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Journal of Histochemistry and Cytochemistry, Vol. 49, 1177-1182, September 2001, Copyright © 2001, The Histochemical Society, Inc.

BRIEF REPORT

Simultaneous Detection of RNA and Protein by In Situ Hybridization and Immunological Staining

Hideyuki Nagasoa, Takehide Murataa, Noel Daya, and Kazunari K. Yokoyamaa
a Gene Engineering Division, BioResource Center, RIKEN (The Institute of Physical and Chemical Research), Tsukuba Science City, Ibaraki, Japan

Correspondence to: Kazunari K. Yokoyama, RIKEN (The Institute of Physical and Chemical Research), Bioresource Center, 3-1-1 Koyadai, Tsukuba, Ibaraki 305-0074, Japan. E-mail: kazu@rtc.riken.go.jp

Proteinase K is widely used in methods for detection of transcripts in biological specimens by in situ hybridization (ISH). However, treatment with proteinase K hampers detection of RNA and protein simultaneously. We have developed a method for double staining of transcripts and proteins by ISH and IHC staining in imaginal discs and embryos of Drosophila. Instead of treatment with proteinase K, samples are treated with ethanol plus xylene and with acetone. Acetone renders cell membranes permeable to probes and antibodies without damaging tissue integrity, whereas treatment with proteinase K sometimes damages tissues. Treatment of samples with acetone allows hybridization of probe with transcripts in tissue. It is also effective for immunological staining of samples after ISH with a riboprobe. Thus, our method allows detection not only of transcripts but also of specific proteins in relatively intact single samples. (J Histochem Cytochem 49:1177–1182, 2001)

Key Words: in situ hybridization, immunological staining, Drosophila, imaginal disc, embryo


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