Fig.3. Working conditions optimization of S-iELISA.
(A-B) Coating antigen concentrations and serum sample dilutions. (C-D)
Second antibody dilutions and sealing times. (E-F) Sealing buffers and
the incubation times of antigen and serum. (G-H) The incubation times of
second antibody and exposure times of TMB.
Fig.4. Determination of the
cut-off value, repeatability and specificity of S-iELISA.
(A) Forty SADS-CoV-negative serum samples were tested using the S-iELISA
and the mean OD450nm value of SADS-CoV-negative serum
samples plus three standard deviations (SDs) was used to calculated the
cut-off value. SADS-CoV-positive serum as control. (B) Six control serum
samples (5 IFA-positive samples and 1 IFA-negative samples) were tested
using the S-iELISA and the OD450nm values of serum
samples were used to calculate the coefficient of variation (CV) to
determine the intra- and inter-assay reproducibility. (C) Porcine
positive sera against SADS-CoV, PEDV, PDCoV, TGEV, PCV2, CSFV, PRV,
FMDV, PRRSV, and ASFV were tested using the S-iELISA and the mean
OD450nm value was calculated to determine whether the
sample was positive or negative based on cut-off value. SPF pig serum as
control.