Alum/CpG adjuvant promotes immunogenicity of inactivated SARS-CoV-2 Omicron vaccine through enhanced humoral and cellular immunity.

The SARS-CoV-2 Omicron variant, which was classified as a variant of concern (VOC) by the World Health Organization on 26 November 2021, has attracted worldwide attention for its high transmissibility and immune evasion ability. The existing COVID-19 vaccine has been shown to be less effective in preventing Omicron variant infection and symptomatic infection, which brings new challenges to vaccine development and application. Here, we evaluated the immunogenicity and safety of an Omicron variant COVID-19 inactivated vaccine containing aluminum and CpG adjuvants in a variety of animal models. The results showed that the vaccine candidate could induce high levels of neutralizing antibodies against the Omicron variant virus and binding antibodies, and significantly promoted cellular immune response. Meanwhile, the vaccine candidate was safe. Therefore, it provided more foundation for the development of aluminum and CpG as a combination adjuvant in human vaccines.

Enhancing Inactivated Yellow Fever 17D Vaccine-Induced Immune Responses in Balb/C Mice Using Alum/CpG.

There are some concerns about the safety of live attenuated yellow fever vaccines (YF-live), particularly viscerotropic adverse events, which have a high mortality rate. The cellular production of the vaccine will not cause these adverse effects and has the potential to extend applicability to those who have allergic reactions, immunosuppression, and age. In this study, inactivated yellow fever (YF) was prepared and adsorbed with Alum/CpG. The cellular and humoral immunities were investigated in a mouse model. The results showed that Alum/CpG (20 µg/mL) could significantly increase the binding and neutralizing activities of the antibodies against YF. Moreover, the antibody level at day 28 after one dose was similar to that of the attenuated vaccine, but significantly higher after two doses. At the same time, Alum/CpG significantly increased the levels of IFN-γ and IL-4 cytokines.

BMP9 is a potent inducer of chondrogenesis, volumetric expansion and collagen type II accumulation in bovine auricular cartilage chondroprogenitors.

Reconstruction of the outer ear currently requires harvesting of cartilage from the posterior of the auricle or ribs leading to pain and donor site morbidity. An alternative source for auricular reconstruction is in vitro tissue engineered cartilage using stem/progenitor cells. Several candidate cell-types have been studied with tissue-specific auricular cartilage progenitor cells (AuCPC) of particular interest. Whilst chondrogenic differentiation of competent stem cells using growth factor TGFß1 produces cartilage this tissue is frequently fibrocartilaginous and lacks the morphological features of hyaline cartilage. Recent work has shown that growth factor BMP9 is a potent chondrogenic and morphogenetic factor for articular cartilage progenitor cells, and we hypothesised that this property extends to cartilage-derived progenitors from other tissues. In this study we show monoclonal populations of AuCPCs from immature and mature bovine cartilage cultured with BMP9 produced cartilage pellets have 3-5-fold greater surface area in sections than those grown with TGFß1. Increased volumetric growth using BMP9 was due to greater sGAG deposition in immature pellets and significantly greater collagen accumulation in both immature and mature progenitor pellets. Polarised light microscopy and immunohistochemical analyses revealed that the organisation of collagen fibrils within pellets is an important factor in the growth of pellets. Additionally, chondrocytes in BMP9 stimulated cell pellets had larger lacunae and were more evenly dispersed throughout the extracellular matrix. Interestingly, BMP9 tended to normalise the response of immature AuCPC monoclonal cell lines to differentiation cues whereas cells exhibited more variation under TGFß1. In conclusion, BMP9 appears to be a potent inducer of chondrogenesis and volumetric growth for AuCPCs a property that can be exploited for tissue engineering strategies for reconstructive surgery though with the caveat of negligible elastin production following 21-day treatment with either growth factor.

Iridoids and active ones in patrinia: A review.

