Rh. Chen et Sv. Fuggle, IN-SITU CDNA POLYMERASE CHAIN-REACTION A NOVEL TECHNIQUE FOR DETECTING MESSENGER-RNA EXPRESSION, The American journal of pathology, 143(6), 1993, pp. 1527-1534
We report a novel method for detecting intracellular messenger RNA by
combining the techniques of in situ hybridization and polymerase chain
reaction (PCR) (in situ CDNA PCR). The technique could detect low abu
ndancy signals and distinguish different levels of gene expression. We
examined the expression of the functional markers of activated cytoto
xic T lymphocytes, granzyme A, and perforin in human lymphocytes from
in vitro cultures. The amplification products were found in the cells
and the supernatants, with the distribution critically affected by the
protease digestion conditions. The specificity of amplification was c
onfirmed by electrophoretic analysis and Southern blotting. We conclud
e that the in situ CDNA PCR technique offers a sensitive method of mea
suring the frequency of signal-expressing cells and has significant re
search and clinical applications.