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. 2022 May 18;8(20):eabn7382. doi: 10.1126/sciadv.abn7382

Fig. 3. Hierarchical self-uncloaking property of RNCOs in vitro.

Fig. 3.

(A) Cas13a RNP activities of NCs and nNCs at pH 7.4 and 5.4 via the trans-cleavage of F-R-Q as fluorescent signal (λex 492 nm, λem 518 nm). (B) Fluorescence spectra analysis of Cas13a RNP recognition specificity via the trans-cleavage of F-R-Q as fluorescent signal (λex 492 nm, λem 518 nm). (C) Fluorescence spectra analysis of Cas13a/crRNA responding to T (from i to vi: 0, 50, 100, 200, 300, and 400 nM). (D) DOX release from RNCOs and nRNCOs at pH 7.4 and 5.4. (i) RNCOs + T (pH 5.4), (ii) nRNCOs + T (pH 5.4), (iii) RNCOs + mT (pH 7.4), (iv) RNCOs + T (pH 7.4), and (v) nRNCOs + T (pH 7.4). Data are means ± SD (n = 3). (E) TEM characterization of hierarchical self-uncloaking of RNCOs. Scale bars, 100 nm. (F) PAGE analysis of RNCO dissociation efficiency. (G) Schematic illustration of multisite trans-cleavage of RNSs (left) and DOX release profiles of RNCOs with incremental NC payloads (right) (from i to iv: 100 nM, 500 nM, 1 μM, and 2 μM).