Iridoid is a special class of monoterpenoids, whose basic skeleton is the acetal derivative of antinodilaldehyde with a bicyclic H-5/H-9ß, ß-cisfused cyclopentan pyran ring. They were often existed in Valerianaceae, Rubiaceae, Scrophulariaceae and Labiaceae family, and has various biological activities, such as anti-inflammatory, hypoglycemic, neuroprotection, and soon. In this review, iridoids from Patrinia (Valerianaceae family), and the active ones as well as their mechanisms in recent 20 years were summarized. Up to now, a total of 115 iridoids had been identified in Patrinia, among which 48 had extensive biological activities mainly presented in anti-inflammatory, anti-tumor and neuroprotective. And the mechanisms involved in MAPK, NF-κB and JNK signal pathways. The summary for iridoids and their activities will provide the evidence to exploit the iridoids in Patrinia.

HI-6-loaded PEGylated liposomes: an on-site first-aid strategy for acute organophosphorus agent poisoning.

Organophosphorus agents, also known as nerve agents, are very dangerous chemicals that were used as chemical warfare agents. HI-6 is one of the most promising reactivators which is effective in reactivating AChE inhibited by many nerve agents. However, the fast in-vivo clearance of HI-6 became a large barrier for first aid use under some sophisticated circumstances. In this study, PEGylated liposomes loading HI-6 were prepared and evaluated in vitro and in vivo. For PEG-LP-HI-6, the optimal formulation's loading efficiency and encapsulation efficiency were 6.47 ± 0.10% and 71.2 ± 1.15%, respectively. According to the pharmacokinetic results, compared with free HI-6 and LP-HI-6, the intravenous injection of PEG-LP-HI-6 significantly extended t1/2 (1.47 ± 0.29 h), MRT (1.44 ± 0.07 h), and improved the AUC of HI-6 in vivo. Drug concentrations in the CNS also increased after the intravenous administration of PEG-LP-HI-6. For in vivo treatment study, twenty minutes after poison exposure, the survival rate of animals in saline, free HI-6, LP-HI-6 and PEG-LP-HI-6 groups were 0, 0, 30% and 70%, respectively. Compared with the non-PEGylated liposomes group and free HI-6, PEG-LP-HI-6 could prolong the survival time of experimental animals and alleviate the neurotoxic symptoms, which demonstrated great potential as a first-aid strategy for acute organophosphorus agent poisoning.

Articular Cartilage Chondroprogenitors: Isolation and Directed Differentiation.

Experimental data suggests that tissue-specific progenitors are present within hyaline articular cartilage with the potential to contribute to growth, maintenance, and repair. In this chapter, we show how colony-forming progenitor-like cells can be isolated from bovine articular cartilage using differential adhesion to fibronectin. Furthermore, we describe the optimal conditions and factors required to differentiate these progenitor cells to produce hyaline articular cartilage.

Safety and immunogenicity of a combined DTacP-sIPV-Hib vaccine in animal models.

The DTacP-sIPV-Hib combination vaccine can replace the single-component acellular pertussis, diphtheria, tetanus, polio, and Haemophilus influenzae type B vaccines. In this study, we evaluated the safety and immunogenicity of a newly developed DTacP-sIPV-Hib combination vaccine in animal models. We used 40 mice and 46 cynomolgus monkeys to evaluate acute and long-term toxicity. Thirty-six guinea pigs were used for sensitization assessment. For immunogenicity assessment, 50 NIH mice and 50 rats were equally randomized to receive 3 doses of 3 different batches of the tested vaccine at an interval of 21 d, or physiological saline solution (0.5 mL). Orbital blood was collected at an interval of 21 d post inoculation to detect related antibody titers or neutralizing antibody titers against poliovirus. Gross autopsy and histopathological examination revealed no abnormal toxicity or irritation in mice and cynomolgus monkeys. Sensitization assessment in guinea pigs indicated the lack of evident allergic symptoms in the high- and low-dose vaccine groups within 30 min after repeated stimulation. The DTacP-sIPV-Hib combination vaccine induced significant immune responses in mice, rats, and cynomolgus monkeys, with 100% seroconversion rates after 3 doses. The DTacP-sIPV-Hib combination vaccine is safe and immunogenic in animal models. Three doses of the vaccine elicited satisfactory antibody responses in mice, rats, and cynomolgus monkeys.

