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Updated: May 20, 2026

Processing the Loblolly Pine PtGen2 cDNA Microarray
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Solid-phase PCR in a picowell array for immobilizing and arraying 100,000 PCR products to a microscope slide.

Jochen Hoffmann1, Martin Trotter, Felix von Stetten

  • 1Laboratory for MEMS Applications, Department of Microsystems Engineering - IMTEK, University of Freiburg, Georges-Koehler-Allee 103, Freiburg, 79110, Germany.

Lab on a Chip
|July 24, 2012
PubMed
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We developed a picowell array solid-phase PCR (PWA-SP-PCR) method for highly parallel DNA amplification and spatial immobilization. This technique enables digital PCR and microarray-based analysis, outperforming traditional emulsion PCR.

Area of Science:

  • Molecular Biology
  • Biotechnology
  • Genomics

Background:

  • Traditional emulsion-based PCR (emPCR) immobilizes DNA onto beads, limiting spatial analysis.
  • Picowell arrays offer high-throughput reaction volumes but require efficient product immobilization.
  • Solid-phase PCR (SP-PCR) allows covalent immobilization but scaling remains a challenge.

Purpose of the Study:

  • To develop a method for highly parallel PCR reactions with simultaneous covalent and spatially-resolved immobilization of products.
  • To demonstrate digital solid-phase PCR in a microarray format for massively parallel analysis.
  • To compare the novel PWA-SP-PCR technique with existing emPCR methods.

Main Methods:

  • Utilized picowell arrays (PWAs) with 100,000 wells cm(-2) and 19 pL reaction volumes.

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  • Performed covalent immobilization of PCR products onto a microscope slide via SP-PCR.
  • Detected immobilized DNA using streptavidin-Cy5 or direct hybridization with labeled probes.
  • Demonstrated amplification and immobilization of DNA fragments from 100 bp to 1513 bp.

    • Main Results:

    • Achieved positive signals in 97.2% of 110,000 wells.
    • Successfully amplified and immobilized single DNA molecules, demonstrating digital SP-PCR.
    • Obtained direct spatial registration of immobilized PCR products in a microarray format.
    • Showcased successful amplification and immobilization across a range of DNA template lengths.

    Conclusions:

    • PWA-SP-PCR enables highly parallel amplification and spatially resolved immobilization of PCR products.
    • The technique facilitates digital PCR and subsequent microarray-based analyses.
    • PWA-SP-PCR offers advantages over emPCR by providing direct spatial registration of amplified DNA.