Fig 1.
Examples of candidate biomarkers.
(A) Scatterplot showing the discrimination of the early infection and control groups using m/z 3720 at F6ISL (Fraction 6, IMAC chip, Low laser intensity). Blue dots: control. Red dotes: early stage of infection. (B) Scatterplot showing the discrimination of the acute and chronic groups using m/z 13407.2 at F6CSH (Fraction 6, CM10 chip, High laser intensity). Blue dots: acute. Red dotes: chronic. (C) Example of a candidate biomarker increased in infected mice over time. Serum SELDI-TOF MS mass spectra obtained for F6ISL (Fraction 6, IMAC chip, Low laser intensity) from infected mice (top 3 spectra) versus non-infected mice (bottom spectrum). A candidate biomarker as 7081 Da is gradually up-regulated from 3, to 6, to 12 weeks after infection (P value 0.006). (D) Example of a candidate biomarker increased in infected mice regardless the parasite burden. Serum SELDI-TOF MS mass spectra obtained for F6CSL (Fraction 6, CM10 chip, Low laser intensity) from infected mice (bottom 4 spectra) versus non-infected mice (top spectrum). A candidate biomarker as 5566.31 Da is up regulated in infected mice dependent on the infection rather than the dose of infected agents (P value 0.006). G1 infected with 200 cercariae, G2 infected with 150 cercariae, G3 infected with 100 cercariae, G5 infected with 50 cercariae.
Fig 2.
Biomarker pattern software based on CART analysis.
A nonparametric procedure, was used to generate candidate diagnostic algorithms. The CART based on a series of binary decision trees that recursively partition a data set into blocks of predicted positive and negative samples. The CART procedure pursues to minimize a cost function that balances prediction errors in false-positive or false-negative results as well as the total number of biomarkers used. An example of decision tree classification using infected (G1) vs controls is shown. In this algorithm, the intensities of the 46106 Da biomarker establish the splitting rules. The samples have an intensity of ≤0.423 are placed in the left daughter node, and samples that have an intensity of ≥0.423 go to the right daughter node. Terminal red boxes = uninfected; blue boxes = acute infection.
Table 1.
Eight proteins were increased in the sera of mice during both the acute and chronic stages of schistosomiasis compared to controls.
Table 2.
Shows these proteins ordered by Spectrum Mill; generally the higher abundant identified proteins.
Table 3.
Overlap between the three methods.
This table shows the number of peaks detected in serum by MALDI and the overlap between them and the similar peaks detected by the other two methods Orbitrap and SELDI.
Fig 3.
Immunologic validation of A1AT as a candidate biomarker.
(A) Representative Western blot of A1AT in pooled sera from S. mansoni-infected mice (6 and 12 weeks post-infection) and controls. (B) Ponceau stained gel and (C) Western blot of GAPDH served as a loading controls. (D) Relative density of A1AT proteins levels normalized to GAPDH.
Fig 4.
Immunologic validation of transferrin as a candidate biomarker.
(A) Representative Western blot of transferrin in pooled sera from S. mansoni-infected mice (6 and 12 weeks post-infection) and controls. (B) Ponceau stained gel and (C) Western blot of GAPDH served as a loading controls. (D) Relative density of transferrin proteins levels normalized to GAPDH.
Fig 5.
Immunologic validation of GST as a candidate biomarker.
(A) Representative Western blot of transferrin in pooled sera from S. mansoni-infected mice (3, 6 and 12 weeks post-infection as well as sample from 6 weeks post infection from different study) and controls. (B) Western blot of GAPDH served as a loading controls. (C) Relative density of GST proteins levels normalized to GAPDH.
Fig 6.
Immunologic validation of actin as a candidate biomarker.
(A) Representative Western blot of actin in pooled sera from S. mansoni-infected mice (6 and 12 weeks post-infection) and controls. (B) Ponceau stained gel and (C) Western blot of GAPDH served as a loading controls. (D) Relative density of actin proteins levels normalized to GAPDH.