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The remarkable material properties of spider silk, such as its high toughness and tensile strength combined with its low density, make it a highly sought-after material with myriad applications. In addition, the biological nature of spider silk makes it a promising, potentially sustainable alternative to many toxic or petrochemical-derived materials. Therefore, interest in the heterologous production of spider silk proteins has greatly increased over the past few decades, making recombinant spider silk an important frontier in biomanufacturing. This has resulted in a diversity of potential host organisms, a large space for sequence design, and a variety of downstream processing techniques and product applications for spider silk production. Here, we highlight advances in each of these technical aspects as well as white spaces therein, still ripe for further investigation and discovery. Additionally, industry landscaping, patent analyses, and interviews with Key Opinion Leaders help define both the research and industry landscapes. In particular, we found that though textiles dominated the early products proposed by companies, the versatile nature of spider silk has opened up possibilities in other industries, such as high-performance materials in automotive applications or biomedical therapies. While continuing enthusiasm has imbued scientists and investors alike, many technical and business considerations still remain unsolved before spider silk can be democratized as a high-performance product. We provide insights and strategies for overcoming these initial hurdles, and we highlight the importance of collaboration between academia, industry, and policy makers. Linking technical considerations to business and market entry strategies highlights the importance of a holistic approach for the effective scale-up and commercial viability of spider silk bioproduction.

Pyriform (or piriform) silk is a fibrous spider silk used in a composite material to attach web silks to substrates and disparate silk types together. Argiope argentata pyriform silk is formed from a protein with a central repetitive region consisting of a 234-amino acid unit (the "Py unit") repeated 21 times. We have previously shown that the Py unit behaves modularly, with a structured core of 6 α-helices that retains its structuring upon truncation of N- and C-terminal intrinsically disordered segments. Here, we apply solution-state nuclear magnetic resonance (NMR) spectroscopy to investigate the structured core of the Py unit. Based on 15N spin relaxation measurements and reduced spectral density mapping at two field strengths, the α-helical core of the Py unit exhibits minimal backbone motion beyond the tumbling of the globular domain except for one inter-helical linker that exhibits a notably higher degree of dynamics. Evaluation of rotational diffusion based on 15N spin relaxation and translational diffusion by pulsed field gradient diffusion NMR demonstrates that the core of the Py unit behaves as a compact structure, with hydrodynamic behaviour consistent with a well-packed protein. Pyriform silk protein undergoes a structural transformation upon conversion from the soluble-state to the fibrous-state, with a loss of α-helical content and gain of β-sheet content. The more dynamic segment we observe midway through the structured core of the Py core, especially in the context of the intrinsically disordered segments that flank the core, would provide an initiation point for decompaction, structural transformation, and/or intermolecular interactions. [ABSTRACT FROM AUTHOR]

Recent, rapid arthropod declines have been tied to human activity. Yet, the propensity for human‐produced noise to disturb the substrate‐borne (vibratory) channel remains virtually untested despite arthropods' ubiquitous use of vibratory information. Characterizing the vibratory landscape at biologically relevant scales is an essential initial step. We aimed to measure vibratory noise across space and time to assess its potential overlap with the urban–rural range and season of a funnel‐weaving spider, Agelenopsis pennsylvanica (C.L. Koch) (Araneae: Agelenidae), a common urban arthropod. We assessed 24‐h ambient vibrations (hereafter vibratory "noise") at 23 sites across urban–rural Lincoln, Nebraska (NE, USA), during the adult season of A. pennsylvanica (August–October). Vibratory noise (amplitude, Leq) varied by ~15 dB across sites overlapping with frequencies within A. pennsylvanica's sensitivity (20–1000 Hz). Urban vibratory noise was positively correlated with the principal component containing estimates of traffic‐induced vibrations (e.g., traffic volume, road proximity, and impervious cover), whereas rural noise levels correlated with probable harvest times. Our findings indicate spatial variation in noise in urban areas and seasonal noise variability in rural areas. We also tested how A. pennsylvanica—collected from four distinct noise survey sites—use their vibratory microhabitat. We assessed daily spider position and dry silk mass within a microcosm that played loud and quiet vibrations (white noise differing by ~15 dB) in separate but connected chambers. Age affected the chamber choice (hereafter "microhabitat use") of spiders collected from the loudest site, as younger adults associated with and used more silk in the loud chamber, but this effect decreased with age. As our data demonstrate that vibratory noise varies with A. pennsylvanica's microhabitat and season with a high potential for impacting behavior, we hypothesize that this environmental noise likely impacts other arthropods as well. [ABSTRACT FROM AUTHOR]

Silk‐based materials are sought after across various industries due to their remarkable properties, including high strength and flexibility. However, their practical application depends largely on how well these properties are maintained under different environmental conditions. Despite significant advancements in the large‐scale production of artificial silk fibers, the effects of temperature on their mechanical behavior are understudied. In this study, the mechanical properties of artificial spider silk fibers between −80 and +120 °C are examined and compared to both synthetic and natural silk fibers. The findings reveal that artificial silk fibers maintain their strength up to +120 °C, though the strain at break slightly decreases, remaining above 60%. At −80 °C, the fibers exhibit increased strength, but the strain at break is reduced. While these artificial fibers closely mimic the behavior of natural silk, they show a noticeable reduction in extensibility at low temperatures. Complementing experimental data, differential scanning calorimetry, and thermogravimetric analysis are also conducted, proposing a simple physical model to explain the observed temperature‐induced softening. Encouragingly, the degradation temperature of artificial silk is comparable to that of native silkworm and spider silk. This study underscores the importance of enhancing the mechanical robustness of artificial silk to expand its applications. [ABSTRACT FROM AUTHOR]

Before humans and allegedly any animal group, spiders developed "functionally graded toothed blades" that cut one of the toughest biological materials: silk. Here, this work reveals the importance of micro‐structured serrations in spiders' fangs that allow these animals to cut silk and artificial high‐performance fibers, such as carbon or Kevlar. The importance of serrations revolves around the stress concentration at the interface between the fang and the fibers, resulting in a cutting efficiency superior to that of a razor blade. This efficiency is increased by the presence of pretension in the fibers and is high also for fibers with different diameters like silk, because of the serration grading that allows a smart positioning of the fiber in the optimal cutting condition. This work proposes that when the silk fiber is grasped by the fang, it slides along the serrated edge till it gets locked in the serration with a comparable size, where the load to cut is minimal. These results provide a new perspective on cutting mechanisms and set the roots for spider fang‐inspired cutting tools. [ABSTRACT FROM AUTHOR]