Gene Digital Circuits Based on CRISPR-Cas Systems and Anti-CRISPR Proteins.

Synthetic gene Boolean gates and digital circuits have a broad range of applications, from medical diagnostics to environmental care. The discovery of the CRISPR-Cas systems and their natural inhibitors-the anti-CRISPR proteins (Acrs)-provides a new tool to design and implement in vivo gene digital circuits. Here, we describe a protocol that follows the idea of the "Design-Build-Test-Learn" biological engineering cycle and makes use of dCas9/dCas12a together with their corresponding Acrs to establish small transcriptional networks, some of which behave like Boolean gates, in Saccharomyces cerevisiae. These results point out the properties of dCas9/dCas12a as transcription factors. In particular, to achieve maximal activation of gene expression, dSpCas9 needs to interact with an engineered scaffold RNA that collects multiple copies of the VP64 activation domain (AD). In contrast, dCas12a shall be fused, at the C terminus, with the strong VP64-p65-Rta (VPR) AD. Furthermore, the activity of both Cas proteins is not enhanced by increasing the amount of sgRNA/crRNA in the cell. This article also explains how to build Boolean gates based on the CRISPR-dCas-Acr interaction. The AcrIIA4 fused hormone-binding domain of the human estrogen receptor is the core of a NOT gate responsive to ß-estradiol, whereas AcrVAs synthesized by the inducible GAL1 promoter permits to mimic both YES and NOT gates with galactose as an input. In the latter circuits, AcrVA5, together with dLbCas12a, showed the best logic behavior.

Comparative multiomics analysis of cell physiological state after culture in a basket bioreactor.

Bioreactors are one of the most important, basic pieces of equipment in the biopharmaceutical industry. Understanding the effects of mechanical damage and other factors on the physiological state of cells during cell matrix culture is the basis for continuously achieving greater efficiency and higher product quality. In this study, Vero cells were used as a model and apoptosis, senescence, transcriptomics, proteomics, and metabolomics were carried out for analysis at the cellular and molecular levels. The results showed that compared with cells cultured in the simulated natural state, the cells cultured in the basket bioreactor displayed no obvious senescence. Additionally, the proportion of early apoptotic cells increased, but the proportions of damaged, late apoptotic and dead cells did not change significantly. The transcription levels of aminoacyl-tRNA synthetase and cyclin D1 and the expression levels of DNA replication licensing factor, methenyltetrahydrofolate cyclohydrolase, arachidonic acid and other metabolites of cells cultured in the basket bioreactor were significantly increased. These results suggest that DNA replication, protein translation and the metabolic activities in cells cultured in basket bioreactors are more active, which is more conducive to cell amplification and target product production. In this study, the growth and physiological state of cells in a basket bioreactor were characterized at the molecular level for the first time. Additionally, a tool to evaluate the physiological state of cells in a bioreactor was established, which can be used to guide the development and optimization of cell matrix culture conditions in industrial production and improve the production efficiency of the target products.

Defective ryanodine receptor N-terminus inter-subunit interaction is a common mechanism in neuromuscular and cardiac disorders.

