E. Mezey et al., POTENTIAL PROBLEMS IN USING [S-35] DATP-TAILED OLIGONUCLEOTIDES FOR DETECTING MESSENGER-RNAS IN CERTAIN CELLS OF THE IMMUNE-SYSTEM, The Journal of histochemistry and cytochemistry, 42(9), 1994, pp. 1277-1283
In this study we examined the cause of unusually intense signals obtai
ned in immune cells by in situ hybridization histochemistry using S-35
-labeled oligonucleotides. We verified that the phenomenon is an ampli
fication of a specific signal due to a series of chemical interactions
after the probe binds to a specific mRNA in the tissue. The presence
of oxidative enzymes in the tissue seems to be necessary for this reac
tion to occur. Therefore, most cells of the immune system (e.g., macro
phages, neutrophil and eosinophil leukocytes), being rich in oxidative
enzymes, will show some signal amplification. The intensification of
the signal can be avoided if MgCl2 is substituted for CoCl2 in the syn
thesis of [S-35]-thiophosphate-labeled probes, if 2,3-dimercaptopropan
ol [British anti-Lewisite (BAL)] is added to the hybridization buffer,
or if [P-33]-phosphate is used instead of [S-35]-thiophosphate in the
labeling of the probes.