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Journal of Histochemistry and Cytochemistry, Vol. 48, 1587-1592, December 2000, Copyright © 2000, The Histochemical Society, Inc.

RAPID COMMUNICATION

Sensitive and Quantitative Co-detection of Two mRNA Species by Double Radioactive In Situ Hybridization

Hélène Salina, Serge Maitrejeanb, Jacques Malleta, and Sylvie Dumasa
a Laboratoire de Génétique Moléculaire de la Neurotransmission et des Processus Neurodégénératifs, UMR C9923, CNRS, Hôpital de la Pitié Salpêtrière, Paris, France
b Biospace Mesures, Paris, France

Correspondence to: Sylvie Dumas, LGN, Bât. CERVI, 5ème étage, Hôpital de la Pitié-Salpêtrière, 83, bd de l'Hôpital, 75013 Paris, France. E-mail: sdumas@infobiogen.fr

A better understanding of biological phenomena involving modulations of gene expression requires quantitative analysis of the expression of several genes in the same structure. For this purpose, we have developed a novel in situ hybridization method to quantify two different mRNA species in the same tissue section simultaneously. Two probes labeled with radioelements of significantly different energies (3H and 33P or 35S) were used to detect the mRNA species. Radioactive images corresponding to the detected mRNA species were acquired with a Micro Imager, a real-time, high-resolution digital autoradiography system. An algorithm was used to process the data such that the initial radioactive image acquired was filtered into two subimages, each representative of the hybridization result specific to one probe. This novel method allows local discrimination and quantification of the respective contributions of each label to each pixel and can therefore be used for quantitative analysis of two mRNAs with a resolution of 15–20 µm. (J Histochem Cytochem 48:1587–1591, 2000)

Key Words: in situ hybridization, double radioactive labeling, Micro Imager, co-detection, gene expression


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