Dose-Dependent Production of Anti-PEG IgM after Intramuscular PEGylated-Hydrogenated Soy Phosphatidylcholine Liposomes, but Not Lipid Nanoparticle Formulations of DNA, Correlates with the Plasma Clearance of PEGylated Liposomal Doxorubicin in Rats.

PEGylated lipid nanoparticle-based Covid-19 vaccines, including Pfizer's BNT162b2 and Moderna's mRNA-1273, have been shown to stimulate variable anti-PEG antibody production in humans. Anti-PEG antibodies have the potential to accelerate the plasma clearance of PEGylated therapeutics, such as PEGylated liposomes and proteins, and compromise their therapeutic efficacy. However, it is not yet clear whether antibody titers produced by PEGylated Covid-19 vaccines significantly affect the clearance of PEGylated therapeutics. This study examined how anti-PEG IgM levels affect the pharmacokinetics of PEGylated liposomal doxorubicin (PLD) and compared the immunogenicity of a lipid nanoparticle formulation of linear DNA (DNA-LNP) to standard PEG-HSPC liposomes. DNA-LNP was prepared using the same composition and approach as Pfizer's BNT162b2, except linear double-stranded DNA was used as the genetic material. PEGylated HSPC-based liposomes were formulated using the lipid rehydration and extrusion method. Nanoparticles were dosed IM to rats at 0.005-0.5 mg lipid/kg body weight 1 week before evaluating the plasma pharmacokinetics of clinically relevant doses of PLD (1 mg/kg, IV) or PEGylated interferon α2a (Pegasys, 5 µg/kg, SC). Plasma PEG IgM was compared between pre- and 1-week post-dose blood samples. The results showed that anti-PEG IgM production increased with increasing PEG-HSPC liposome dose and that IgM significantly correlated with the plasma half-life, clearance, volume of distribution, and area under the curve of a subsequent dose of PLD. The plasma exposure of Pegasys was also significantly reduced after initial delivery of 0.005 mg/ml PEG-HSPC liposome. However, a single 0.05 mg/kg IM dose of DNA-LNP did not significantly elevate PEG IgM and did not alter the IV pharmacokinetics of PLD. These data showed that PEGylated Covid-19 vaccines are less immunogenic compared to standard PEGylated HSPC liposomes and that there is an antibody threshold for accelerating the clearance of PEGylated therapeutics.

Potentiating the Cross-Reactive IFN-γ T Cell and Polyfunctional T Cell Responses by Heterologous GX-19N DNA Booster in Mice Primed with Either a COVID-19 mRNA Vaccine or Inactivated Vaccine.

Waning vaccine-induced immunity, coupled with the emergence of SARS-CoV-2 variants, has inspired the widespread implementation of COVID-19 booster vaccinations. Here, we evaluated the potential of the GX-19N DNA vaccine as a heterologous booster to enhance the protective immune response to SARS-CoV-2 in mice primed with either an inactivated virus particle (VP) or an mRNA vaccine. We found that in the VP-primed condition, GX-19N enhanced the response of both vaccine-specific antibodies and cross-reactive T Cells to the SARS-CoV-2 variant of concern (VOC), compared to the homologous VP vaccine prime-boost. Under the mRNA-primed condition, GX-19N induced higher vaccine-induced T Cell responses but lower antibody responses than the homologous mRNA vaccine prime-boost. Furthermore, the heterologous GX-19N boost induced higher S-specific polyfunctional CD4+ and CD8+ T cell responses than the homologous VP or mRNA prime-boost vaccinations. Our results provide new insights into booster vaccination strategies for the management of novel COVID-19 variants.

NETosis in Parasitic Infections: A Puzzle That Remains Unsolved.

Neutrophils are the key players in the innate immune system, being weaponized with numerous strategies to eliminate pathogens. The production of extracellular traps is one of the effector mechanisms operated by neutrophils in a process called NETosis. Neutrophil extracellular traps (NETs) are complex webs of extracellular DNA studded with histones and cytoplasmic granular proteins. Since their first description in 2004, NETs have been widely investigated in different infectious processes. Bacteria, viruses, and fungi have been shown to induce the generation of NETs. Knowledge is only beginning to emerge about the participation of DNA webs in the host's battle against parasitic infections. Referring to helminthic infections, we ought to look beyond the scope of confining the roles of NETs solely to parasitic ensnarement or immobilization. Hence, this review provides detailed insights into the less-explored activities of NETs against invading helminths. In addition, most of the studies that have addressed the implications of NETs in protozoan infections have chiefly focused on their protective side, either through trapping or killing. Challenging this belief, we propose several limitations regarding protozoan-NETs interaction. One of many is the duality in the functional responses of NETs, in which both the positive and pathological aspects seem to be closely intertwined.