The ryanodine receptor (RyR) is a homotetrameric channel mediating sarcoplasmic reticulum Ca2+ release required for skeletal and cardiac muscle contraction. Mutations in RyR1 and RyR2 lead to life-threatening malignant hyperthermia episodes and ventricular tachycardia, respectively. In this brief report, we use chemical cross-linking to demonstrate that pathogenic RyR1 R163C and RyR2 R169Q mutations reduce N-terminus domain (NTD) tetramerization. Introduction of positively-charged residues (Q168R, M399R) in the NTD-NTD inter-subunit interface normalizes RyR2-R169Q NTD tetramerization. These results indicate that perturbation of NTD-NTD inter-subunit interactions is an underlying molecular mechanism in both RyR1 and RyR2 pathophysiology. Importantly, our data provide proof of concept that stabilization of this critical RyR1/2 structure-function parameter offers clear therapeutic potential.

IACRA: Lifetime Optimization by Invulnerability-Aware Clustering Routing Algorithm Using Game-Theoretic Approach for Wsns.

Energy limitation is one of the intrinsic shortcomings of wireless sensor networks (WSNs), although it has been widely applied in disaster response, battlefield surveillance, wildfire monitoring, radioactivity detection, etc. Due to the large amount of energy consumed for data transmission, how to prolong the network lifespan by designing various hierarchical routing protocols has attracted more and more attention. As a result, numerous achievements have emerged successively. However, these presented mechanisms can rarely guarantee the satisfactory quality of service (QoS), while lowering the energy cost level of WSNs. Meanwhile, invulnerability is undoubtedly an excellent quantitative index to assess QoS. Therefore, it is critical to develop a practical routing method to optimize network lifetime by considering both invulnerability and energy efficiency. Game theory is suitable for such a critical problem as it can be used in node or at network level to encourage the decision-making capabilities of WSNs. In this paper, a novel invulnerability-aware clustering routing algorithm (IACRA) using game-theoretic method is proposed to solve the predicament. The core features of the addressed game-theory-based routing protocol include integral invulnerability awareness, optimal cluster head selection in hierarchical routing, distance-aware cluster head discovery, and cluster rotation update mechanism for lifetime optimization. Particularly, the integral network invulnerability based on weighted fusion is constructed for further defining the profit model by combining the invulnerability indicators used to evaluate the local and whole network. Meanwhile, the optimal probability function of every node elected as CH in per cluster is established through the game between invulnerability and node energy consumption. In addition, the cluster update mechanism base on cluster rotation is proposed to avoid the rapid death of nodes with large energy consumption for maximizing network lifetime. The experimental results indicated a significant improvement in energy balance as well as in invulnerability compared with the other three kinds of well-known clustering routing protocols including GEEC (Game-theory-based energy efficient clustering routing protocol), HGTD (Hybrid, game-theory-based distributed clustering protocol), and EEGC (Efficient energy-aware and game-theory-based clustering protocol). Concretely, at the 400 communication rounds, the invulnerability of IACRA was higher than that of GEEC, HGTD, and EEGC by 77.56%, 29.45% and 15.90%, respectively, and the average residual energy of IACRA was 8.61%, 18.35% and 6.36% larger than that of GEEC, HGTD, and EEGC, respectively. Based on these results, the proposed protocol can be utilized to increase the capability of WSNs against deterioration of QoS and energy constraints.

DHPA Protects SH-SY5Y Cells from Oxidative Stress-Induced Apoptosis via Mitochondria Apoptosis and the Keap1/Nrf2/HO-1 Signaling Pathway.

