Evaluation of the biotinylated (Blugene) vs 32P-labeled cDNA probes of beta-glucocerebrosidase: relative sensitivities in genomic and other systems
Strasberg, P.
Clinical Chemistry 35(7): 1512-1516
1989
ISSN/ISBN: 0009-9147 PMID: 2758598 Document Number: 342738
The sensitivity, rapidity, and ease of use of biotinylated (Blugene, Bethesda Research Laboratories) and 32P cDNA probes have been compared, the probe being the cDNA for beta-glucocerebrosidase (EC 3.1.2.45). With the Blugene kit I could detect 2 pg of biotinylated DNA on dot blots. However, under conditions of hybridization, the lower limit of detection for unlabeled cDNA (transblotted onto nitrocellulose) by its labeled counterpart was 5000-fold smaller (10 pg vs 50 ng) for the isotopically labeled probe. 32P- and Blugene-probes hybridized detectably with 0.5 and 10 micrograms, respectively, of transblotted EcoR 1-digested genomic DNA, making the radioactive method 20 times as sensitive. However, color development was complete within 30 min to 3 h, whereas radioautoradiography required 12 h to one week. Blugene was also safer, easy to use, and effective under appropriate conditions. The 32P method is expensive, hazardous, time-consuming, and technically difficult. This nonisotopic procedure represents a desirable improvement in biotechnology.