Small Oligonucleotides Detection in Three-Dimensional Polymer Network of DNA-PEG Hydrogels

Mazzarotta, Alessia; Caputo, Tania Mariastella; Raiola, Luca; Battista, Edmondo; Netti, Paolo Antonio; Causa, Filippo

doi:10.3390/gels7030090

Cited by 5 publications

(3 citation statements)

References 57 publications

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“…To remove non-cross-linked DNA, we then washed the gel with aqueous buffer for 12 h using the washing protocol detailed in Dorsey et al The change in fluorescence of the hydrogel when comparing the gel directly after photopatterning to the same gel after the 12-h wash indicated a cross-linking efficiency of DNA of 55 ± 3.9% ( n = 3 gels, mean ± s.d.). This cross-linking efficiency is consistent with similar photopatterning processes − (SI Section S7 and Figure S11). We thus assumed half of the Payloads would remain conjugated to the hydrogel after photopolymerization in subsequent experiments.…”

supporting

confidence: 84%

Sequential Activation of Spatially Localized Oligonucleotides

Rubanov

Dorsey

Scalise

et al. 2022

ACS Materials Lett.

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The programmable spatiotemporal activation of molecules could be used to create stimuli-responsive biomaterials capable of executing multistage sensing and computational processes. Here we develop a method for the activation of different DNA molecules in precisely specified locations and times within hydrogels. The activation locations are determined by patterning molecules to be released in their inactive form within hydrogels at resolutions of tens of microns, whereas the time of activation is controlled by a DNA strand displacement reaction cascade that releases the target DNA oligonucleotides at precisely specified time intervals. This programmed activation of DNA from hydrogel substrates thus enables the scalable development of DNA-based reaction-diffusion systems that regulate DNA strand availability in space and time. This system could be used as a platform to autonomously activate predefined chemical signals embedded within a soft material.

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supporting

confidence: 84%

Sequential Activation of Spatially Localized Oligonucleotides

Rubanov

Dorsey

Scalise

et al. 2022

ACS Materials Lett.

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“…Higher molecular weight dextran at a higher concentration experiences much slower self-diffusion of dextran entanglements and creates an obstruction effect on the smaller solutes within the droplet [29,30]. With dextran acting as a physical obstacle, diffusion of mannitol and AdHu5 within the bulk droplet is hindered and may explain why AdHu5 is less likely to partition on the particle surface.…”

Section: Predicted Component Distribution In Spray Dried Particlesmentioning

confidence: 99%

Dextran Mass Ratio Controls Particle Drying Dynamics in a Thermally Stable Dry Powder Vaccine for Pulmonary Delivery

et al. 2022

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Purpose Thermally stable, spray dried vaccines targeting respiratory diseases are promising candidates for pulmonary delivery, requiring careful excipient formulation to effectively encapsulate and protect labile biologics. This study investigates the impact of dextran mass ratio and molecular weight on activity retention, thermal stability and aerosol behaviour of a labile adenoviral vector (AdHu5) encapsulated within a spray dried mannitol-dextran blend. Methods Comparing formulations using 40 kDa or 500 kDa dextran at mass ratios of 1:3 and 3:1 mannitol to dextran, in vitro quantification of activity losses and powder flowability was used to assess suitability for inhalation. Results Incorporating mannitol in a 1:3 ratio with 500 kDa dextran reduced viral titre processing losses below 0.5 log and displayed strong thermal stability under accelerated aging conditions. Moisture absorption and agglomeration was higher in dextran-rich formulations, but under low humidity the 1:3 ratio with 500 kDa dextran powder had the lowest mass median aerodynamic diameter (4.4 µm) and 84% emitted dose from an intratracheal dosator, indicating strong aerosol performance. Conclusions Overall, dextran-rich formulations increased viscosity during drying which slowed self-diffusion and favorably hindered viral partitioning at the particle surface. Reducing mannitol content also minimized AdHu5 exclusion from crystalline regions that can force the vector to air-solid interfaces where deactivation occurs. Although increased dextran molecular weight improved activity retention at the 1:3 ratio, it was less influential than the ratio parameter. Improving encapsulation ultimately allows inhalable vaccines to be prepared at higher potency, requiring less powder mass per inhaled dose and higher delivery efficiency.

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“…The tuneable network confers size exclusion ability as the polymer mesh size can be finely tuned to specifically allow diffusion and reaction of target molecules of different size, improving the assay sensitivity [22]. Moreover, it has been proved that a 3D functionalized polymer network offers not only enhanced sensitivity, but also better specificity, since probes spaced further apart hybridize less with mismatch sequences [16,23].…”

Section: Introductionmentioning

confidence: 99%

Hydrogel Microparticles for Fluorescence Detection of miRNA in Mix-Read Bioassay

Mazzarotta

Caputo

Battista

et al. 2021

Sensors

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Herein we describe the development of a mix-read bioassay based on a three-dimensional (3D) poly ethylene glycol—(PEG)-hydrogel microparticles for the detection of oligonucleotides in complex media. The key steps of hydrogels synthesis and molecular recognition in a 3D polymer network are elucidated. The design of the DNA probes and their density in polymer network were opportunely optimized. Furthermore, the diffusion into the polymer was tuned adjusting the polymer concentration and consequently the characteristic mesh size. Upon parameters optimization, 3D-PEG-hydrogels were synthetized in a microfluidic system and provided with fluorescent probe. Target detection occurred by double strand displacement assay associated to fluorescence depletion within the hydrogel microparticle. Proposed 3D-PEG-hydrogel microparticles were designed for miR-143-3p detection. Results showed 3D-hydrogel microparticles with working range comprise between 10−6–10−12 M, had limit of detection of 30 pM and good specificity. Moreover, due to the anti-fouling properties of PEG-hydrogel, the target detection occurred in human serum with performance comparable to that in buffer. Due to the approach versatility, such design could be easily adapted to other short oligonucleotides detection.

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Small Oligonucleotides Detection in Three-Dimensional Polymer Network of DNA-PEG Hydrogels

Cited by 5 publications

References 57 publications

Sequential Activation of Spatially Localized Oligonucleotides

Sequential Activation of Spatially Localized Oligonucleotides

Dextran Mass Ratio Controls Particle Drying Dynamics in a Thermally Stable Dry Powder Vaccine for Pulmonary Delivery

Hydrogel Microparticles for Fluorescence Detection of miRNA in Mix-Read Bioassay

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