Adaptation of a radioactive L. donovani complex DNA probe to a chemiluminescent detection system gives enhanced sensitivity for diagnostic and epidemiological applications

Publisher Website
Google Scholar
Abstract
The cDNA probe, Lmet2, was labelled with digoxigenin and used in a chemiluminescent system to detect fewer than 100 membrane-immobilized Leishmania parasites. The probe was found to hybridize primarily with members of the L. donovani complex but a slight cross-reaction was also observed with greater than 5 x 104L. major. This cross-reaction was reduced by hybridizations in 50% formamide at 37 °C. Formamide also significantly reduced non-specific binding of the digoxigenin-labelled probe to the membrane support which, in hybridizations without formamide, masked the specific hybridization signal. This background was not observed with the corresponding radio-isotope labelled probe. With hybridizations in formamide the sensitivity achieved by the chemiluminescent system after exposure to film for 3 h was greater than that achieved by the isotopic system even after autoradiography for 24 h.