用原位杂交组化方法探测肾上腺中合成前脑啡肽原的mRNA

DETECT mRNAs ENCODING PREPROENKEPHALIN A IN THE ADRENAL GLANDS USING THE TECHNIQUE OF IN SITU HYBRIDIZATION HISTOCHEMISTRY

取经Zambonis’固定液固定的豚鼠肾上腺作冰冻切片.切片经预杂交后,用含生物素标记的前脑啡肽原A(PPA)的cDNA探针的杂交液孵育,然后孵于链霉抗生物素-辣根过氧化物酶复合物溶液中,用DAB呈色.结果:约90％的肾上腺髓质细胞呈阳性反应,阳性反应产物位于细胞质中。

cDNA probes encoding preproenkephalin A (PPA) were used to detect the mRNA codings within the sections of adrenal glands of guinea pigs, processing according to the following procedures; perfused the animals with Zambonis' or 4% paraformaldehyde fixative; rinsed the block of adrenal glands for 12 hours in O.1mol/L phosphate sodium solution; the block were cut at -15℃ in a cryostat; Then the sections were prehybridized and hybridized with the biotinylated cDNA encoding PPA. The sections were incubated in the solution containing streptavi- din-biotinylated HRP and chromostained with DAB. The positive reactive produ- cts appeared in the cytoplasm of about 90% of adrenal medullary cells.