Tuning Hydrophobicity of Paper Substrates for Effective Colorimetric detection of Glucose and Nucleic acids.

This study investigated the colorimetric response of standard glucose, serum glucose, and nucleic acid assays on various paper surfaces with different wettability, including hydrophilic, hydrophobic, and nearly superhydrophobic surfaces. Water contact angles (WCA) formed by water droplets on each surface were measured using ImageJ software. The hydrophilic surface showed no contact angle, while the hydrophobic and nearly superhydrophobic surfaces exhibited contact angles of 115.667° and 133.933°, respectively. The colorimetric sensitivity of the standard glucose assay was analyzed on these surfaces, revealing enhanced sensitivity on the nearly superhydrophobic surface due to the high molecular crowding effect owing to its non-wetting behavior and eventually confined reaction product at the sample loading zone. The hydrophobic nature of the surface restricts the spreading and diffusion of the reaction product, leading to a controlled and localized concentration of the assay product leading to moderate colorimetric intensity. On the other hand, the hydrophilic surface showed the least enhancement in colorimetric sensitivity; this is attributed to the high wettability of the hydrophilic surface causing the reaction product to spread extensively, resulting in a larger area of dispersion and consequently a lower colorimetric intensity. The measured limit of detection (LOD) for nucleic acid on nearly superhydrophobic surfaces was found to be 16.15 ng/µL, which was almost four-fold lower than on hydrophilic surfaces (60.08 ng/µL). Additionally, the LODs of standard glucose and clinical serum samples were two-fold lower on nearly superhydrophobic surfaces compared to hydrophilic surfaces. Our findings clearly highlight the promising potential of utilizing superhydrophobic surfaces to significantly enhance colorimetric sensitivity in paper-based diagnostic applications. This innovative approach holds promise for advancing point-of-care diagnostics and improving disease detection in resource-limited settings.

Aptamers: Potential Diagnostic and Therapeutic Agents for Blood Diseases.

Aptamers are RNA/DNA oligonucleotide molecules that specifically bind to a targeted complementary molecule. As potential recognition elements with promising diagnostic and therapeutic applications, aptamers, such as monoclonal antibodies, could provide many treatment and diagnostic options for blood diseases. Aptamers present several superior features over antibodies, including a simple in vitro selection and production, ease of modification and conjugation, high stability, and low immunogenicity. Emerging as promising alternatives to antibodies, aptamers could overcome the present limitations of monoclonal antibody therapy to provide novel diagnostic, therapeutic, and preventive treatments for blood diseases. Researchers in several biomedical areas, such as biomarker detection, diagnosis, imaging, and targeted therapy, have widely investigated aptamers, and several aptamers have been developed over the past two decades. One of these is the pegaptanib sodium injection, an aptamer-based therapeutic that functions as an anti-angiogenic medicine, and it is the first aptamer approved by the U.S. Food and Drug Administration (FDA) for therapeutic use. Several other aptamers are now in clinical trials. In this review, we highlight the current state of aptamers in the clinical trial program and introduce some promising aptamers currently in pre-clinical development for blood diseases.

Simple and rapid peptide nanoprobe biosensor for the detection of Legionellaceae.

This study demonstrates the development of a sensitive, specific, and quantitative peptide-based nanoprobe prototype assay for the detection of Legionellaceae in a simple way and in a short time. In this work, proteases present in the culture supernatants of Legionella spp. were used as a biomarker. Fluorogenic peptide substrates, specific to Legionella strains culture supernatant proteases, were identified. Peptidases produced a significant increase in the fluorescence intensity following the cleavage of the dipeptide fluorogenic substrates. The specific substrates were identified and coupled with carboxyl-terminated nano-magnetic particles (NMPs). On the other hand, the C-terminal was conjugated with the cysteine residue to covalently integrate with a gold sensing platform via the Au-S linkage. Four different sensors were fabricated from the four specific substrates, which were treated with the protesase of six different species of Legionella. In the presence of specific protease, the peptide sequence is digested and the magnetic nanobeads moved out of the gold surface, resulting in the apparence of gold color. One of the nanoprobes sensitivity detects as low as 60 CFU mL-1 of Legionella anisa, Legionella micdadei, and Fluoribacter dumoffii. The cross-reactivity of the sensors was tested using other closely associated bacterial species and no significant cross-reactivity of the sensors was found. It is envisaged that this assay could be useful for screening purposes or might be supportive for the fast and easy detection of Legionella protease activity for water monitoring purposes.

Determination of minimal sequence for zearalenone aptamer by computational docking and application on an indirect competitive electrochemical aptasensor.