In many animals, early-life decisions influence long-term fitness. Mantidflies are spider egg predators; their tiny larvae climb aboard spiders to find eggs, but little is known about how they find spiders. We tested the hypothesis that mantidfly larvae (Dicromantispa sayi) detect and respond to substrate-borne spider cues (e.g., silk and/or excreta). We presented larvae with filter paper exposed to different types of spiders (adult female, adult male, or juvenile Habronattus trimaculatus jumping spiders) versus a no-cue control. Larvae spent more time on filter paper with spider cues. We then tested the hypothesis that mantidflies make finer distinctions between spiders when given direct choices between these cues. Larvae did not discriminate between sexes but spent more time (and exhibited more phoretic behavior) on filter paper with female or male cues compared with juvenile cues. While this suggests that mantidflies actively seek out adult spiders, we also found that adult spiders were more active than juveniles and may have simply deposited more silk and excreta, providing a stronger cue to detect. We discuss these findings in the context of the risks and benefits of different spider hosts, and how early-life spider-searching strategies may shape a mantidfly's long-term fitness. [ABSTRACT FROM AUTHOR]

Liquid-liquid phase separation (LLPS) of proteins can be considered an intermediate solubility regime between disperse solutions and solid fibers. While LLPS has been described for several pathogenic amyloids, recent evidence suggests that it is similarly relevant for functional amyloids. Here, we review the evidence that links spider silk proteins (spidroins) and LLPS and its role in the spinning process. Major ampullate spidroins undergo LLPS mediated by stickers and spacers in their repeat regions. During spinning, the spidroins droplets shift from liquid to crystalline states. Shear force, altered ion composition, and pH changes cause micelle-like spidroin assemblies to form an increasingly ordered liquid-crystalline phase. Interactions between polyalanine regions in the repeat regions ultimately yield the characteristic β-crystalline structure of mature dragline silk fibers. Based on these findings, we hypothesize that liquid-liquid crystalline phase separation (LLCPS) can describe the molecular and macroscopic features of the phase transitions of major ampullate spidroins during spinning and speculate whether other silk types may use a similar mechanism to convert from liquid dope to solid fiber. Liquid-liquid phase separation (LLPS) of proteins can be considered an intermediate solubility regime between disperse solutions and solid fibers, relevant to both pathogenic and functional amyloids. Here, the authors review the evidence that links spider silk proteins (spidroins) and LLPS and its role in the spinning process. [ABSTRACT FROM AUTHOR]

Bagworm silk proteins, which contain crystalline structures with large β‐nanocrystal sizes, are ideal candidates for biosynthetic high‐performance fibers. However, the extremely high glycine content and greater molecular weight limit their heterologous expression efficiency and further application exploration. Here, a multi‐module assembly strategy is developed to engineer novel chimeric structural proteins by incorporating the mechanical functional domains of bagworm silk proteins with the C‐terminal self‐assembly domains of spider silk proteins. By selecting a single repetitive unit of the functional region of bagworm silk proteins, the challenge of low heterologous expression efficiency is successfully addressed. Furthermore, the content and ordering of β‐sheet structure are enhanced in the chimeric proteins through the alignment mediated by the spider silk C‐terminal domain and ligation facilitated by split inteins, resulting in a remarkable Young's modulus of ≈15 GPa. This surpasses many artificial protein fibers, synthetic polymer fibers, and even natural spider silk. Notably, these protein fibers are drawn into surgical sutures and demonstrate superior wound healing effects compared to clinical suture in a skin wound model. This research presents a novel strategy for developing high‐performance protein fibers, which will expand the scope of their mechanically demanding applications. [ABSTRACT FROM AUTHOR]

The thermal transport properties of polymer fibers are important for heat dissipation in apparel, matrix of wearable devices, and so on. However, it is difficult to precisely determine the thermal properties of a single polymer fiber because of non-negligible thermal radiation due to its relatively low thermal conductivity and high surface-area-to-volume ratio. Here, we developed a system in which the apparent thermal diffusivity and size of microfibers can be measured to estimate their intrinsic thermal diffusivity. We determined the thermal diffusivities of three fibrous materials: silk, spider silk, and cellulose microfibers to be 4.2 (± 0.8) × 10 − 7 , 1.8 (± 0.7) × 10 − 7 , and 4.7 (± 0.5) × 10 − 7 , respectively. For all fibers, the apparent thermal diffusivity strongly depended on the fiber size, indicating that eliminating the radiation effect is indispensable for determining the thermal transport properties of polymer microfibers. [ABSTRACT FROM AUTHOR]

Articulus wound infection is a threat to human health. Existing medical materials have poor biocompatibility and may contain harmful chemicals, causing allergies and secondary infections. Therefore, there is an urgent need to develop innovative medical materials. Materials made of artificial spider silk proteins have been widely used in wound healing because of their good biocompatibility, biodegradability, cell adhesion and bioelectronic properties. [ABSTRACT FROM AUTHOR]

This article extensively explores various bio-inspired techniques of interfacial strengthening. Bio-inspired composites fabricated by emulating five natural materials: nacre, byssus thread, plant roots, bone, and spider silk have been discussed. A detailed account of the strengthening mechanisms active in these natural materials responsible for improving fiber/matrix adhesion in artificial composites is presented. Dendritic growth on fiber surfaces is crucial in enhancing the mechanical properties via interlocking in root-inspired composites. Micro-waviness, nano-asperity interlocking, shear deformation, and mineral bridge fracture are the main strengthening mechanisms active in the nacre-inspired composites. Byssal threads have exceptional adhesive properties due to the sacrificial bonds, hidden length, and strong metal coordination cross-links. The typical ‘Bouligand’ microstructure of bone has inspired different composite materials with excellent crack deflection and arresting. Spider silk fibers have a highly oriented proteinous structure surrounded by a semi-amorphous matrix. Their skin-core structure provides tensile strength and toughness to these fibers, inspiring various artificial composites. This article explores additive manufacturing and chemical modification as the implementation techniques for bio-inspired strengthening. Applications of bio-inspired composites include biomedical, marine, aerospace, and tissue engineering. Challenges, such as optimal mechanical properties, sustainability, simulation, and integration with Industry 5.0, must be addressed systematically to augment their applicability. [ABSTRACT FROM AUTHOR]

