Assays used to detect MSP1 and antibodies to MSP1.

a<p>Polyclonal anti-MSP1₄₂ was resuspended in carbonated-bicarbonated buffer (pH 9.6) and <b>used to coat the ELISA plates as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0007986#s2" target="_blank">materials and methods</a> section. As a standard, recombinant MSP1₄₂ was resuspended in the same media used for the test samples (i.e. plasma from N. American subjects or perfusates) and added to the wells coated with anti-MSP1₄₂ at serial two-fold dilutions starting at 4 ng/ml</b>. Sensitivity of the assay was approximately 200 pg/ml.</p>b<p>Plates were coated with rMSP1₄₂ in PBS.</p>c<p>Plates were coated with streptavidin in PBS (Pierce Biotechnology Inc). Standards used for this experiment were bMSP1₄₂ or pooled human plasma containing anti-MSP1 (25%) combined with bMSP1₄₂ as described in the text. The assay had a sensitivity ∼175–200 pg/ml <b>to detect the bMSP1₄₂ in the samples</b>.</p>d<p>Perfusate was added at 1∶5 dilution to wells coated with streptavidin and incubated overnight at 4°C. Plates were washed thoroughly (6 times) with 0.1% Tween as described in the text to remove any unbound antibody prior to adding goat anti-human IgG. Control experiments using MSP1₄₂ incubated with normal serum had <0.15 OD and the mean+2 SD was subtracted from background described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0007986#pone-0007986-g004" target="_blank">figure 4</a>.