A quantitative electrochemiluminescence assay for Clostridium perfringens alpha toxin.
Described is a rapid direct sandwich format electrochemiluminescence assay for identifying and assaying Clostridium perfringens alpha toxin. Biotinylated antibodies to C. perfringens alpha toxin bound to streptavidin paramagnetic beads specifically immunoadsorbed soluble sample alpha toxin which subsequently selectively immunoadsorbed ruthenium (Ru)-labeled detection antibodies. The ruthenium chelate of detection antibodies chemically reacted in the presence of tripropylamine and upon electronic stimulation emitted photons (electrochemiluminescence) that were detected by the photodiode of the detector. Elevated toxin concentrations increased toxin immunoadsorption and the specific immunoadsorption of Ru-labeled antibodies to alpha toxin, which resulted in increased dose-dependent electrochemiluminescent signals. The standardized assay was rapid (single 2.5-h coincubation of all reagents), required no wash steps, and had a sensitivity of about 1 ng/ml of toxin. The assay had excellent accuracy and precision and was validated in buffer, serum, and urine with no apparent matrix effects.
Merrill
Gerald A
GA
Department of Clinical Investigation, Brooke Army Medical Center, 3400 Rawley E. Chambers Ave, STE A, Ft. Sam Houston, TX 78234-6315, USA. Gerald.Merrill@Amedd.Army.Mil
Rivera
Victor R
VR
Neal
Dwayne D
DD
Young
Charles
C
Poli
Mark A
MA
D000906:Antibodies; D001427:Bacterial Toxins / Q000032:analysis* / Q000276:immunology; D002135:Calcium-Binding Proteins / Q000032:analysis* / Q000276:immunology; D003016:Clostridium perfringens; D004563:Electrochemistry / Q000379:methods*; D008163:Luminescent Measurements / Q000379:methods*; D008958:Models, Molecular; D012680:Sensitivity and Specificity; D010738:Type C Phospholipases / Q000032:analysis* / Q000276:immunology
D000906:Antibodies; D001427:Bacterial Toxins; D002135:Calcium-Binding Proteins; D010738:Type C Phospholipases; C040649:alpha toxin, Clostridium perfringens