ABSTRACT Spiders are a hyperdiverse taxon and among the most abundant predators in nearly all terrestrial habitats. Their success is often attributed to key developments in their evolution such as silk and venom production and major apomorphies such as a whole‐genome duplication. Resolving deep relationships within the spider tree of life has been historically challenging, making it difficult to measure the relative importance of these novelties for spider evolution. Whole‐genome data offer an essential resource in these efforts, but also for functional genomic studies. Here, we present de novo assemblies for three spider species: Ryuthela nishihirai (Liphistiidae), a representative of the ancient Mesothelae, the suborder that is sister to all other extant spiders; Uloborus plumipes (Uloboridae), a cribellate orbweaver whose phylogenetic placement is especially challenging; and Cheiracanthium punctorium (Cheiracanthiidae), which represents only the second family to be sequenced in the hyperdiverse Dionycha clade. These genomes fill critical gaps in the spider tree of life. Using these novel genomes along with 25 previously published ones, we examine the evolutionary history of spidroin gene and structural hox cluster diversity. Our assemblies provide critical genomic resources to facilitate deeper investigations into spider evolution. The near chromosome‐level genome of the ‘living fossil’ R. nishihirai represents an especially important step forward, offering new insights into the origins of spider traits. [ABSTRACT FROM AUTHOR]

Natural materials possess inherent multilength‐scale structures, showcasing outstanding mechanical properties such as strain‐stiffening, high damping, and shape‐memory features under ambient conditions. Such integrated properties are highly desirable for advanced materials in biomedical devices and soft robots but remain challenging in synthetic materials. Herein, a novel strategy of sequential ion treatment is employed to introduce micro/nanoscale‐ordered structures and molecular‐scale stimulus responses into hydrogel‐derived self‐assembly fibers under ambient conditions. The treated fibers exhibit strain‐stiffening properties with a high toughness of 257 MJ m−3 and 73% damping capacity comparable to spider silk. Owing to their cross‐linking network and reversible secondary structure, those fibers exhibit outstanding water‐stability, wet stretchability, and hydration‐responsive shape‐memory performance, with ultimate elongation of 407%, shape‐fixity ratio of 94%, and shape‐recovery rate of 97%. This work furthers the green fabrication of smart materials with multifunction and presents promising applications in diverse fields, including biomimetics and biomedicine. [ABSTRACT FROM AUTHOR]

Cribellate silks, produced by ancient spiders, are fascinating because they feature a highly sophisticated, 3D hierarchical structure consisting of filaments with different diameters and shapes. Here, the smallest and thinnest constituents of the cribellate silk are investigated: nanofibrils that form a dense mesh that is supported by larger fibers. Analysis of their structure via atomic force and transmission electron microscopies shows that they are flattened fibrils, only ≈5 nm thick — thinner than any other natural spider silk fibrils previously reported. In this work, the first mechanical tensile testing experiments on these fibrils are carried out, which reveals that the fibrils show an outstanding extensibility of at least 1100%, almost twice as much as the most stretchable spider silk previously reported. Based on these extraordinary findings, this work significantly expands the parameter space of materials properties attainable by spider silks and provides further insights into their nanomechanics. [ABSTRACT FROM AUTHOR]

Nature has evolved a myriad of dedicate‐designed structures that assist in prey capture. Spiders employ their web structures for vibrational sensing and prey capture, analogous to a component of a neurosensory system. Here, we propose a spider web‐inspired sensor with integrated energy‐absorption and force perception capabilities by 3D printing. The conductive carbon black/TPU composite made from TPU granules and carbon black particles are employed for 3D printing. We analyze the underlying sensing mechanism of conductive CB/TPU composites stemming from the enhanced dispersion of the CB particles. Through 3D printing method, we are allowed to regulate the structure's form, filling density, arrangement, and printing path to tailor the static mechanical properties, initial resistance, and force sensitivity of the spider‐web inspired structure. The static mechanical model for single silk of the spider‐web inspired structure is constructed. We explore the energy absorption capability of spider‐web inspired structures with different structural parameters, and demonstrate its impact perceiving characteristics at different falling height. Additionally, we assembled a circuit to visually reflect the falling height of the impacting ball through the brightness of the LED light. This study provides insights to endowing structural materials with intelligent characteristics, contributing to their practical applications in the fields of biomedical, soft robotics, and artificial skin, etc. Highlights: A spider web‐inspired sensor is 3D printed with CB/TPU composites.The sensor has integrated force perception and energy‐absorption capabilities.The sensitivity capabilities can be regulated by parameter design.The mechanical model of the bioinspired structure is constructed.Its impact perceiving characteristics are demonstrated by two applications. [ABSTRACT FROM AUTHOR]

High-strength biodegradable polymer composites have potential applications in a variety of biomedical applications. This study investigates the influence of silk fiber on the properties of the commonly used biodegradable polylactic acid-based composites, focusing on mechanical, rheological, morphological, and degradation characteristics. Mechanical tests revealed that the addition of 2.5 wt% silk fibers enhanced the ductility of PLA composites, increasing tensile strain at break from 7.39% for pure PLA to 11.51% for the composite. However, higher silk contents (≥10 wt%) resulted in lower elongation at breaks but higher moduli, indicating a trade-off between flexibility and the structural rigidity of the composite. Rheological tests demonstrated that the presence of silk fibers up to 7.5% improved the storage modulus, reflecting better network formation within the PLA matrix. Scanning Electron Microscopy (SEM) photomicrographs illustrated improved fiber dispersion, while higher contents introduced voids and stress concentrations, adversely affecting mechanical properties. Degradation tests in phosphate-buffered saline at 37 °C showed that silk additions slowed PLA degradation, suggesting controlled degradation suitable for biomedical applications. The optimal silk fiber content for balancing mechanical integrity and flexibility was identified to be ca 7.5 wt%, providing insights into the design of PLA/silk composites for enhanced performance in practical applications. [ABSTRACT FROM AUTHOR]

The cosmetics industry is undergoing a shift towards sustainability and efficacy, driven by consumer demand for eco-friendly and safe products. This paper introduces SVX, a spider silk-inspired raw material intended to transform cosmetic formulations. Produced through fermentation, SVX is a biopolymer composed of self-assembled proteins characterized by a porous structure for delivering active ingredients safely to the skin. The study utilized in vitro and ex vivo methods to assess SVX's ability to protect against oxidative stress, enhance skin hydration, and support ingredient delivery. Safety assays, including the HET-CAM, patch test, and HRIPT, demonstrated that SVX is non-irritating and safe for topical application. Additionally, FTIR analysis confirmed SVX's capacity for sustained release of active ingredients, such as hyaluronic acid, over an 8 h period. Results showed that SVX significantly improved skin barrier protection and exhibited superior antioxidant properties compared to control formulations. Its biocompatibility, along with a vegan and biodegradable composition, aligns with the principles of sustainability, with over 60% biodegradability achieved within 10 days. Furthermore, SVX displayed antioxidant efficacy approximately 130 times greater than L-ascorbic acid, based on DPPH assay results. These findings suggest that SVX offers a versatile and sustainable solution for skincare formulations, combining environmental responsibility with benefits for skin health and performance. [ABSTRACT FROM AUTHOR]

