Polyacrylamide gel electrophoresis of RNA compared with polyclonal- and monoclonal-antibody-based enzyme immunoassays for rotavirus.
Polyacrylamide gel electrophoresis (PAGE) of rotaviral RNA, a sensitive and highly specific test for detecting rotavirus in stool, was compared with two commercially available enzyme immunoassays (EIAs), monoclonal (Pathfinder) and polyclonal (Rotazyme II). Stool samples from 204 children with nosocomial diarrhea were tested for rotavirus by both EIAs and by PAGE of RNA extracted from raw stools or 10% stool suspensions. Samples which tested positive by either EIA but were negative by PAGE were subjected to blocking EIA with rabbit or goat anti-SA11. Rotavirus was detected by PAGE and Pathfinder in 62 stools, but only 47 of these were positive by Rotazyme II. Blocking assays of EIA-positive, PAGE-negative samples suggested the presence of rotavirus in four additional stools. Sensitivity and specificity measured against PAGE and blocking assays were: Pathfinder, 0.985 and 0.934; and Rotazyme II, 0.731 and 0.927, respectively. False-positive rates were 0.134 for Pathfinder and 0.149 for Rotazyme II. The specificity and rate of false-positive results of Pathfinder were improved by using an adjusted optical density cutoff 4.36 times greater than that recommended by the manufacturer (specificity, 0.993; sensitivity, 0.985; false-positive rate, 0.015).
Pacini
D L
DL
Department of Pediatrics, College of Medicine, Ohio State University, Columbus 43210.
Brady
M T
MT
Budde
C T
CT
Connell
M J
MJ
Hamparian
V V
VV
Hughes
J H
JH
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral / Q000032:analysis; D002648:Child; D003428:Cross Infection / Q000175:diagnosis*; D003967:Diarrhea / Q000175:diagnosis; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007124:Immunoenzyme Techniques; D011237:Predictive Value of Tests; D011446:Prospective Studies; D012367:RNA, Viral / Q000032:analysis*; D011933:Reagent Kits, Diagnostic; D012401:Rotavirus / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D012400:Rotavirus Infections / Q000175:diagnosis*
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D012367:RNA, Viral; D011933:Reagent Kits, Diagnostic