Fig. 2: Validation of the CRISPR system and off-wearable strategy targeting EBV cfDNA on a solid commercial microelectrode. | Nature Communications

Fig. 2: Validation of the CRISPR system and off-wearable strategy targeting EBV cfDNA on a solid commercial microelectrode.

From: Programmable CRISPR-Cas9 microneedle patch for long-term capture and real-time monitoring of universal cell-free DNA

Fig. 2

a In vitro cleavage ability of dRNP validated by polyacrylamide gel electrophoresis (PAGE), S and P refer to the sample and product, respectively. b Next-generation sequencing of CRISPR-Cas9 gene editing in EBV cfDNA. c CRISPR-Cas9 system representative real-time i–t curve raw data for detecting EBV cfDNA targets (sample interval 0.1 s, sampling time 1200 s); the red line represents the fitting curve (polynomial order = 3). d The current signal output of the CRISPR-Cas9 system in the presence of EBV cfDNA, analyzed by two-way ANOVA, p value = 0.038, n = 3 independent experiments, data presented as mean values ± standard deviation (SD). e CV plots and f EIS spectra of the microelectrode under different conditions. I, II, III, and IV refer to the bare microelectrode, graphene-modified microelectrode, CRISPR microelectrode, and CRISPR microelectrode targeting 2 × 10−10 M EBV cfDNA, respectively, using 0.05 M [Fe(CN)6]3−/4− as the probe. g Specificity of the CRISPR-based microelectrode; 100 times of interferences showed a slight influence, 0.05 M [Fe(CN)6]3−/4− as the probe, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, analyzed by two-way ANOVA, p value of 0.0000025, 0.0000018, 0.000017 for WENV, JPEV, DENV respectively, data presented as mean values ± SD, n = 3 independent experiments. h Real-time CRISPR-based microelectrode I response targeting variable concentrations of EBV cfDNA. Regions 1, 2, 3, and 4 refer to the phase before the time threshold, the phase after the time threshold, the stable period, and the rinsing step, respectively. i Slope of the plots from (h). j Calibration curve of the real-time I response from (h), data presented as mean values ± SD, n = 3 independent experiments. k The interaction of dRNP and EBV cfDNA. l UV-vis absorption changes; inset: calibration curve, dRNP with EBV cfDNA; I, II, III, and IV: R0.1, R0.25, R0.5, and R1.0, respectively, data presented as mean values ± SD, n = 3 replicated measurements.

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