Table 1.

Comparison of procedures used for the two IELISAs and two CELISAsa

ParameterAssay
IAEA-IELISAADRI-IELISACELISA-OCCELISA-sLPS
MicroplateNunc PolysorbNunc 69620Nunc 69620Nunc 69620
AntigensLPSsLPSO chainsLPS
 Concn (μg/ml)1.01.02.01.0
 Buffer0.05 M CO3 0.05 M CO3 0.05 M CO3 0.05 M CO3
 Incubation temp (°C)4204 (frozen)20
 Incubation time (h)≥18≥18≥18≥18
Wash buffer0.002 M PO4, 0.15 M NaCl, 0.05% Tween 20, pH 7.40.01 M PO4, 0.15 M NaCl, 0.05% Tween 20, pH 7.20.01 M PO4, 0.15 M NaCl, 0.05% Tween 20, pH 7.20.01 M PO4, 0.15 M NaCl, 0.05% Tween 20, pH 7.2
No. of wash cycles3444
Serum diluent0.01 M PBS, 0.05% Tween 20, pH 7.4Wash buffer plus EDTA-EGTA, pH 6.3Same as wash bufferWash buffer plus EDTA-EGTA, pH 6.3
Serum (control/test)
 Serum dilution1:2001:501:501:20
 Incubation temp (°C)37202020
 Incubation time (min)603012030
 AgitationYesNo3 min3 min
Competing antibodyNANAYsT9-HRPM84
Detecting antibodyMAb to bovine IgG1-HRPMAb to bovine IgG1-HRPSame as competingGaMIgG-HRP (diluted in wash buffer)
 Incubation time (min)6030NA30
 Incubation temp (°C)3720NA20
 AgitationYesNoNANo
Substrate/chromogen
 Incubation time (min)10101010
 Incubation temp (°C)37202020
 AgitationYesYesYesYes
 Wavelength (nm)405414414414
  • a Abbreviations: PBS, phosphate-buffered saline; NA, not applicable; HRP, horseradish peroxidase; MAb, monoclonal antibody; IgG1, immunoglobulin G1; GaM, goat anti-mouse IgG-HRP.