Quantifying Myeloperoxidase-DNA and Neutrophil Elastase-DNA Complexes from Neutrophil Extracellular Traps by Using a Modified Sandwich ELISA.

Certain stimuli, such as microorganisms, cause neutrophils to release neutrophil extracellular traps (NETs), which are basically web-like structures composed of DNA with granule proteins, such as myeloperoxidase (MPO) and neutrophil elastase (NE), and cytoplasmic and cytoskeletal proteins. Although interest in NETs has increased recently, no sensitive, reliable assay method is available for measuring NETs in clinical settings. This article describes a modified sandwich enzyme-linked immunosorbent assay to quantitatively measure two components of circulating NETs, MPO-DNA and NE-DNA complexes, which are specific components of NETs and are released into the extracellular space as breakdown products of NETs. The assay uses specific monoclonal antibodies for MPO or NE as the capture antibodies and a DNA-specific detection antibody. MPO or NE binds to one site of the capture antibody during the initial incubation of samples containing MPO-DNA or NE-DNA complexes. This assay shows good linearity and high inter-assay and intra-assay precision. We used it in 16 patients with COVID-19 with accompanying acute respiratory distress syndrome and found that the plasma concentrations of MPO-DNA and NE-DNA were significantly higher than in the plasma obtained from healthy controls. This detection assay is a reliable, highly sensitive, and useful method for investigating the characteristics of NETs in human plasma and culture supernatants.

Cytoplasmic DNAs: Sources, sensing, and roles in the development of lung inflammatory diseases and cancer.

Cytoplasmic DNA is emerging as a pivotal contributor to the pathogenesis of inflammatory diseases and cancer, such as COVID-19 and lung carcinoma. However, the complexity of various cytoplasmic DNA-related pathways and their crosstalk remains challenging to distinguish their specific roles in many distinct inflammatory diseases, especially for the underlying mechanisms. Here, we reviewed the latest findings on cytoplasmic DNA and its signaling pathways in inflammatory lung conditions and lung cancer progression. We found that sustained activation of cytoplasmic DNA sensing pathways contributes to the development of common lung diseases, which may result from external factors or mutations of key genes in the organism. We further discussed the interplays between cytoplasmic DNA and anti-inflammatory or anti-tumor effects for potential immunotherapy. In sum, this review aids in understanding the roles of cytoplasmic DNAs and exploring more therapeutic strategies.

Accelerating Cell-Free Biosensors by Entropy-Driven Assembly of Transcription Templates.

Due to its high efficiency and selectivity, cell-free biosynthesis has found broad utility in the fields of bioproduction, environment monitoring, and disease diagnostics. However, the practical application is limited by its low productivity. Here, we introduce the entropy-driven assembly of transcription templates as dynamic amplifying modules to accelerate the cell-free transcription process. The catalytic DNA circuit with high sensitivity and enzyme-free format contributes to the production of large amounts of transcription templates, drastically accelerating the as-designed cell-free transcription system without interference from multiple enzymes. The proposed approach was successfully applied to the ultrasensitive detection of SARS-CoV-2, improving the sensitivity by 3 orders of magnitude. Thanks to the high programmability and diverse light-up RNA pairs, the method can be adapted to multiplexing detection, successfully demonstrated by the analysis of two different sites of the SARS-CoV-2 gene in parallel. Further, the flexibility of the entropy-driven circuit enables a dynamic responding range by tuning the circuit layers, which is beneficial for responding to targets with different concentration ranges. The strategy was also applied to the analysis of clinical samples, providing an alternative for sensitively detecting the current SARS-CoV-2 RNA that quickly mutates.

Immune response enhancement with GLS-5310 DNA primary vaccine against SARS-CoV-2 followed by administration of an mRNA vaccine heterologous boost.

