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. 2012 Aug 22;7(8):e43533. doi: 10.1371/journal.pone.0043533

Figure 6. Pull down assay with T. cruzi trypomastigotes from spiked plasma samples.

Figure 6

Streptavidin beads, loaded with Apt68 or scrambled aptamer (Apt68 Sc) prepared in SELEX buffer were incubated with a suspension of T. cruzi trypomastigotes, prepared in human plasma, at a concentration of 100 parasites/ml or 10 parasites/ml. The parasite-bead aggregates were then concentrated using a magnet and washed with SELEX buffer. DNA was extracted using the Direct Lysis Buffer (Sigma, Mo.) and aliquots used for real time PCR. The data plotted is a representative of 3 independent experiments. Mean Ct values are plotted on the y-axis with error bars representing the standard deviation of 7 replicates. There was a significant difference (p<0.001) between the amount of parasite DNA amplified from the samples, when Apt68 coated beads were used, compared to the scrambled aptamer coated beads, low Ct values indicating higher DNA concentration. This trend is observed at both concentrations of the spiked parasites, 100 parasites/ml and 10 parasites/ml. Negative controls, where nuclease free water was used for amplification, showed Ct >40.