This review focuses on the innovative use of laser techniques in developing and functionalizing biomimetic surfaces, emphasizing their potential applications in the medical and biological fields. Drawing inspiration from the remarkable properties of various natural systems, such as the water-repellent lotus leaf, the adhesive gecko foot, the strong yet lightweight spider silk, and the unique optical structures of insect wings, we explore the potential for replicating these features through advanced laser surface modifications. Depending on the nature and architecture of the surface, particular techniques have been designed and developed. We present an in-depth analysis of various methodologies, including laser ablation/evaporation techniques, such as Pulsed Laser Deposition and Matrix-Assisted Pulsed Laser Evaporation, and approaches for laser surface structuring, including two-photon lithography, direct laser interference patterning, laser-induced periodic surface structures, direct laser writing, laser-induced forward transfer, and femtosecond laser ablation of metals in organic solvents. Additionally, specific applications are highlighted with the aim of synthesizing this knowledge and outlining future directions for research that further explore the intersection of laser techniques and biomimetic surfaces, paving the way for advancements in biomedical applications. [ABSTRACT FROM AUTHOR]

Spiders produce webs, which are still a largely unexplored source of antibacterial compounds, although the reports of its application in the medical field. Therefore, this study aims to present an integrative review of the antibacterial activity of spider webs. The research was conducted using Google Scholar, Scielo, Web of Science, PubMed, ScienceDirect, Medline EBSCO, LILACS, and Embase. The inclusion criteria were original articles written in English that studied the antibiotic properties of the web or isolated compounds tested. The studies were compared according to the spider species studied, the type of web, treatment of the sample, type of antimicrobial test, and the results obtained. Nine hundred and seventy-three publications were found, and after applying the inclusion and exclusion criteria, sixteen articles were selected. Bacterial inhibition was found in seven studies against various species of bacteria such as Escherichia coli, Klebsiella pneumoniae, Staphylococcus aureus, Salmonella Typhi, Bacillus megaterium, Listeria monocytogenes, Acinetobacter baumannii, Streptococcus pneumoniae, Pasteurella multocida, and Bacillus subtilis. Additionally, there was no apparent relationship between the proximity of the spider species evaluated in the studies and the presence or absence of activity. Methodological problems detected may affected the reproducibility and reliability of the results in some studies, such as the lack of description of the web or microorganism strain, as well as the absence of adequate controls and treatments to sterilize the sample. Spider webs can be a valuable source of antibiotics; however, more studies are needed to confirm the real activity of the web or components involved. [ABSTRACT FROM AUTHOR]

Abstract In many animals, early-life decisions influence long-term fitness. Mantidflies are spider egg predators; their tiny larvae climb aboard spiders to find eggs, but little is known about how they find spiders. We tested the hypothesis that mantidfly larvae (Dicromantispa sayi) detect and respond to substrate-borne spider cues (e.g., silk and/or excreta). We presented larvae with filter paper exposed to different types of spiders (adult female, adult male, or juvenile Habronattus trimaculatus jumping spiders) versus a no-cue control. Larvae spent more time on filter paper with spider cues. We then tested the hypothesis that mantidflies make finer distinctions between spiders when given direct choices between these cues. Larvae did not discriminate between sexes but spent more time (and exhibited more phoretic behavior) on filter paper with female or male cues compared with juvenile cues. While this suggests that mantidflies actively seek out adult spiders, we also found that adult spiders were more active than juveniles and may have simply deposited more silk and excreta, providing a stronger cue to detect. We discuss these findings in the context of the risks and benefits of different spider hosts, and how early-life spider-searching strategies may shape a mantidfly’s long-term fitness.

The availability of appropriate conduits remains an obstacle for successful reconstruction of long-distance nerve defects. In previous sheep trials, we were able to bridge 6 cm nerve gaps with nerve conduits based on spider silk fibers with full functional outcomes. Here, we describe the first application of spider silk for nerve repair in humans.

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Spun silk is found only in arthropods, where it plays critical roles in feeding, protection, and reproduction. It has evolved repeatedly within the arthropods, providing an opportunity for comparative analyses to reveal the mechanisms driving convergent evolution within this taxon. Silk in crustaceans has received much less attention than silk in arachnids and insects but may have unique and useful properties. We combine data on the morphology of silk and silk systems with ancestral state reconstruction and find that silk has likely evolved convergently at least six times within Crustacea. We also break down the components of crustacean silk as a composite trait by discussing the silk properties, the silk production systems, and the silk genes. We find that amphipod and tanaid silks are similar to silkworm and spider silks but have their own unique silk system morphology. Much remains unknown about the origins and diversification of silk in crustaceans; we propose that the molecular investigation of silk genes and systems will provide a path forward. By investigating the evolution of silk production in crustaceans, we may discover insights into predispositions for silk production in arthropods and valuable substrates for biomaterial research. [ABSTRACT FROM AUTHOR]

Hydrogels are useful drug release systems and tissue engineering scaffolds. However, synthetic hydrogels often require harsh gelation conditions and can contain toxic by‐products while naturally derived hydrogels can transmit pathogens and in general have poor mechanical properties. Thus, there is a need for a hydrogel that forms under ambient conditions, is non‐toxic, xeno‐free, and has good mechanical properties. A recombinant spider silk protein‐derived hydrogel that rapidly forms at 37 °C is recently developed. The temperature and gelation times are well‐suited for an injectable in situ polymerising hydrogel, as well as a 3D cell culture scaffold. Here, it is shown that the diffusion rate and the mechanical properties can be tuned by changing the protein concentration and that human fetal mesenchymal stem cells encapsulated in the hydrogels show high survival and viability. Furthermore, mixtures of recombinant spider silk proteins and green fluorescent protein (GFP) form gels from which functional GFP is gradually released, indicating that bioactive molecules are easily included in the gels, maintain activity and can diffuse through the gel. Interestingly, encapsulated ARPE‐19 cells are viable and continuously produce the growth factor progranulin, which is detected in the cell culture medium over the study period of 31 days. [ABSTRACT FROM AUTHOR]