Heterologous boost regimens are being increasingly considered against SARS-CoV-2. We report results for the 32 of 45 participants in the Phase 1 CoV2-001 clinical trial (Kim et al., Int J Iinfect Dis 2023, 128:112-120) who elected to receive an EUA-approved SARS-CoV-2 mRNA vaccine 6 to 8 months following a two-dose primary vaccination with the GLS-5310 bi-cistronic DNA vaccine given intradermally and followed by application of suction using the GeneDerm device. Receipt of EUA-approved mRNA vaccines after GLS-5310 vaccination was well-tolerated, with no reported adverse events. Immune responses were enhanced such that binding antibody titers, neutralizing antibody titers, and T-cell responses increased 1,187-fold, 110-fold, and 2.9-fold, respectively. This paper is the first description of the immune responses following heterologous vaccination with a DNA primary series and mRNA boost.

Multiplexable and Biocomputational Virus Detection by CRISPR-Cas9-Mediated Strand Displacement.

Recurrent disease outbreaks caused by different viruses, including the novel respiratory virus SARS-CoV-2, are challenging our society at a global scale; so versatile virus detection methods would enable a calculated and faster response. Here, we present a novel nucleic acid detection strategy based on CRISPR-Cas9, whose mode of action relies on strand displacement rather than on collateral catalysis, using the Streptococcus pyogenes Cas9 nuclease. Given a preamplification process, a suitable molecular beacon interacts with the ternary CRISPR complex upon targeting to produce a fluorescent signal. We show that SARS-CoV-2 DNA amplicons generated from patient samples can be detected with CRISPR-Cas9. We also show that CRISPR-Cas9 allows the simultaneous detection of different DNA amplicons with the same nuclease, either to detect different SARS-CoV-2 regions or different respiratory viruses. Furthermore, we demonstrate that engineered DNA logic circuits can process different SARS-CoV-2 signals detected by the CRISPR complexes. Collectively, this CRISPR-Cas9 R-loop usage for the molecular beacon opening (COLUMBO) platform allows a multiplexed detection in a single tube, complements the existing CRISPR-based methods, and displays diagnostic and biocomputing potential.

Metagenomics of gut microbiome for migratory seagulls in Kunming city revealed the potential public risk to human health.

BACKGROUND: Seagull as a migratory wild bird has become most popular species in southwest China since 1980s. Previously, we analyzed the gut microbiota and intestinal pathogenic bacteria configuration for this species by using 16S rRNA sequencing and culture methods. To continue in-depth research on the gut microbiome of migratory seagulls, the metagenomics, DNA virome and RNA virome were both investigated for their gut microbial communities of abundance and diversity in this study. RESULTS: The metagenomics results showed 99.72% of total species was bacteria, followed by viruses, fungi, archaea and eukaryota. In particular, Shigella sonnei, Escherichia albertii, Klebsiella pneumonia, Salmonella enterica and Shigella flexneri were the top distributed taxa at species level. PCoA, NMDS, and statistics indicated some drug resistant genes, such as adeL, evgS, tetA, PmrF, and evgA accumulated as time went by from November to January of the next year, and most of these genes were antibiotic efflux. DNA virome composition demonstrated that Caudovirales was the most abundance virus, followed by Cirlivirales, Geplafuvirales, Petitvirales and Piccovirales. Most of these phages corresponded to Enterobacteriaceae and Campylobacteriaceae bacterial hosts respectively. Caliciviridae, Coronaviridae and Picornaviridae were the top distributed RNA virome at family level of this migratory animal. Phylogenetic analysis indicated the sequences of contigs of Gammacoronavirus and Deltacoronavirus had highly similarity with some coronavirus references. CONCLUSIONS: In general, the characteristics of gut microbiome of migratory seagulls were closely related to human activities, and multiomics still revealed the potential public risk to human health.

A dual-labeled fluorescence quenching lateral flow assay based on one-pot enzyme-free isothermal cascade amplification for the rapid and sensitive detection of pathogens.

