A sensitive non-radioactive in situ hybridization method for the detection of chicken IgG gamma-chain mRNA: a technique suitable for detecting of variety of mRNAs in tissue sections.
We established a sensitive non-radioactive in situ hybridization (ISH) method for the detection of chicken IgG gamma-chain mRNA in paraffin sections. RNA probes were transcribed in vitro from cloned chicken IgG CH1 nucleotide sequences with SP6/T7 RNA polymerases in the presence of DIG-UTP. These probes were used for hybridization and were immunodetected using anti-DIG antibodies conjugated to horseradish peroxidase. The immunoreactive products were visualized with DAB-H(2)O(2). IgG gamma-chain mRNA-expressing cells were localized in both the spleen and oviductal tissues. This method demonstrated an excellent sensitivity since the ISH signal was clear and the background was negligible. We found that in the spleen IgG gamma-chain mRNA-expressing cells were present mainly in the red pulp, whereas in the oviduct they appeared mainly in the mucosal stroma and not in the mucosal epithelium.
Graduate School for International Development and Cooperation. Hiroshima University, Higashi-Hiroshima, 739-8528. Japan.Faculty of Applied Biological Science. Hiroshima University, Higashi-Hiroshima, 739-8528. Japan. yyosimu@hiroshima-u.ac.jp
Weiming
W
Zheng
Junichi
J
Izaki
Shuichi
S
Furusawa
Yukinori
Y
Yoshimura