Artificial spider silk is an attractive material for many technical applications since it is a biobased fiber that can be produced under ambient conditions but still outcompetes synthetic fibers (e.g., Kevlar) in terms of toughness. Industrial use of this material requires bulk‐scale production of recombinant spider silk proteins in heterologous host and replication of the pristine fiber's mechanical properties. High molecular weight spider silk proteins can be spun into fibers with impressive mechanical properties, but the production levels are too low to allow commercialization of the material. Small spider silk proteins, on the other hand, can be produced at yields that are compatible with industrial use, but the mechanical properties of such fibers need to be improved. Here, the literature on wet‐spinning of artificial spider silk fibers is summarized and analyzed with a focus on mechanical performance. Furthermore, several strategies for how to improve the properties of such fibers, including optimized protein composition, smarter spinning setups, innovative protein engineering, chemical and physical crosslinking as well as the incorporation of nanomaterials in composite fibers, are outlined and discussed. [ABSTRACT FROM AUTHOR]

Over the past two decades, significant advancements have been made in the scalable production and commercialization of microbially‐produced recombinant protein polymers. This perspective presents the evolution from early research efforts to the development of market‐ready products, with a focus on recombinant silk‐like proteins. Initial attempts to synthesize spider silk proteins in microbial hosts faced challenges with solubility, stability, and yield. Recent advancements in synthetic biology, protein engineering, and bioprocess development have enabled the substantial progress on these challenges. Early commercial efforts highlight the complexities and high costs involved in silk production and more recent strategies have shifted toward processes with better scalability, techno‐economics, and product properties. Significant commercial progress has been made, with products launched in textiles and personal care. Although market penetration is limited so far, substantial groundwork is laid for future success. Key challenges remain, such as continued high production costs and the need for cost‐effective purification and fiber spinning techniques. However, the convergence of scientific, technological, and market developments –including a growing number of product launches – suggests that recombinant silk and protein polymers can soon become widespread sustainable materials across various industries. [ABSTRACT FROM AUTHOR]

Spider dragline silk's exceptional mechanical performance, coupled with its benign production pathway, have inspired the design of diverse smart materials. Remarkably, relatively little is known about the self‐assembly process of major ampullate spidroin 1 (MaSp1) – the main protein constituent of dragline fiber – during its rapid conversion from soluble precursor protein into hierarchically structured mature silk fibers. Herein, the biomimetic potential of MaSp1 is explored using a new recombinant platform with full domain representation. MaSp1 is found to undergo efficient liquid‐liquid phase separation (LLPS) in response to kosmotropic ions, with notably higher propensity compared to MaSp2, and with MaSp1 exhibiting a more complex solubility profile relative to levels of sodium chloride. To further the understanding of silk spinning, the rapid assembly of mesoscale structures is monitored in real‐time in response to ion and pH gradients, revealing a progression from LLPS droplets toward aligned hierarchical fibers. Furthermore, using this biomimetic system, insoluble macroscopic MaSp1 fibers can be readily generated, wherein the application of mechanical deformation triggers the emergence of β‐sheet secondary structures. The insights gained from this study can have broader application in efforts to mimic the formation of biomaterials with spatially organized features across multiple length scales. [ABSTRACT FROM AUTHOR]

Spider silk is one of the strongest and toughest fibrous materials available in nature. Its superior properties make it a good candidate for applications in various fields ranging from protective body armor to scaffolds for medical implants. However, harvesting a substantial amount of natural spider silk remains challenging because spiders cannot be easily bred. With the development of synthetic biology and its integration with materials science, considerable research has been directed toward engineering and production of synthetic spider silk materials. Here the study overviews general strategies on molecular design, mesoscopic assembly, and macroscopic regulation of artificial spider silk materials. The insights into the correlation between silk protein sequences, mesoscopic assemblies, and macroscopic material properties are provided for guiding de novo design and engineering of next‐generation spider silk materials. This review also emphasizes the challenges and future perspectives for advancing the translational research on these designer functional materials for diverse applications. [ABSTRACT FROM AUTHOR]

Hydration‐induced shape‐morphing behavior has been discovered in many natural fiber‐based materials, yet this smart behavior in regenerated fibers from biopolymers lacks investigation. Here, hierarchically structured silk fibers are developed with anisotropic long‐range molecular organization and water‐responsive effects resembling natural spider silk. The regenerated silk fibers exhibit the water‐triggered shape‐memory effect and a water‐driven cyclic response. The reversible hydrogen bonds and transformation in the metastable secondary structure from α‐helices/random coils to β‐sheets are explored as the mechanisms responsible for the water‐responsiveness. The silk fibers obtained possess a tensile strength higher than 104 MPa at a fracture strain of ≈100%, showing noticeable toughness. The water‐responsive silk fibers exhibit a shape recovery rate of ∼83% and generate a maximum actuation stress of up to 18 MPa during the water‐driven cyclic contraction that outperforms most traditional natural textile fibers. The regenerated silk fibers show potential for use in water‐driven actuators, artificial muscle, and smart fabrics based on the integration of suitable mechanical properties and water responsiveness. [ABSTRACT FROM AUTHOR]

All animal species, from arthropods to vertebrates, must deal with occasional stressors in their lives, though most research on this has been focused on vertebrates. Meanwhile, our understanding of stress reactions in arthropod species like spiders is nascent. In the United States, a non‐native orb‐weaving spider, Trichonephila clavata (‘jorō’ spider), is spreading as is its already‐established cousin in the United States, T. clavipes (golden silk spider). Prior study has revealed how these two species have a unique behavioural reaction to physical stressors, whereby they remain in a thanatosis state for a prolonged period compared with other species. Here, we investigate the physiological stress reactions of these Trichonephila spiders by evaluating how each species' dorsal vessel contractions (heart rates) become elevated after being subjected to a non‐lethal stressor. For comparison, we also evaluate two similarly‐sized orb weavers, Argiope aurantia (garden spider) and A. trifasciata (banded garden spider). We record baseline heart rates of inactive, resting, spiders in our lab, then restrain them under an electronic, ‘optocardiographic’, sensor for 10 min, to record their ‘stressed’ heart rates. Argiope aurantia has a pronounced heart rate elevation, for reasons unknown. We observe that all spider heart rates increase during restraint, though each has a species‐specific pattern of elevation over time under restraint. Notably, heart rates of both Trichonephila spiders are less variable under stress, since they tend not to struggle during restraint. Meanwhile, both Argiope spiders frequently struggle, leading to marked fluctuations in cardiac output. The stress reactions of Trichonephila spiders could be characterized as ‘even‐tempered’, which may factor into their ability to live in habitats with frequent disturbances. [ABSTRACT FROM AUTHOR]