Rapid detection of nucleic acids is integral for clinical diagnostics, especially if a major public-health emergency occurs. However, such detection cannot be carried out efficiently in remote areas limited by medical resources. Herein, a dual-labeled fluorescence resonance energy transfer (FRET) lateral flow assay (LFA) based on one-pot enzyme-free cascade amplification was developed for rapid, convenient, and sensitive detection of open reading frame (ORF)1ab of severe acute respiratory syndrome-coronavirus-2. The catalyzed hairpin assembly (CHA) reaction of two well-designed hairpin probes was initiated by a target sequence and generated a hybridization chain reaction (HCR) initiator. Then, HCR probes modified with biotin were initiated to produce long DNA nanowires. After two-level amplification, the cascade-amplified product was detected by dual-labeled lateral flow strips. Gold nanoparticles (AuNPs)-streptavidin combined with the product and then ran along a nitrocellulose membrane under the action of capillary force. After binding with fluorescent microsphere-labeled-specific probes on the T line, a positive signal (red color) could be observed. Meanwhile, AuNPs could quench the fluorescence of the T line, and an inverse relationship between fluorescence intensity and the concentration of the CHA-HCR-amplified product was formed. The proposed strategy achieved a satisfactory limit of detection of 2.46 pM for colorimetric detection and 174 fM for fluorescent detection, respectively. Benefitting from the features of being one-pot, enzyme-free, low background, high sensitivity, and selectivity, this strategy shows great potential in bioanalysis and clinical diagnostics upon further development.

The CRISPR/Cas System: A Customizable Toolbox for Molecular Detection.

Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated proteins (Cas) are promising molecular diagnostic tools for rapidly and precisely elucidating the structure and function of genomes due to their high specificity, programmability, and multi-system compatibility in nucleic acid recognition. Multiple parameters limit the ability of a CRISPR/Cas system to detect DNA or RNA. Consequently, it must be used in conjunction with other nucleic acid amplification techniques or signal detection techniques, and the reaction components and reaction conditions should be modified and optimized to maximize the detection performance of the CRISPR/Cas system against various targets. As the field continues to develop, CRISPR/Cas systems have the potential to become an ultra-sensitive, convenient, and accurate biosensing platform for the detection of specific target sequences. The design of a molecular detection platform employing the CRISPR/Cas system is asserted on three primary strategies: (1) Performance optimization of the CRISPR/Cas system; (2) enhancement of the detection signal and its interpretation; and (3) compatibility with multiple reaction systems. This article focuses on the molecular characteristics and application value of the CRISPR/Cas system and reviews recent research progress and development direction from the perspectives of principle, performance, and method development challenges to provide a theoretical foundation for the development and application of the CRISPR/CAS system in molecular detection technology.

Accurate quantification of DNA using on-site PCR (osPCR) by characterizing DNA amplification at single-molecule resolution.

Despite the need in various applications, accurate quantification of nucleic acids still remains a challenge. The widely-used qPCR has reduced accuracy at ultralow template concentration and is susceptible to nonspecific amplifications. The more recently developed dPCR is costly and cannot handle high-concentration samples. We combine the strengths of qPCR and dPCR by performing PCR in silicon-based microfluidic chips and demonstrate high quantification accuracy in a large concentration range. Importantly, at low template concentration, we observe on-site PCR (osPCR), where only certain sites of the channel show amplification. The sites have almost identical ct values, showing osPCR is a quasi-single molecule phenomenon. Using osPCR, we can measure both the ct values and the absolute concentration of templates in the same reaction. Additionally, osPCR enables identification of each template molecule, allowing removal of nonspecific amplification during quantification and greatly improving quantification accuracy. We develop sectioning algorithm that improves the signal amplitude and demonstrate improved detection of COVID in patient samples.

Monkeypox viral nucleic acids detected using both DNA and RNA extraction workflows.

Molecular methods have been used to detect human pathogens in wastewater with sampling typically performed at wastewater treatment plants (WWTP) and upstream locations within the sewer system. A wastewater-based surveillance (WBS) program was established at the University of Miami (UM) in 2020, which included measurements of SARS-CoV-2 levels in wastewater from its hospital and within the regional WWTP. In addition to the development of a SARS-CoV-2 quantitative PCR (qPCR) assay, qPCR assays to detect other human pathogens of interest were also developed at UM. Here we report on the use of a modified set of reagents published by the CDC to detect nucleic acids of Monkeypox virus (MPXV) which emerged during May of 2022 to become a concern worldwide. Samples collected from the University hospital and from the regional WWTP were processed through DNA and RNA workflows and analyzed by qPCR to detect a segment of the MPXV CrmB gene. Results show positive detections of MPXV nucleic acids in the hospital and wastewater treatment plant wastewater which coincided with clinical cases in the community and mirrored the overall trend of nationwide MPXV cases reported to the CDC. We recommend the expansion of current WBS programs' methods to detect a broader range of pathogens of concern in wastewater and present evidence that viral RNA in human cells infected by a DNA virus can be detected in wastewater.

