Table 1.
Monoclonal antibody (mAb) library against Y. pestis LcrV and F1 antigens.
Fig 1.
Western blot analysis of anti-LcrV monoclonal antibodies (mAbs) against Yersinia pestis Harbin-35 lysate.
Horseradish peroxidase (HRP) conjugated LcrV mAbs (1 μg/mL) were used to probe (A) reduced and (B) non-reduced Y. pestis Harbin-35 bacterial lysate (1.5x106 cells/lane) by direct Western blot.
Fig 2.
Western blot analysis of anti-F1 monoclonal antibodies (mAbs) against Yersinia pestis Harbin-35 lysate.
Anti-F1 mAbs (1 μg/mL) were used to probe (A) reduced and (B) non-reduced Y. pestis Harbin-35 bacterial lysate (1.5x106 cells/lane) by indirect Western blot. HRP-conjugated goat anti-mouse Ig was used for detection of F1 mAb binding.
Table 2.
Affinity and kinetics analysis of Y. pestis mAbs by surface plasmon resonance.
Table 3.
Assay sensitivity of the top 4 mAb pairs (LcrV or F1) by lateral flow immunoassay.
Fig 3.
Sensitivity of Y. pestis lateral flow immunoassays (LFI) using recombinant LcrV and F1.
LFI prototypes were tested with recombinant (A) LcrV and (B) F1 serial diluted into pooled normal human serum ranging from 0.25 to 256 ng/mL. Assay signal was evaluated and quantitated by optical density using a Qiagen ESE reader. Intensity ≥ 20 mm*mV scores as positive.
Fig 4.
Specificity testing of dual Yersinia pestis lateral flow immunoassay (LFI) against clinically relevant bacterial panel.
The dual LFI prototype containing test lines specific for LcrV (8F10/6F10) and F1 (11C7/3F2) was tested against a panel of bacterial lysates. Bacterial lysate (50 μL at OD600 = 0.5) was applied onto the conjugate pad and chased with buffer. LFIs were imaged after 20 minutes.
Table 4.
Limit of detection of enzyme-linked immunosorbent assays using recombinant LcrV and F1 antigens in buffer.