Single-molecule Pcr in a Picowell Array Simultaniously Immobilizing Pcr Products to a Pdms Coverslide

For the first time, we amplified single DNA-molecules (“Digital PCR”) randomly distributed in a picowell array and simultaneously immobilized the generated PCR-products to the surface of a PDMS coverslide (“solid-phase PCR”) which was used as sealing of the picowells during PCR. First, by this unprecedented technique PCR-products can be recovered for both, further reactions/analysis and counting the number of initial DNA-molecules from digital signals with a good signal to noise ratio of 10. Second, our experiments demonstrate the currently smallest low-volume on-chip PCR in an array of 18.5 pL wells. This may enable single-cell PCR experiments by filling PCR microreactors with truly single-cells due to geometrical constriction.