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Titolo:
Fluorescent in situ hybridisation on tissue sections: a quantitative approach with confocal laser scanning microscopy
Autore:
Neri, TM; Carnevali, L; Orlandini, G; Gatti, R; Scandroglio, R; Savi, M; Allegri, L;
Indirizzi:
Univ Parma, Dept Clin Med Nephrol & Hlth Sci, I-43100 Parma, Italy Univ Parma Parma Italy I-43100 Nephrol & Hlth Sci, I-43100 Parma, Italy Univ Parma, Inst Histol & Gen Embryol, I-43100 Parma, Italy Univ Parma Parma Italy I-43100 istol & Gen Embryol, I-43100 Parma, Italy
Titolo Testata:
EUROPEAN JOURNAL OF HISTOCHEMISTRY
fascicolo: 2, volume: 44, anno: 2000,
pagine: 193 - 198
SICI:
1121-760X(2000)44:2<193:FISHOT>2.0.ZU;2-J
Fonte:
ISI
Lingua:
ENG
Soggetto:
NONRADIOACTIVE INSITU HYBRIDIZATION; MESSENGER-RNA; LABELED OLIGONUCLEOTIDES; IMAGE-ANALYSIS; EXPRESSION; PROBES; RAT;
Keywords:
in situ hybridisation; fluorescent oligonucleotides; confocal microscopy;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
18
Recensione:
Indirizzi per estratti:
Indirizzo: Neri, TM Univ Parma, Dept Clin Med Nephrol & Hlth Sci, Via Gramsci 14, I-43100 Parma, Italy Univ Parma Via Gramsci 14 Parma Italy I-43100 43100 Parma, Italy
Citazione:
T.M. Neri et al., "Fluorescent in situ hybridisation on tissue sections: a quantitative approach with confocal laser scanning microscopy", EUR J HIST, 44(2), 2000, pp. 193-198

Abstract

The use of fluorescent detection methods in association with digital microscopy technologies is an innovative approach for tissue localisation of messenger RNA. The success of such methods relies on the tissue preservation, local availability of the probe and on the existence of high resolution tridimensional analysis systems. Cryostatic sections, mild denaturation, shortoligonucleotide probes (20mer) and confocal laser scanning microscopy allow the fullfillment of all these conditions avoiding photobleaching and tissue autofluorescence. In this paper, we describe in detail a method for in situ hybridisation set up with digoxigenin-coupled oligonucleotide complementary to p-actin mRNA as a probe and an anti-hapten fluorescent antibody as second step for detecting specific hybridisation. Fluorescence was analysedby means of a confocal laser scanning microscope (CLSM) that provides images with low out-of-focus blurring also with relatively low numerical aperture (NA) objectives. We propose also an easy method to perform semi-quantitative thresholding analysis which allows to discriminate between background and specific signal.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 21/09/20 alle ore 05:42:27