Oxidative stress in the brain is highly related to the pathogenesis of Alzheimer's disease (AD). It could be induced by the overproduction of reactive oxygen species (ROS), produced by the amyloid beta (Aß) peptide and excess copper (Cu) in senile plaques and cellular species, such as ascorbic acid (AA) and O2. In this study, the protective effect of 5-hydroxy-7-(4'-hydroxy-3'-methoxyphenyl)-1-phenyl-3-heptanone (DHPA) on Aß(1-42)/Cu2+/AA mixture-treated SH-SY5Y cells was investigated via in vitro and in silico studies. The results showed that DHPA could inhibit Aß/Cu2+/AA-induced SH-SY5Y apoptosis, OH· production, intracellular ROS accumulation, and malondialdehyde (MDA) production. Further research demonstrated that DHPA could decrease the ratio of Bax/Bcl-2 and repress the increase of mitochondrial membrane potential (MMP) of SH-SY5Y cells, to further suppress the activation of caspase-3, and inhibit cell apoptosis. Meanwhile, DHPA could inhibit the Aß/Cu2+/AA-induced phosphorylation of Erk1/2 and P38 in SH-SY5Y cells, and increase the expression of P-AKT. Furthermore, DHPA could bind to Keap1 to promote the separation of Nrf2 to Keap1 and activate the Keap1/Nrf2/HO-1 signaling pathway to increase the expression of heme oxygenase-1 (HO-1), quinone oxidoreductase-1 (NQO1), glutathione (GSH), and superoxide dismutase (SOD). Thus, our results demonstrated that DHPA could inhibit Aß/Cu2+/AA-induced SH-SY5Y apoptosis via scavenging OH·, inhibit mitochondria apoptosis, and activate the Keap1/Nrf2/HO-1 signaling pathway.

Alum/CpG Adjuvanted Inactivated COVID-19 Vaccine with Protective Efficacy against SARS-CoV-2 and Variants.

Since the beginning of the COVID-19 pandemic, numerous variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have emerged, including five variants of concern (VOC) strains listed by the WHO: Alpha, Beta, Gamma, Delta and Omicron. Extensive studies have shown that most of these VOC strains, especially the currently dominant variant Omicron, can escape the host immune response induced by existing COVID-19 vaccines to different extents, which poses considerable risk to the health of human beings around the world. In the present study, we developed a vaccine based on inactivated SARS-CoV-2 and an adjuvant consisting of aluminum hydroxide (alum) and CpG. The immunogenicity and safety of the vaccine were investigated in rats. The candidate vaccine elicited high titers of SARS-CoV-2-spike-specific IgG antibody and neutralizing antibody in immunized rats, which not only neutralize the original SARS-CoV-2, but also showed great cross-neutralization activity against the Beta, Delta and Omicron variants.

A Mutated Nme1Cas9 Is a Functional Alternative RNase to Both LwaCas13a and RfxCas13d in the Yeast S. cerevisiae.

CRISPR-Cas systems provide powerful biological tools for genetic manipulation and gene expression regulation. Class 2 systems, comprising type II, type V, and type VI, have the significant advantage to require a single effector Cas protein (Cas9, Cas12, and Cas13 respectively) to cleave nucleic acids upon binding the crRNA. Both Cas9 and Cas12 recognize DNA and induce a double-strand break in it. In contrast, Cas13 bind and cleave RNA exclusively. However, some Cas9 homologs have shown RNase activity as well. Here, we harnessed Nme1Cas9, LwaCas13a, and RfxCas13d to carry out gene downregulation in Saccharomyces cerevisiae by triggering mRNA degradation. To avoid potential DNA damage, we mutated Nme1Cas9 into d16ANme1Cas9 that lost the nuclease activity of the RuvC domain but retained the active HNH domain, able to act on the target DNA strand and, therefore, on the corresponding transcript. Our results showed that d16ANme1Cas9 is a functional RNase in vivo, although with moderate activity since it provoked a fluorescence reduction from 21% to 32%. Interestingly, d16ANme1Cas9 works in a PAM-independent way nor demands helper PAMmer molecules. LwaCas13a and RfxCas13d appeared substantially unfunctional in S. cerevisiae, though they were shown to perform well in mammalian cells. To the best of our knowledge, this is the first report about the working in vivo of a variant of Nme1Cas9 as an RNase and the issues connected with the usage of Cas13 proteins in S. cerevisiae.

In Vitro Preparation of Actively Maturing Bovine Articular Cartilage Explants for X-Ray Phase Contrast Imaging.