Clinical characteristics of the host DNA-removed metagenomic next-generation sequencing technology for detecting SARS-CoV-2, revealing host local immune signaling and assisting genomic epidemiology.

Background: Metagenomic next-generation sequencing (mNGS) technology has been central in detecting infectious diseases and helping to simultaneously reveal the complex interplay between invaders and their hosts immune response characteristics. However, it needs to be rigorously assessed for clinical utility. The present study is the first to evaluate the clinical characteristics of the host DNA-removed mNGS technology for detecting SARS-CoV-2, revealing host local immune signaling and assisting genomic epidemiology. Methods: 46 swab specimens collected from COVID-19 patients were assayed by two approved commercial RT-qPCR kits and mNGS. The evolutionary tree of SARS-CoV-2 was plotted using FigTree directly from one sample. The workflow of removing the host and retaining the host was compared to investigate the influence of host DNA removal on the performances of mNGS. Functional enrichment analysis of DEGs and xCell score were used to explore the characteristics of host local immune signaling. Results: The detection rate of mNGS achieved 92.9% (26/28) for 28 samples with a Ct value ≤ 35 and 81.1% (30/37) for all 46 samples. The genome coverage of SARS-CoV-2 could reach up to 98.9% when the Ct value is about 20 in swab samples. Removing the host could enhance the sensitivity of mNGS for detecting SARS-CoV-2 from the swab sample but does not affect the species abundance of microbes RNA. Improving the sequencing depth did not show a positive effect on improving the detection sensitivity of SARS-CoV-2. Cell type enrichment scores found multiple immune cell types were differentially expressed between patients with high and low viral load. Conclusions: The host DNA-removed mNGS has great potential utility and superior performance on comprehensive identification of SARS-CoV-2 and rapid traceability, revealing the microbiome's transcriptional profiles and host immune responses.

A polyethylene glycol enhanced ligation-triggered self-priming isothermal amplification for the detection of SARS-CoV-2 D614G mutation.

We presented a polyethylene glycol (PEG) enhanced ligation-triggered self-priming isothermal amplification (PEG-LSPA) for the detection D614G mutation in S-glycoprotein of SARS-CoV-2. PEG was employed to improve the ligation efficiency of this assay by constructing a molecular crowding environment. Two hairpin probes (H1 and H2) were designed to contain 18 nt and 20 nt target binding site at their 3' end and 5' end, respectively. In presence of target sequence, it complemented with H1 and H2 to trigger ligation by ligase under molecular crowding condition to form ligated H1-H2 duplex. Then 3' terminus of the H2 would be extended by DNA polymerase under isothermal conditions to form a longer extended hairpin (EHP1). 5' terminus of EHP1 with phosphorothioate (PS) modification could form hairpin structure due to the lower Tm value. The resulting 3' end overhang would also fold back as a new primer to initiate the next round of polymerization, resulting in the formation of a longer extended hairpin (EHP2) containing two target sequence domains. In the circle of LSPA, long extended hairpin (EHPx) containing numerous target sequence domains was produced. The resulting DNA products can be monitored in real-time fluorescence signaling. Our proposed assay owns an excellent linear range from 10 fM to 10 nM with a detection limit down to 4 fM. Thus, this work provides a potential isothermal amplification method for monitoring mutations in SARS-CoV-2 variants.

Unwinding mechanism of SARS-CoV helicase (nsp13) in the presence of Ca2+, elucidated by biochemical and single-molecular studies.

The recent outbreak of COVID-19 has created a serious health crisis with fatFal infectious viral diseases, such as Severe Acute Respiratory Syndrome (SARS). The nsp13, a helicase of coronaviruses is an essential element for viral replication that unwinds secondary structures of DNA and RNA, and is thus considered a major therapeutic target for treatment. The replication of coronaviruses and other retroviruses occurs in the cytoplasm of infected cells, in association with viral replication organelles, called virus-induced cytosolic double-membrane vesicles (DMVs). In addition, an increase in cytosolic Ca2+ concentration accelerates viral replication. However, the molecular mechanism of nsp13 in the presence of Ca2+ is not well understood. In this study, we applied biochemical methods and single-molecule techniques to demonstrate how nsp13 achieves its unwinding activity while performing ATP hydrolysis in the presence of Ca2+. Our study found that nsp13 could efficiently unwind double stranded (ds) DNA under physiological concentration of Ca2+ of cytosolic DMVs. These findings provide new insights into the properties of nsp13 in the range of calcium in cytosolic DMVs.

