Fig 1.
2-D gel electrophoresis, silver staining and western blotting of Toxocara canis Excretory Secretory Antigens (TES-Ag).
The TES-Ag sample was separated and analyzed using 2D gel electrophoresis and western blotting. Three of the 2DE gels were transferred to nitrocellulose membranes and probed with a strong EIA positive Toxocara human sera pool (A), a negative human serum sample (B), and Baylisascaris procyonis positive serum (C).A reference gel was stained using silver stain (D). The circled spots in D represent proteins that were excised and subjected to mass spectrometry analysis.
Table 1.
Identified proteins from mass spectrometry analysis.
Table 2.
Identified mRNA sequences from a T. canis expressed sequenced tag database.
Fig 2.
Purity and antigenicity of purified recombinant proteins.
SDS-PAGE of the three recombinant proteins. A. Tc-CTL-1; B. Tc-TES-26; C. Tc-MUC-3. The recombinant protein samples at a concentration of: 1–6.25 ng/mm; 2–3.125 ng/mm; 3–1.6 ng/mm, and 4–0.8 ng/mm was treated with SDS and heated at 65°C for 15 minutes, separated and analyzed using SDS gel electrophoresis and western blotting. Two gels were transferred to nitrocellulose membranes and probed with a strong EIA positive Toxocara human sera pool, a negative human serum sample diluted 1:100 in PBS/Tween 0.3%/5% milk, and one gel was incubated with protein staining, Colloidal Gold Total Protein Stain (Bio-Rad, Cat. # 170–6527). M = Precision Plus Protein Dual Xtra Standards (Biorad, Cat. #161–0377)
Fig 3.
ROC Curves of Tc-CTL-1 and Tc-TES-26, and optimal linear combination for visceral larval migrans.
ROC Curves of Tc-CTL-1, Tc-TES-26, and the optimal linear combination were constructed based on Luminex-derived mean fluorescence intensity antigen for visceral larval migrans from 288 negative U.S. serum samples + 134 heterologous parasitic infected serum samples and 204 visceral larval migrans positive serum samples; thresholds are calculated with the highest sum of sensitivity and specificity and are represented on the plot with black dots and corresponding 95% confidence intervals.
Table 3.
Performance of Tc-CTL-1 and Tc-TES-26 Luminex based assays.
Table 4.
Specificity of the Toxocariasis Luminex assay.