Understanding the mechanisms that underpin post-natal maturation of articular cartilage is of crucial importance for designing the next generation of tissue engineering strategies and potentially repairing diseased or damaged cartilage. In general, postnatal maturation of the articular cartilage, which is a wholesale change in collagen structure and function of the tissue to accommodate growth of the organism, occurs over a timescale ranging from months to years. Conversely dissolution of the structural organization of the cartilage that also occurs over long timescales is the hallmark of tissue degeneration. Our ability to study these biological processes in detail have been enhanced by the findings that growth factors can induce precocious in vitro maturation of immature articular cartilage. The developmental and disease related changes that occur in the joint involve bone and cartilage and an ability to co-image these tissues would significantly increase our understanding of their intertwined roles. The simultaneous visualization of soft tissue, cartilage and bone changes is nowadays a challenge to overcome for conventional preclinical imaging modalities used for the joint disease follow-up. Three-dimensional X-ray Phase-Contrast Imaging methods (PCI) have been under perpetual developments for 20 years due to high performance for imaging low density objects and their ability to provide additional information compared to conventional X-ray imaging. In this protocol we detail the procedure used in our experiments from biopsy of the cartilage, generation of in vitro matured cartilage to data analysis of image collected using X-ray phase contrast imaging.

Interaction of Bare dSpCas9, Scaffold gRNA, and Type II Anti-CRISPR Proteins Highly Favors the Control of Gene Expression in the Yeast S. cerevisiae.

Type II CRISPR-(d)SpCas9 and anti-CRISPR proteins (AcrIIs) show evidence of coevolution and competition for survival between bacteria and phages. In biotechnology, CRISPR-(d)SpCas9 is utilized for gene editing and transcriptional regulation. Moreover, its activity is controlled by AcrIIs. However, studies of dSpCas9/AcrII-based transcription regulation in Saccharomyces cerevisiae are rare. In this work, we used dSpCas9 as a template to engineer new transcription activators. We found that the most performant activation system requires the use of bare dSpCas9 in conjunction with scaffold gRNA (scRNA). This means that activation domains shall not be fused to dSpCas9 but rather interact with scRNA. We showed that a low amount of sgRNA is not a limiting factor in dSpCas9-driven transcription regulation. Moreover, a high quantity of sgRNA does not improve, generally, activation (and repression) efficiency. Importantly, we analyzed the performance of AcrIIA2, AcrIIA4, and AcrIIA5 in S. cerevisiae in depth. AcrIIA4 is the strongest of the three AcrIIs and also the only one able to induce high inhibition at low concentrations. However, the activation domains fused to dSpCas9 hindered interactions with the AcrIIs as well and limited their control of gene transcription regulation, confirming that bare dSpCas9 is the best solution for building synthetic genetic networks in yeast.

High performance near-infrared phototransistors via enhanced electron trapping effect.

A high performance near-infrared organic phototransistor is achieved via introducing a small molecule acceptor as an electron trapping site into the narrow-bandgap conjugated polymer films. With only 10% (wt) addition of the acceptor molecule, the photoresponse to light of 850 nm has been significantly improved with a best photoresponsivity up to 2000 A W-1, high detectivity of 1016 Jones and fairly good photosensitivity in the order of 106.

Supramolecular Drug Delivery System from Macrocycle-Based Self-Assembled Amphiphiles for Effective Tumor Therapy.