The Plateau–Rayleigh instability (PRI) of a liquid column, which always spontaneously breaks into droplets to minimize surface energy, underlies a variety of fascinating phenomena in daily life and industrial applications. Different from the free liquid column, the evolution of the liquid film on a fiber involves solid‐liquid interfaces which allow for regulating the PRI. Recently, natural fibers with certain topologies have witnessed various dynamic liquid behaviors from droplet manipulation to continuous liquid films. Tailoring the topology of local curved liquid film by the structural‐confinement is the key to manipulating liquids, where the asymmetric Laplace pressure generated by the asymmetric/non‐spherical liquid film promotes the liquid motion. In nature, the spider silk collects droplets efficiently by the spindle‐knot structure; while the mulberry silk enables a smooth surface‐coating on dual‐parallel fibers. Drawing inspirations, many artificial structured fibers are developed to precisely regulate liquid behaviors in the form of either separated droplets or continuous liquid films, which have demonstrated applications as fluidic coating, micro‐patterning, and materials fabrication. Here, recent research progress on bioinspired fibrous PRI from the viewpoints of tailoring the Laplace pressure by rationally designing the surface topology, which offers inspiration for liquid manipulation using open fibrous media, is reviewed. [ABSTRACT FROM AUTHOR]

Multifunctional fibers represent a cornerstone of human civilization, playing a pivotal role in numerous aspects of societal development. Natural biomaterials, in contrast to synthetic alternatives, offer environmental sustainability, biocompatibility, and biodegradability. Among these biomaterials, natural silk is favored in biomedical applications and smart fiber technology due to its accessibility, superior mechanical properties, diverse functional groups, controllable structure, and exceptional biocompatibility. This review delves into the intricate structure and properties of natural silk fibers and their extensive applications in biomedicine and smart fiber technology. It highlights the critical significance of silk fibers in the development of multifunctional materials, emphasizing their mechanical strength, biocompatibility, and biodegradability. A detailed analysis of the hierarchical structure of silk fibers elucidates how these structural features contribute to their unique properties. The review also encompasses the biomedical applications of silk fibers, including surgical sutures, tissue engineering, and drug delivery systems, along with recent advancements in smart fiber applications such as sensing, optical technologies, and energy storage. The enhancement of functional properties of silk fibers through chemical or physical modifications is discussed, suggesting broader high-end applications. Additionally, the review addresses current challenges and future directions in the application of silk fibers in biomedicine and smart fiber technologies, underscoring silk's potential in driving contemporary technological innovations. The versatility and sustainability of silk fibers position them as pivotal elements in contemporary materials science and technology, fostering the development of next-generation smart materials. [ABSTRACT FROM AUTHOR]

Silk fibroin is highly susceptible to damage during the extraction process. To address this issue, this study proposes an integrated degumming solution that uses a composite system of Na2CO3 and the surfactant SDS (sodium dodecyl sulfate), combined with ultrasonic treatment. Silk fibroin fibers treated with this method exhibit a smoother and more even surface, with markedly less fiber damage. Compared to the traditional 100 °C alkaline degumming (30 min complete degumming), this scheme completes degumming in only 10 min at 75 °C. Nanofibrous membranes were prepared from these degummed fibers using electrospinning technology, and their β-sheet crystalline content increased to 49.58% after ethanol treatment, along with high tensile strength (5.27 MPa) and elongation at break (123.49%). Over a 5-day cell culture period, the membranes have supported cell adhesion, spreading and proliferation, demonstrating excellent cytocompatibility. These research findings provide a novel solution for the production of high-performance silk fibroin nanofibrous membranes and are expected to advance the application and development of silk fibroin materials in the biomedical field. [ABSTRACT FROM AUTHOR]

Collagen is one main component of the extracellular matrix (ECM) in natural tissues and is, therefore, well suited as a biomaterial for tissue engineering. In this study, a method is presented to 3D‐bioprint collagen into a precipitation bath comprising recombinantly produced spider silk protein eADF4(C16) yielding a composite with excellent mechanical properties. The spider silk precipitation bath induced assembly of the collagen into fibrils, and subsequent addition of potassium phosphate buffer lead to the formation of silk particles and stabilization of the collagen fibrils. The produced collagen‐silk composite scaffolds show an internal structure of homogeneously distributed and interacting collagen fibrils and spider silk particles with significantly better mechanical properties compared to plain collagen scaffolds. Further, enzymatic degradation assays of the scaffolds over a 7‐day period show higher stability of the collagen‐silk scaffolds compared to plain collagen scaffolds in the presence of wound proteases. Using the spider silk variant eADF4(C16‐RGD) further increases compressive stress and elastic modulus compared to that of the unmodified variant. Finally, it is shown that the unique collagen‐spider silk composite scaffolds comprising the cell‐binding domains of collagen and the RGD sequence in the spider silk variant represent a promising material for soft tissue regeneration. [ABSTRACT FROM AUTHOR]

Despite impressive progress in the field, there are still several major bottlenecks in producing fibers from recombinantly produced spider‐silk‐like proteins to replicate the extraordinary mechanical properties of spider major ampullate silk. The conventional artificial fiber spinning processes rely primarily on organic solvents to coagulate proteins into fibers and require complex post‐treatments to obtain fibers with valuable properties. This is due to challenges in obtaining soluble silk proteins, but also because the native silk spinning process leading to the hierarchical organization of the silk proteins is not fully understood and is hard to replicate in a manner applicable to industrial settings. Here, recombinant spider‐silk fusion proteins are efficiently produced and processed into as‐spun fibers with a toughness modulus of 120 MJ m−3 and extensibility of 255% using solely aqueous solutions. The spider‐silk fusion proteins assemble in a manner similar to that reported for native spider silk: they phase separate induced by salting out, followed by alignment and a secondary structure transition triggered by shear forces and dehydration. Finally, the design of the fusion silk proteins enables straightforward functionalization of the fibers under mild all‐aqueous conditions via a simple biomolecular click reaction both pre‐ and post‐spinning. [ABSTRACT FROM AUTHOR]

