15th January 2019
Aptamers have a great potential in cancer biomarker discovery via selection against cancer cells following counter selection against healthy cells. Aptamers were identified to bind to cancer specific biomarkers on cells and then subsequently isolated and analysed.
Prostate cancer cells were lysed and incubated with streptavidin magnetic beads pre-immobilized with a biotinylated single stranded aptamer. The aptamer loaded beads were then used to ‘pull-down’ the captured target. After washing the beads and collecting unbound material, biomarker that was bound by the immobilised aptamers, was eluted. This was performed alongside a negative control sample with aptamer-free streptavidin beads. Recovered fractions of unbound material, final wash and elution were analysed by SDS-PAGE, and visualised by sliver staining.
Figure 1: Sliver stained SDS-PAGE gel showing the binding and elution of the unknown biomarker expressed by prostate cancer cells. For both aptamer positive (right of Ladder) and aptamer negative beads (left of Ladder), unbound, final washes and elutions were collected. The streptavidin band is highlighted by a black arrow and the unknown biomarker in red.
The positive elution shows the presence of an additional protein with an approximate molecular weight of 37kDa, which was not visible in the initial cell lysate. Following, the biomarker was recovered from the gel by extraction and analysed by mass spectrometry (data not shown). Aptamer Group has successfully demonstrated specific aptamer development to prostate cancer biomarkers, and subsequent lysate capture and elution utilizing standard techniques comparable to immunoprecipitation.
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