Optimization of CRISPR-Based Viral Diagnostics Using Quantum Nanoparticle Perovskites
2022 Annual Meeting
Optimization of CRISPR-Based Viral Diagnostics Using Quantum Nanoparticle Perovskites
CRISPR-based viral diagnostics uses CRISPR-Cas13 engineered complex and an additional cleavage reporter molecule that is made of a quencher molecule bound via a short, single-stranded RNA molecule to an organic dye. Upon cleavage of this RNA strand, the organic dye (now no longer bound to the quencher molecule) emits photoluminescence. This photoluminescence is then measured as a metric of whether the sample of cells contained the target viral sequence. The most recent advancement in this technology was the design of CARMEN (Combinatorial Arrayed Reactions for Multiplexed Evaluation of Nucleic Acids). CARMEN tests multiple combinations of infected cell samples against multiple CRISPR-Cas13 target sequences in a microwell array without user instrumentation. The principle of a solution-based fluorescent color code remains as the identifier of the viral sequence in each respective infected cell sample. Thus, my researchâs objective is to optimize this multiplexed viral detection in CARMEN.
In CARMENâs solution-based fluorescent color code, the number of pairwise combinations of droplets in the microwell array is a combination of three organic dyes used in detection. To increase the combinatorial array of these fluorescent color combinations, a type of nanoparticle called perovskites are introduced. For this research, the color codes are based on inorganic-organic metal halide perovskites, which have four unit cells (each with a Pb core cation and stabilizing anions) that form an octahedral structure with a Methyl-Ammonium cation center. The electronic bandgap determines the emission wavelength of the perovskite and tuning the anions used in perovskite synthesis produces a range of emission in the visible light. Changing the anion composition produces a range of photoluminescent Methylammonium Lead perovskites. Currently, we are working on designing encapsulation protocols to use these perovskites in aqueous solution with CRISPR-Cas13 and DNA samples.
