Chapter 13
In Situ Hybridization for RNA: Nonradioactive
Probe: Oligo-DNA Probe: Digoxigenin (II)
MASANOBU MIYAZAKI, YOSHIYUKI OZONO, TAKASHI HARADA
AND SHIGERU KOHNO
Introduction
Nonradioactive in situ hybridization is thought to be less sensitive than the radioactive method, especially when used with oligo-DNA probe. However, the sensitivity of nonradioactive in situ
hybridization using oligo-DNA is sufficient for localization of
mRNA at cellular level even in human biopsy specimens. The
method of nonradioactive in situ hybridization was developed
in our laboratory in 1994 (Miyazaki et al. 1994) and was applied
to identify and localize mRNA of several factors synthesized by
renal cells in normal individuals and patients with a variety of
renal diseases (Suzuki et al. 1995; Miyazaki et al. 1996). Nonradioactive in situ hybridization with oligo-DNA has several advantages compared with the radioactive method, including a
high resolution, good probe stability and no exposure to radiation, as described in detail by Koji and Nakane (1996). Compared
with other types of probes, such as RNA or eDNA probe, oligoDNA has several benefits. First, oligo-DNA is easier to prepare.
While several years ago, a special DNA synthesizer that was available only in certain laboratories, was necessary for the preparation of oligo-DNA, such synthesizers are less expensive nowadays. Alternatively, custom made probes can be purchased
nowadays from several commercial sources. Second, a special
knowledge of molecular technology and expertise, e.g., cloning
techniques, is not necessary. Finally, the probes can be easily deCorrespondence to Masanobu Miyazaki, Nagasaki University School of
Medicine, The Second Department of Internal Medicine, 1-7-1 Sakamoto,
Nagasaki, 852-8501, Japan (phone +81-95-849-7273; fax +81-95-849-7285;
e-mail msnbmiya-ngs@umin.ac.jp)
In Situ Hybridization for RNA: Nonradioactive
Probe: Oligo-DNA Probe: Digoxigenin (II)
MASANOBU MIYAZAKI, YOSHIYUKI OZONO, TAKASHI HARADA
AND SHIGERU KOHNO
Introduction
Nonradioactive in situ hybridization is thought to be less sensitive than the radioactive method, especially when used with oligo-DNA probe. However, the sensitivity of nonradioactive in situ
hybridization using oligo-DNA is sufficient for localization of
mRNA at cellular level even in human biopsy specimens. The
method of nonradioactive in situ hybridization was developed
in our laboratory in 1994 (Miyazaki et al. 1994) and was applied
to identify and localize mRNA of several factors synthesized by
renal cells in normal individuals and patients with a variety of
renal diseases (Suzuki et al. 1995; Miyazaki et al. 1996). Nonradioactive in situ hybridization with oligo-DNA has several advantages compared with the radioactive method, including a
high resolution, good probe stability and no exposure to radiation, as described in detail by Koji and Nakane (1996). Compared
with other types of probes, such as RNA or eDNA probe, oligoDNA has several benefits. First, oligo-DNA is easier to prepare.
While several years ago, a special DNA synthesizer that was available only in certain laboratories, was necessary for the preparation of oligo-DNA, such synthesizers are less expensive nowadays. Alternatively, custom made probes can be purchased
nowadays from several commercial sources. Second, a special
knowledge of molecular technology and expertise, e.g., cloning
techniques, is not necessary. Finally, the probes can be easily deCorrespondence to Masanobu Miyazaki, Nagasaki University School of
Medicine, The Second Department of Internal Medicine, 1-7-1 Sakamoto,
Nagasaki, 852-8501, Japan (phone +81-95-849-7273; fax +81-95-849-7285;
e-mail msnbmiya-ngs@umin.ac.jp)