Protein-based bio-products such as vaccines, antibodies, enzymes, and plasma are crucial in public health and life sciences, yet their efficacy is frequently compromised by temperature fluctuations, especially repeated freeze-thaw cycles during storage and transport. While monitoring freeze-thaw damage is critical for the quality control of these bio-products, current methods lack the capability to indicate the exact number of freeze-thaw cycles. Here, structural color liquids enable visualized freeze-thaw monitoring (FT-SCLs) are introduced by harnessing their irreversible sol-gel phase transition under repeated freeze-thaw cycles, which are constructed by assembling periodically structured poly(styrene-acrylic acid) colloidal particles within a poly(vinyl alcohol) suspension. The FT-SCLs undergo irreversible sol-gel transition and therefore unidirectional alteration of their periodic structures during freeze-thaw cycling, imparting stepwise and unrecoverable color change (from red to green) to indicate the exact number of freeze-thaw cycles. Through modulating the sol-gel transition, the FT-SCLs are constructed with adjustable sensitivity across practically relevant temperature ranges (−80–−4 °C) and customizable response thresholds for diverse application scenarios. Leveraging their unique capabilities of freeze-thaw monitoring via non-tampered optical signals, such FT-SCLs exhibit broad applicability in vaccine storage, whole blood preservation, and enzyme stability monitoring, which can further be extended for cell cryopreservation and the food industry.
This work aimed to investigate the covalent crosslinking sites induced by transglutaminase (TGase, EC 2.3.2.13) in gelatin hydrogel and their impact on the gelatin molecular conformation. LC-MS/MS results demonstrated that Lys 644 and Gln 1351 of α 1 chain and Gln 19 of α 2 chain were the main sites. The results of molecular dynamic (MD) simulation indicated that at the moderate degree of covalent crosslinking (13.55%), Radius of gyration (Rg) values decreased from 11.5 to 10.0 nm at 4 °C and led to a more compact structure, therefore enhancing structural tightness and gel strength. As the degree of crosslinking rose, the number of crosslinking sites gradually increased, with a predominant distribution in the C-terminal pro-peptide of gelatin chain. At the high crosslinking degree (37.48%), Rg value decreased at 100 °C and gelatin chains were connected through a “head to tail” linkage configuration, resulting in a significant reduction in the triple helix-like structure. The structure maintained the gel-forming ability at high temperature while lowering the aggregation energy of water molecules from −2.359 × 10 5 to −2.455 × 10 5 kJ/mol, leading to thermal irreversibility of gelatin hydrogel.
The development of nanozymes with minimized color interference and target-specific recognition capability represents a critical advancement in colorimetric test strip. Here, white nanozymes are facilely synthesized by coordination polymerization. The optimization of surfactant modification endowed these nanozymes with enhanced alkaline phosphatase (ALP)-like activity by a larger specific surface area, and electrostatic interactions. Notably, the ALP-like activity of the nanozyme can be specifically inhibited by emamectin benzoate (EMB) rather than other pesticides. A reversible uncompetitive inhibition mechanism is systematically investigated at both enzymological and molecular levels, establishing a framework for studying nanozyme-small molecule interactions. Further, an EMB test strip is developed. Crucially, the whiteness of nanozyme effectively minimized nanozyme-induced background interference and shows a whitening effect on the paper substrate of test strip, significantly enhancing the visibility of colorimetric signal. Due to EMB-specific recognition, high ALP-like activity, and the whiteness of nanozyme, the developed nanozyme-based test strip demonstrated excellent selectivity, high sensitivity, and reliable performance for the real-time detection in real samples. This work not only demonstrates the seamless integration of white nanozymes with colorimetric test strips, but also provides a promising solution for developing enzyme-free, visual, and rapid detection platforms by screening highly specific nanozyme recognition elements for commercial pesticides.
危险品化学品经营许可证（带存储）