Intelligent drug delivery systems (DDSs) that can improve therapeutic outcomes of antitumor agents and decrease their side effects are urgently needed to satisfy special requirements of treatment of malignant tumors in clinics. Here, the fabrication of supramolecular self-assembled amphiphiles based on the host-guest recognition between a cationic water-soluble pillar[6]arene (WP6A) host and a sodium decanesulfonate guest (G) is reported. The chemotherapeutic agent doxorubicin hydrochloride (DOX) can be encapsulated into the formed vesicle (G/WP6A) to construct supramolecular DDS (DOX@G/WP6A). WP6A affords strong affinities to G to avoid undesirable off-target leakage during delivery. Nanoscaled DOX@G/WP6A is capable of preferentially accumulating in tumor tissue via enhanced permeability and retention (EPR) effect. After internalization by tumor cells, the abundant adenosine triphosphate (ATP) binds competitively with WP6A to trigger the disintegration of self-assembled vesicles with the ensuing release of DOX. In vitro and in vivo research confirmed that DOX@G/WP6A is not only able to promote antitumor efficacy but also reduce DOX-related systemic toxicity. The above favorable findings are ascribed to the formation of ternary self-assembly, which profits from the combination of the factors of the EPR effect and the ATP-triggered release.

A Dirac nodal surface semi-metallic carbon-based structure as a universal anode material for metal-ion batteries with high performance.

The rapid development of electronic devices requires high power storage batteries. However, reported 3D carbon-based materials are semiconductors or metals and are used in Li- or Na-ion batteries with low capacities. Thus, it is of interest to discover whether there is a universal semi-metallic material for use in high performance Li-, Na-, and K-ion batteries. Inspired by the recent synthesis of 3D carbon-based materials, in the research reported here, a 3D regular porous structure (bct-C56) is designed using graphene sheets. The porous carbon-based material has mechanical, dynamic, thermal, and mechanical stabilities. Interestingly, bct-C56 exhibits semi-metallic features with two Dirac nodal surfaces with mirror symmetry, as well as high Fermi velocities, indicating high electron-transport abilities. More excitingly, its theoretical capacities are 743.8, 478.2, and 425.0 mA h g-1, with diffusion barriers of 0.05-0.12, 0.07-0.12, and 0.03-0.05 eV, average OCVs of 0.31, 0.45, and 0.59 V, and volume expansion levels of 1.2%, 0.02%, and 3.1%, in Li-, Na-, and K-ion batteries, respectively. All these excellent characteristics suggest that semi-metallic bct-C56 is a universal anode material for use in metal-ion batteries with a fast charge-discharge rate. In this research, not only was a new material with a Dirac nodal surface feature designed, but it also offers an approach for the creation of high performance and universal metal-ion battery anodes with 3D porous carbon materials.

Role of hydrogen peroxide in intra-operative wound preparation based on an in vitro fibrin clot degradation model.

Three per cent hydrogen peroxide (H2O2) is widely used to irrigate acute and chronic wounds in the surgical setting and clinical experience tells us that it is more effective at removing dried-on blood than normal saline alone. We hypothesise that this is due to the effect of H2O2 on fibrin clot architecture via fibrinolysis. We investigate the mechanisms and discuss the clinical implications using an in vitro model. Coagulation assays with normal saline (NaCl), 1% and 3% concentrations of H2O2 were performed to determine the effect on fibrin clot formation. These effects were confirmed by spectrophotometry. The effects of 1%, 3% and 10% H2O2 on the macroscopic and microscopic features of fibrin clots were assessed at set time intervals and compared to a NaCl control. Quantitative analysis of fibrin networks was undertaken to determine the fibre length, diameter, branch point density and pore size. Fibrin clots immersed in 1%, 3% and 10% H2O2 demonstrated volume losses of 0.09-0.25mm3/min, whereas those immersed in the normal saline gained in volume by 0.02±0.13 mm3/min. Quantitative analysis showed that H2O2 affects the structure of the fibrin clot in a concentration-dependent manner, with the increase in fibre length, diameter and consequently pore sizes. Our results support our hypothesis that the efficacy of H2O2 in cleaning blood from wounds is enhanced by its effects on fibrin clot architecture in a concentration- and time-dependent manner. The observed changes in fibre size and branch point density suggest that H2O2 is acting on the quaternary structure of the fibrin clot, most likely via its effect on cross-linking of the fibrin monomers and may therefore be of benefit for the removal of other fibrin-dependent structures such as wound slough.