI, GIOVANNA ANGELA, MANARESI, ELISABETTA, DI FURIO, FRANCESCA, MUSIANI, MONICA, ZERBINI, MARIALUISA, B. Guerra, SIMONAZZI, GIULIANA, C. Puccetti, M. Contoli, F.Bonvicini, G.Gallinella, G.A.Gentilomi, E.Manaresi, F.Di Furio, B.Guerra, G.Simonazzi, C.Puccetti, M.Contoli, M.Musiani, and M.Zerbini.

Subjects
B19 antibody detection
Published
2011

31. Infezione da Parvovirus B19 nella donna in gravidanza, nel feto e nel neonato

Author

BONVICINI, FRANCESCA, GALLINELLA, GIORGIO, MANARESI, ELISABETTA, DI FURIO, FRANCESCA, GUERRA, BRUNELLA, SIMONAZZI, GIULIANA, PUCCETTI, CHIARA, MUSIANI, MONICA, ZERBINI, MARIALUISA, GENTILOMI, GIOVANNA ANGELA, Bonvicini F., Gallinella G., Gentilomi G.A., Manaresi E, Di Furio F., Guerra B., Simonazzi G., Puccetti C., Musiani M., and Zerbini M.

Published
2010

32. Parvovirus B19 infection during pregnancy and evaluation of fetal outcomes

Author

BONVICINI, FRANCESCA, GALLINELLA, GIORGIO, GUERRA, BRUNELLA, SIMONAZZI, GIULIANA, PUCCETTI, CHIARA, MUSIANI, MONICA, ZERBINI, MARIALUISA, GENTILOMI, GIOVANNA ANGELA, Bonvicini F., Gallinella G., Gentilomi G.A., Guerra B., Simonazzi G., Puccetti C., Musiani M., and Zerbini M.

Published
2010

34. Bio-chemiluminescent miniaturized biosensors

Author

MIRASOLI, MARA, GUARDIGLI, MASSIMO, MICHELINI, ELISA, SIMONI, PATRIZIA, DOLCI, LUISA STELLA, CEVENINI, LUCA, MEZZANOTTE, LAURA, BURAGINA, ANGELA, BONVICINI, FRANCESCA, MUSIANI, MONICA, RODA, ALDO, Mirasoli M., Guardigli M., Michelini E., Simoni P., Dolci L. S., Cevenini L., Mezzanotte L., Buragina A., Bonvicini F., Musiani M., and Roda A.

Subjects
POINT-OF-CARE TESTING, LAB ON A CHIP, BIO-CHEMILUMINESCENCE, LUMINESCENCE IMAGING, PARVOVIRUS B19
Abstract

The Point-Of-Care (POC) approach is based on portable devices suitable to perform the analysis directly where it is required or where the sample is obtained. POC, which can provide significant advantages in the clinical field for its ability to provide accurate diagnosis at the patient bedside, can find applications in other fields, such as critical medicine, bio-terrorism, developing countries, food and environmental analyses, space station and telemedicine. A POC device should combine portability, minimum sample pre-treatment and ability to perform highly sensitive multiplexed assays in a short time. Microfluidic integrated systems relying on biospecific recognition reactions (e.g., immunological reactions, nucleic acids hybridization) and ultrasensitive bio-chemiluminescence (BL-CL) detection techniques represent one of the most promising options. Although several miniaturized analytical systems were developed, in most cases the miniaturization of the analytical format was not paralleled by an adequate miniaturization of the overall analytical device (e.g., use of laboratory instrumentation for signal detection). This work describes the development of POC devices exploiting “contact” BL-CL detection, in which the analytical signal is produced directly in the surface of an imaging light sensor able to localize and quantify the signal. In particular, the analytes are captured and detected in different positions of a transparent solid support, which is in contact, through a tapered fiber optic faceplate, with a portable cooled highly sensitive CCD sensor. “Contact” imaging, which provides high light collection efficiency, has been exploited in different analytical devices. A microfluidics-based device was developed for the diagnosis of parvovirus B19 (B19V) infection. In particular, target B19V DNA is captured by a specific peptide nucleic acid (PNA) probe immobilized on a glass surface, and then revealed by exploiting an enzyme label suitable for CL detection. The limit of detection (50 fmol/mL of B19V DNA) is comparable with that obtained with standard laboratory methodologies, with an overall assay time of 30 min. A new portable biodevice containing genetically engineered BL whole-cell biosensors was developed for multiplexed detection of environmental pollutants (e.g., compounds with hormone-like activity, heavy metals…). The cells were modified with the introduction of the gene encoding for a BL reporter protein (e.g., luciferase), so that its expression is controlled by the ability of the target analyte to interact with a specific regulatory proteins or receptor. The expressed reporter protein can be readily measured and directly related to the analyte concentration in the sample. Different recombinant yeast and bacterial biosensors were immobilized in a modified clear bottom black 384-well microplate to obtain a BL cell array suitable for on-site analysis. The cells array, which can be stored for up to 1 month at 4°C without loosing cell vitality, was characterized by high sensitivity (e.g., a LOD of 0.5 nM for testosterone was obtained with immobilized yeast cell-based biosensor for androgen detection after 1-month storage). The performance of “contact” imaging is evaluated and compared to that of conventional optics-based imaging performed with laboratory instrumentation.

Published
2010

36. Presenza e ruolo della metilazione dei dinucleotidi CpG nel DNA di parvovirus B19

Author

BONVICINI, FRANCESCA, DI FURIO, FRANCESCA, MANARESI, ELISABETTA, VENTUROLI, SIMONA, GENTILOMI, GIOVANNA ANGELA, ZERBINI, MARIALUISA, MUSIANI, MONICA, GALLINELLA, GIORGIO, L. De Falco, F. Chessa, F.Bonvicini, F.Di Furio, L.De Falco, F.Chessa, E.Manaresi, S.Venturoli, G.A.Gentilomi, M.Zerbini, M.Musiani, and G.Gallinella.

Published
2010

38. 'Human parvovirus'

Author

BONVICINI, FRANCESCA, MUSIANI, MONICA, ZERBINI, MARIALUISA, F.Bonvicini, M.Musiani, and M.Zerbini.

Abstract

Capitolo di “Antimicrobial Therapy and Vaccines", Volume I: Microbes, Second Edition

Published
2009

39. 'Human Bocavirus'

Author

BONVICINI, FRANCESCA, MUSIANI, MONICA, ZERBINI, MARIALUISA, F.Bonvicini, M.Musiani, and M.Zerbini.

Abstract

Capitolo di “Antimicrobial Therapy and Vaccines, Volume I: Microbes, Second Edition

Published
2009

46. Luminescence detection in point-of-care testing devices

Author

RODA, ALDO, GUARDIGLI, MASSIMO, MIRASOLI, MARA, MICHELINI, ELISA, DOLCI, LUISA STELLA, SIMONI, PATRIZIA, MUSIANI, MONICA, Roda A., Guardigli M., Mirasoli M., Michelini E., Dolci L.S., Simoni P., and Musiani M.

Published
2008

50. A proteomic approach to the analysis of B19 capsid proteins

Author

MANARESI, ELISABETTA, BONVICINI, FRANCESCA, GENTILOMI, GIOVANNA ANGELA, MUSIANI, MONICA, ZERBINI, MARIALUISA, GALLINELLA, GIORGIO, C. Filippone, E.Manaresi, C.Filippone, F.Bonvicini, G.A. Gentilomi, M.Musiani, M.Zerbini, and G.Gallinella

Published
2007

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