ABSTRACT
A novel nucleic acid hybridization assay with a DNA probe immobilized on 1.25-micron-diameter latex particles was developed. Hybridization of the immobilized probe DNA with sample rRNA was complete in 10 to 15 min. Alkaline phosphatase-labeled anti-DNA-RNA was allowed to bind to the DNA-RNA hybrids on the latex particles. Then the latex was collected on a small glass fiber filter pad, and bound alkaline phosphatase was quantitated by reflectance rate measurement. The method detected a broad range of bacterial species and had a detection limit of 500 cells per assay. The assay was used to screen urine samples for bacteriuria and had a sensitivity of 96.2% compared with conventional culture at a decision level of greater than or equal to 10(4) CFU/ml. The hybridization method could have broad application to the detection of bacteria and viruses.
Subject(s)
Bacteria/isolation & purification , Bacteriuria/diagnosis , DNA, Bacterial/genetics , RNA, Bacterial/analysis , RNA, Ribosomal/analysis , Bacteria/genetics , Chemical Phenomena , Chemistry , DNA, Bacterial/biosynthesis , Escherichia coli/genetics , Humans , Microspheres , Nucleic Acid Hybridization , Predictive Value of TestsABSTRACT
A radioimmunoadsorbent assay for the detection of hepatitis B surface antigen (HBsAG) is described. The method uses small DEAE-cellulose columns to adsorb HBsAg from serum or plasma samples, and it uses 125 I-labeled antibody to HBsAg to detect the adsorbed antigen. A single 2-h incubation at 45 degrees C is used in the procedure. The sensitivity of the method was determined with the Bureau of Biologics HBsAg reference panels and was shown to be equivalent to other third-generation test methods. The specificity of the method was evaluated by testing specimens from blood donors, dialysis patients, and hospital staff in parallel with commercial radioimmunoassay kits for HBsAg. In the tests performed on 2,868 blood donor specimens, 6 positive specimens were identified by both the radioimmunoadsorbent method and the commercial radioimmunoassay kits. These positive specimens were confirmed by a counterimmunoelectrophoresis procedure. One nonconfirmable, repeatably positive specimen was observed with the radioimmunoadsorbent method. In testing 1,250 specimens from dialysis patients and hospital staff, 120 confirmable positive specimens were identified by both the radioimmunoadsorbent method and a commercial test. Overall, the false positive rate for the radioimmunoadsorbent method was found to be less than 1%.