Spider silk is renowned for its exceptional mechanical properties, surpassing those of other natural and many synthetic fibers. Yet, replicating its remarkable properties through synthetic production remains a challenge. The variability in the mechanical properties of synthetic spider silks lacking protective coatings, exacerbated by factors such as spinning conditions and humidity levels, poses an additional challenge, impacting their application potential. Bioconjugation offers a versatile synthetic method to modify protein structures, enhancing their pharmacokinetics, solubility, stability, and immune response. In particular, polyethylene glycol (PEG)‐ylation has emerged as a successful strategy with numerous marketed PEG–protein conjugates. This study introduces synthetic spider silk—multiarm PEG bioconjugates, facilitating spidroin crosslinking, and chemical functionalization while retaining a biomimetic spinning approach. Two different examples demonstrate the potential of this approach to improve the fiber's tensile strength and extensibility, respectively, both leading to an increased toughness modulus. Furthermore, the approach could allow the tuning of fiber mechanical properties without developing a new mini‐spidroin construct and fiber coating with lipids attached to multiarm PEG, potentially mitigating the impact of environmental conditions on synthetic spider silk fibers. [ABSTRACT FROM AUTHOR]

Silk fibroin hydrogels are extensively explored for tissue engineering and regenerative medicine as an artificial extracellular matrix (ECM) that can support tissue growth. However, the nanometer pore size of hydrogels limits adequate cell, tissue, and vascular infiltration. Microgel scaffolds are an emerging class of microporous biomaterials formed by annealing small microscale hydrogels (microgels) into a 3D construct. In this work, silk microgels are generated using a microfluidic device that allows tuning of the microgel diameter (100–400 µm) and are stabilized via visible light‐initiated photo‐crosslinking of native tyrosine residues in silk. Microgels are then covalently annealed using silk solution as glue and the same cytocompatible visible light‐initiated crosslinking to form microgel scaffolds. Unlike the nano‐porosity of bulk photo‐crosslinked silk hydrogels, the microgel scaffolds have an average pore diameter of 29 ± 17 or 192 ± 81 µm depending on the microgel size, with enhanced mechanical properties compared to bulk hydrogels. This microporosity supports enhanced cell spreading and proliferation in vitro and increases scaffold remodeling in vivo, encouraging improved tissue infiltration and matrix deposition. The microgel size and material format also affect inflammatory responses in vivo. This work demonstrates that silk microgels and microgel scaffolds are promising candidates for tissue engineering and regenerative medicine applications. [ABSTRACT FROM AUTHOR]

DNA is organized into chromatin-like structures that support the maintenance and regulation of genomes. A unique and poorly understood form of DNA organization exists in chloroplasts, which are organelles of endosymbiotic origin responsible for photosynthesis. Chloroplast genomes, together with associated proteins, form membrane-less structures known as nucleoids. The internal arrangement of the nucleoid, molecular mechanisms of DNA organization, and connections between nucleoid structure and gene expression remain mostly unknown. We show that Arabidopsis thaliana chloroplast nucleoids have a unique sequence-specific organization driven by DNA binding to the thylakoid membranes. DNA associated with the membranes has high protein occupancy, has reduced DNA accessibility, and is highly transcribed. In contrast, genes with low levels of transcription are further away from the membranes, have lower protein occupancy, and have higher DNA accessibility. Membrane association of active genes relies on the pattern of transcription and proper chloroplast development. We propose a speculative model that transcription organizes the chloroplast nucleoid into a transcriptionally active membrane-associated core and a less active periphery. [ABSTRACT FROM AUTHOR]

The prediction of protein 3D structure from amino acid sequence is a computational grand challenge in biophysics and plays a key role in robust protein structure prediction algorithms, from drug discovery to genome interpretation. The advent of AI models, such as AlphaFold, is revolutionizing applications that depend on robust protein structure prediction algorithms. To maximize the impact, and ease the usability, of these AI tools we introduce APACE, AlphaFold2 and advanced computing as a service, a computational framework that effectively handles this AI model and its TB-size database to conduct accelerated protein structure prediction analyses in modern supercomputing environments. We deployed APACE in the Delta and Polaris supercomputers and quantified its performance for accurate protein structure predictions using four exemplar proteins: 6AWO, 6OAN, 7MEZ, and 6D6U. Using up to 300 ensembles, distributed across 200 NVIDIA A100 GPUs, we found that APACE is up to two orders of magnitude faster than off-the-self AlphaFold2 implementations, reducing time-to-solution from weeks to minutes. This computational approach may be readily linked with robotics laboratories to automate and accelerate scientific discovery. [ABSTRACT FROM AUTHOR]

Proteins mediate their functions through chemical interactions; modeling these interactions, which are typically through sidechains, is an important need in protein design. However, constructing an all-atom generative model requires an appropriate scheme for managing the jointly continuous and discrete nature of proteins encoded in the structure and sequence. We describe an all-atom diffusion model of protein structure, Protpardelle, which represents all sidechain states at once as a "superposition" state; superpositions defining a protein are collapsed into individual residue types and conformations during sample generation. When combined with sequence design methods, our model is able to codesign all-atom protein structure and sequence. Generated proteins are of good quality under the typical quality, diversity, and novelty metrics, and sidechains reproduce the chemical features and behavior of natural proteins. Finally, we explore the potential of our model to conduct all-atom protein design and scaffold functional motifs in a backbone- and rotamer-free way. [ABSTRACT FROM AUTHOR]

Lightweight structural materials are commonly used as effective fillers for advanced composites with high toughness. This study focused on enhancing the toughness of direct‐spun carbon nanotube yarns (CNTYs) by controlling the micro‐textural structure using a water‐gap‐based direct spinning. Drawing inspiration from the structural features of natural spider silk fibroin, characterized by an α‐helix in the amorphous region and β‐sheet in the crystalline region, multiscale bundles within CNTYs are reorganized into a unique nano‐coil‐like structure. This nano‐coiled structure facilitated the efficient dissipation of external mechanical loads through densification with the rearrangement of multiscale bundles, improving specific strength and strain. The resulting CNTYs exhibited exceptional mechanical properties with toughness reaching 250 J g−1, making them promising alternatives to commercially available fibers in lightweight, high‐toughness applications. These findings highlight the significance of nano‐coiling engineering for emulating bio‐inspired micro‐textural structures, achieving remarkable enhancement in the toughness of CNTYs. [ABSTRACT FROM AUTHOR]

