Lab in a Tube: Purification, Amplification, and Detection of DNA Using Poly(2‐oxazoline) Multilayers

Abstract

Fast and easy purifi cation and amplifi cation of DNA are prerequisites for the development of point-of-care diagnostics. For this reason covalent coatings of amine containing poly(2-oxazoline)s (POx) on glass and poly(propylene) surfaces are prepared, to reversibly bind genetic material directly from biological samples. The polymer is deposited in a layer-by-layer process, whereas initial immobilization of macromolecules on the surface is accomplished by the use of an epoxy functionalized siloxane monolayer. Alternating treatment with polymer and cross-linker leads to the construction of amine containing POx multilayers on the substrates. Successful deposition is investigated by confocal laser scanning microscopy (using labeled polymers), contact angle measurements, as well as refl ectometric interference spectroscopy. The interaction of these layer systems with DNA regarding binding and temperature dependent release is studied using labeled genetic material. Finally, polymerase chain reaction (PCR) vessels are coated with POx layers on the inside, and used for quantitative realtime PCR (qPCR) experiments. It is possible to bind genetic material directly from cell lysates to perform qPCR assays from surface adsorbed DNA within the same tube including amplifi cation, as well as detection. The presented system displays an easy to use device for a point of care diagnostic.

Cationic ring-opening polymerization of protected oxazolidine imines resulting in gradient copolymers of poly(2-oxazoline) and poly(urea)

“…Additionally, the formation of micelles as drug carriers, [51] nanogels [44] and surface coatings for bio-analytics [52] supports the increasing importance of POx.…”

Comparison of random and gradient amino functionalized poly(2‐oxazoline)s: Can the transfection efficiency be tuned by the macromolecular structure?

“…Therefore, the investigated polymers are usable for the purification of genetic material or drug delivery applications, however, not for gene delivery by using the investigated conditions. Future studies might concentrate on alternative copolymers with differences in the hydrophobicity and buffer capacity to identify ideal carriers for genetic material, which can ensure biocompatibility while expressing higher transfection efficiencies.…”

Advances in point-of-care nucleic acid extraction technologies for rapid diagnosis of human and plant diseases

“…After the addition of samples to the testing device, the results are displayed within a few minutes. In POC testing, various detection techniques such as nucleic acid testing ( Batule et al, 2020 ; Leiske et al, 2015 ), lateral flow assays (LFA) ( Fang et al, 2014 ; R. H. Tang et al, 2017 ), nanomaterial-based sensors ( Li et al, 2020 ; Liu et al, 2014 ; Ngo et al, 2018 ; Padmavathy et al, 2012 ), colorimetric immunosensors ( Ren et al, 2017 ), volatile organic compound sensors (Z. Li et al, 2019a , Li et al, 2019b ), bio-optical sensors ( Jin et al, 2018 ; Yoo and Lee, 2016 ), and electrochemical sensors ( Dutta et al, 2018 ; W. Liu et al, 2018 ) have been applied for the rapid detection of a broad range of human and plant diseases.…”

“…Several on-chip cell lysis techniques such as chemical lysis ( Ma et al, 2019 ; Yoon et al, 2018 ), mechanical lysis (J. Choi et al, 2015 ; Mahalanabis et al, 2009 ), electrical lysis ( Hügle et al, 2018 ; Kim et al, 2009 ; Nan et al, 2014 ), ultrasonic lysis ( Branch et al, 2017 ), thermal lysis ( Leiske et al, 2015 ; Wang et al, 2011 ; Zhang et al, 2016 ), and enzymatic lysis ( Lounsbury et al, 2013 ; Petralia et al, 2013 ) have been demonstrated for rapid lysis of human and pathogen cells.…”