Solid-phase silica-based extraction leads to underestimation of residual DNA in decellularized tissues

Abstract

Decellularization of animal tissues is a novel route to obtain biomaterials for use in tissue engineering and organ transplantation. Successful decellularization is required as animal DNA causes inflammatory reactions and contains endogenous retroviruses, which could be transmitted to the patient. One of the criteria for successful decellularization is digestion (fragmentation) and elimination (residual quantity) of DNA from the tissue. Quantification of DNA can be done in many ways, but it has recently been shown that silica-based solid-phase extraction methods often do not completely purify in particular small DNA fragments. In the context of decellularization, this means that the measured DNA amount is underestimated, which could compromise safety of the processed tissue for in-patient use. In this article, we review DNA quantification methods used by researchers and assess their influence on the reported DNA contents after decellularization. We find that underestimation of residual DNA amount after silica-based solid-phase extraction may be as large as a factor of ten. We therefore recommend a direct assessment of DNA amount in tissue lysate using dsDNA-specific binding dyes, such as Picogreen, due to their higher accuracy for small fragment detection as well as ease of use and widespread availability.