Coxsackievirus B3 (CVB3) is a member of the enterovirus genus and linked to several diseases, including myocarditis, which can progress to dilated cardiomyopathy. Despite ongoing preclinical efforts, no clinically approved vaccines against CVB3 are currently available, highlighting the urgent need for effective prophylactic solutions. In this study, a nanovaccine platform based on spider minor ampullate silk protein (MiSp) is introduced. This platform utilizes protein nanoparticles engineered from chimeric proteins that incorporate CVB3 antigenic peptides into customized MiSp, subsequently loaded with all‐trans retinoic acid (RA). These functional nanovaccines are capable of eliciting both mucosal and systemic immune responses following subcutaneous administration and demonstrate significant protective effects against CVB3 infection in mice. This study signifies an approach in peptide‐based parenteral vaccine strategies, utilizing engineered MiSp nanoparticles combined with RA. This methodology represents a promising pathway for preventing enterovirus infections by leveraging the unique immunomodulatory properties of spidroins and RA to combat these pathogens effectively. [ABSTRACT FROM AUTHOR]

Spider dragline silks exhibit remarkable mechanical properties, combining both high strength and toughness. These unique characteristics arise from the intricate structure of the silk, which requires atomic‐level information to understand its origins. 13C solid‐state NMR provides this detailed structural insight into spider dragline silk. In this review, 13C CP/MAS, 13C DD/MAS and 13C INEPT NMR spectroscopies are employed to reveal the structure of spider dragline silks together with 13C conformation‐dependent chemical shifts, 2D spin‐diffusion NMR, rotational echo double resonance, dipolar‐assisted rotational resonance, and angle‐dependent NMR. The primary structure of major ampullate of spider dragline silk consists of repeated polyalanine and a glycine‐rich regions. By analyzing the 13C conformation‐dependent chemical shifts and utilizing several solid‐state NMR techniques, it has been proposed that the glycine‐rich region primarily adopts a random coil conformation, including partially β‐sheet and β‐turn structures. This contradicts the previously suggested 31 helix conformation. On the other hand, the polyalanine region exhibits an antiparallel β‐sheet structure with staggered packing arrangements. Additionally, solid‐state NMR has also revealed the structure of fragelliform spider silk. These findings contribute to the understanding of the remarkable properties of spider dragline silks and provide insights into its atomic‐level architecture. [ABSTRACT FROM AUTHOR]

[Display omitted] Due to their superior safety and stability, solid-state electrolytes (SSEs) are a promising alternative to flammable liquid electrolytes in lithium-ion batteries. However, the poor solid–solid contact at the SSEs/electrodes interface remains a significant challenge. To address this issue, inspired by spider silk, we develop a composite polymer electrolyte (SPLZO), which is highly adhesive due to the designed rich hydrogel bond network, containing a supramolecular poly (urethane-urea) (SPU), lithium bis(trifluoromethanesulfonyl)imide (LiTFSI) and Li 6.5 La 3 Zr 1.5 Ta 0.5 O 12. The abundant hydrogen bonds mainly enabled inherently strong adhesion to ensure intimate electrolyte–electrode contact with low interfacial impedance. Besides, the soft polymer segments facilitate Li+ transport, and the hard components enhance the LiTFSI dissociation and accelerate Li+ motion, resulting in a high ionic conductivity of 1.67 × 10−4 S cm−1. The significantly improved interface contact and high ionic conductivity lead to a decent capacity and cycling performance of the fabricated solid-state lithium-ion batteries. Moreover, the designed SPLZO electrolyte exhibits remarkable deformability, and the flexible lithium-ion battery demonstrates outstanding mechanical flexibility and stability with negligible capacity loss when subjected to various dynamic deformations. This adhesive SSE design strategy opens new possibilities for promoting interfaces in solid-state batteries. [ABSTRACT FROM AUTHOR]

Efforts to develop innovative water harvesting strategies offer powerful solutions to alleviate the water crisis, especially in remote and arid areas. Inspired by the hydrophobic/hydrophilic pattern of desert beetles and water self-propulsion property of spider silks, a double-strand hydrophobic PVDF-HFP/hydrophilic PAN nanofibers yarn is proposed by electrospinning and twisting techniques. The double-strand cooperation approach allows for water deposition on hydrophobic PVDF-HFP segment and transport under the asymmetric capillary driving force of hydrophilic PAN segment, thus speeded up the aggregation and growth of droplets. The effects of the composition and the diameter ratio of the two primary yarns were studied and optimized for boosting fog collection performance. The double-strand anisotropic yarn not only provide an effective method for water harvesting, but also hold the potential to inspire innovative design concepts for fog collection materials in challenging environments. [ABSTRACT FROM AUTHOR]

Purpose: Eri silk fiber has superior thermal insulation behavior, better softness than cotton fiber. However, Eri silk's use in the commercial arena has not yet taken off. The purpose of the study is to explore the comfort properties of the fabric, which enhances the commercial acceptance of Eri silk clothing. Design/methodology/approach: In this investigation, three different single knit Eri silk structures were produced with different loop lengths and yarn counts to analyze the influence of process variables on low-stress mechanical properties. To execute the research work, Eri silk spun yarn of three different linear densities (15 tex, 20 tex, 25 tex) were chosen. Three different knitted structures were produced, such as single jersey, popcorn and cellular blister, and two different loop lengths were also selected. Findings: The cellular blister structure has shown appreciable low-stress properties next highest position was attained by the popcorn structure. Yarn fineness and loop length were significant with most of the low-stress properties. Research limitations/implications: The findings of this research will contribute to a greater awareness of Eri silk knitted fabric and its process parameters in relation to its commercial utility. Originality/value: This study was conducted to explore the influence of knit structure, loop length and yarn count on the low-stress properties of Eri silk-based thermal clothing. [ABSTRACT FROM AUTHOR]

There has been significant interest in researching droplet transport behavior on composite wetting surfaces. However, current research is primarily focused on modifying individual droplets and lacks an in‐depth investigation into high‐precision droplet storage. This study introduces a "billiard ball" droplet transport and storage platform (TSP) with differentiated areas. Within this platform, the volume of droplets stored in the area reaches a consistent threshold through droplet "scrambling," inspired by the water‐gathering behavior of spiders. The TSP involves connecting two regions of different sizes using a three‐dimensional stepped wedge angle structure. However, this connection is not seamless, leaving a 2‐mm gap between the regions. This gap is intentionally designed to enable continuous droplet transfer while preventing any static migration. Through systematic experimental and simulation analysis, we investigated the influence of superhydrophilic pattern structures and parameters on quantitative droplet storage. We established a functional relationship between the pattern area and the stored volume, and analyzed the intrinsic mechanism of droplet collision separation. This enabled us to achieve on‐demand quantitative droplet storage and autonomize the storage process. The "billiard ball" droplet transport–storage platform proposed in this study holds promising applications in the fields of biomedical and organic chemistry. [ABSTRACT FROM AUTHOR]