Recent Progress in Development and Application of DNA, Protein, Peptide, Glycan, Antibody, and Aptamer Microarrays.

Microarrays are one of the trailblazing technologies of the last two decades and have displayed their importance in all the associated fields of biology. They are widely explored to screen, identify, and gain insights on the characteristics traits of biomolecules (individually or in complex solutions). A wide variety of biomolecule-based microarrays (DNA microarrays, protein microarrays, glycan microarrays, antibody microarrays, peptide microarrays, and aptamer microarrays) are either commercially available or fabricated in-house by researchers to explore diverse substrates, surface coating, immobilization techniques, and detection strategies. The aim of this review is to explore the development of biomolecule-based microarray applications since 2018 onwards. Here, we have covered a different array of printing strategies, substrate surface modification, biomolecule immobilization strategies, detection techniques, and biomolecule-based microarray applications. The period of 2018-2022 focused on using biomolecule-based microarrays for the identification of biomarkers, detection of viruses, differentiation of multiple pathogens, etc. A few potential future applications of microarrays could be for personalized medicine, vaccine candidate screening, toxin screening, pathogen identification, and posttranslational modifications.

Specific Detection of Proteins by a Nanobody-Functionalized Nanopore Sensor.

Nanopores are label-free single-molecule analytical tools that show great potential for stochastic sensing of proteins. Here, we described a ClyA nanopore functionalized with different nanobodies through a 5-6 nm DNA linker at its periphery. Ty1, 2Rs15d, 2Rb17c, and nb22 nanobodies were employed to specifically recognize the large protein SARS-CoV-2 Spike, a medium-sized HER2 receptor, and the small protein murine urokinase-type plasminogen activator (muPA), respectively. The pores modified with Ty1, 2Rs15d, and 2Rb17c were capable of stochastic sensing of Spike protein and HER2 receptor, respectively, following a model where unbound nanobodies, facilitated by a DNA linker, move inside the nanopore and provoke reversible blockade events, whereas engagement with the large- and medium-sized proteins outside of the pore leads to a reduced dynamic movement of the nanobodies and an increased current through the open pore. Exploiting the multivalent interaction between trimeric Spike protein and multimerized Ty1 nanobodies enabled the detection of picomolar concentrations of Spike protein. In comparison, detection of the smaller muPA proteins follows a different model where muPA, complexing with the nb22, moves into the pore, generating larger blockage signals. Importantly, the components in blood did not affect the sensing performance of the nanobody-functionalized nanopore, which endows the pore with great potential for clinical detection of protein biomarkers.

Viral Nucleases.

Nucleases are ubiquitous hydrolytic enzymes that cleave phosphodiester bond of DNA (DNases), RNA (RNases), or protein-RNA/DNA (phosphodiesterases), within the strand (endonucleases) or from the end (exonucleases) [...].

Classification of nucleic acid amplification on ISFET arrays using spectrogram-based neural networks.

The COVID-19 pandemic has highlighted a significant research gap in the field of molecular diagnostics. This has brought forth the need for AI-based edge solutions that can provide quick diagnostic results whilst maintaining data privacy, security and high standards of sensitivity and specificity. This paper presents a novel proof-of-concept method to detect nucleic acid amplification using ISFET sensors and deep learning. This enables the detection of DNA and RNA on a low-cost and portable lab-on-chip platform for identifying infectious diseases and cancer biomarkers. We show that by using spectrograms to transform the signal to the time-frequency domain, image processing techniques can be applied to achieve the reliable classification of the detected chemical signals. Transformation to spectrograms is beneficial as it makes the data compatible with 2D convolutional neural networks and helps gain significant performance improvement over neural networks trained on the time domain data. The trained network achieves an accuracy of 84% with a size of 30kB making it suitable for deployment on edge devices. This facilitates a new wave of intelligent lab-on-chip platforms that combine microfluidics, CMOS-based chemical sensing arrays and AI-based edge solutions for more intelligent and rapid molecular diagnostics.