Aptamers are short single-stranded oligonucleotides (either DNA or RNA) that can fold into well-defined three-dimensional (3D) spatial structures which enable them to capture their specific target by complementary shape interactions. Aptamers are selected from large random libraries through the SELEX process and only a small fraction of the sequence is involved in direct docking with the target. In this paper, we describe the possible truncation variants of zearalenone (ZEA) aptamer which might be an effective binding region for the target. The originally selected zearalenone (ZEA) aptamer was 80-mer in length and shown to bind the target with a high affinity (Kd = 41 ± 5 nM). Herein, computational docking simulation was performed with 15 truncated variants to determine the predicted binding energy and responsible binding site of the aptamer-analyte complex. The results revealed that 5 truncated variants had binding energy lower than - 7.0 kcal/mol. Circular dichroism analysis was performed on the shortlisted aptamer and the conformational change of aptamers was observed with the presence of an analyte. Aptamer Z3IN (29-mer) was chosen as the most enhanced affinity for its target with a dissociation constant of 11.77 ± 1.44 nM. The aptamer was further applied in the electrochemical aptasensor of ZEA based on an indirect competitive format. The results demonstrated that the truncated aptamer leads to an enhancement of the sensitivity of the biosensor.

Peptide substrate screening for the diagnosis of SARS-CoV-2 using fluorescence resonance energy transfer (FRET) assay.

The novel corona (SARS-CoV-2) virus causes a global pandemic, which motivates researchers to develop reliable and effective methods for screening and detection of SARS-CoV-2. Though there are several methods available for the diagnosis of SARS-CoV-2 such as RT-PCR and ELSIA, nevertheless, these methods are time-consuming and may not apply at the point of care. In this study, we have developed a specific, sensitive, quantitative and fast detection method for SARS-CoV-2 by fluorescence resonance energy transfer (FRET) assay. The total extracellular protease proteolytic activity from the virus has been used as the biomarker. The specific peptide sequences from the library of 115 dipeptides were identified via changes in the fluorescence signal. The fluorogenic dipeptide substrates have the fluorophore and a quencher at the N- and the C- terminals, respectively. When the protease hydrolyzes the peptide bond between the two specific amino acids, it leads to a significant increase in the fluorescence signals. The specific fluorogenic peptide (H-d) produces a high fluorescence signal. A calibration plot was obtained from the changes in the fluorescence intensity against the different concentrations of the viral protease. The lowest limit of detection of this method was 9.7 ± 3 pfu/mL. The cross-reactivity of the SARS-CoV-2-specific peptide was tested against the MERS-CoV which does not affect the fluorescence signal. A significant change in the fluorescence signal with patient samples indicates that this FRET-based assay might be applied for the diagnosis of SARS-CoV-2 patients. Graphical abstract.

Probing high-affinity aptamer binding region and development of aptasensor platform for the detection of cylindrospermopsin.

Cylindrospermopsin (CYN) is one of the most concerning cyanotoxins due to its potential toxicity and spreading to various environments including drinking water. CYN has potential interferences with human and animal metabolic pathways, which influence the functions of organs including liver, kidneys, lungs, etc. CYN is involved in the inhibition of protein synthesis and detachment of ribosomes from the endoplasmic reticulum membrane. It also interacts with soluble proteins, which are associated with protein translations. It is believed that cytochrome 450 is responsible for the rapid toxicity of CYN. Researchers are urged to develop a high-throughput screening method for the detection of CYN in water. Construction of low cost, rapid, and sensitive analytical methods for the detection of CYN is challenging. Here, we used graphene oxide (GO) as the fluorescence sensing platform for probing the high affinity of the short aptamer derived from the wild-type long aptamer-CYN sensing. The biosensor construction involved two steps: first, quenching the fluorescence of fluorescent-labelled truncated aptamer using GO as a quencher and, second, fluorescence recovery in the presence of CYN by competitive binding between the target and GO. One of the truncate aptamers has a 12-fold higher affinity and enhances sensitivity compared to the long aptamer sequence. The limit of detection of the high affinity truncated aptamer is 17 pM which is 6-fold lower than the long aptamer (100 pM). The sensor specifically detects CYN in the presence of other potential interfering toxins. The performance of the sensor was validated using CYN spiked tap water with very good recovery percentage. A rapid and highly sensitive detection of CYN from water resources has been achieved using this method.

Electrochemical determination of zearalenone using a label-free competitive aptasensor.

An electrochemical aptasensor is described for determination of the phytohormone of zearalenone (ZEA). The gold electrode was modified with ZEA via covalent attachment using cysteamine-hydrochloride and 1,4-phenylene diisocyanate linker. A truncated ZEA aptamer with a dissociation constant of 13.4 ± 2.1 nM was used in an aptasensor. The electrochemical property was investigated using square wave voltammetry for monitoring the change in the electron transfer using the ferro/ferricyanide system as redox probe. Under optimal experimental conditions, the response was best measured at a potential of 0.20 V (vs. Ag/AgCl). The signals depended on the competitive mechanism between the immobilised ZEA and free ZEA for the aptamer binding site. The aptasensor works in the range 0.01 to 1000 ng·mL-1 ZEA concentration, with a detection limit of 0.017 ng·mL-1. High degree of cross-reactivity with the other analogues of ZEA was observed, whereas none towards other mycotoxins. The aptasensor was further applied for the determination of ZEA in the extract of maize grain and showed good recovery percentages between 87 and 110%. Graphical abstract Schematic representation of the electrochemical determination of zearalenone based on indirect competitive assay. Step a Immobilisation of ZEA on the surface of gold electrode via covalent attachment, b competition for the ZEA aptamer binding site between immobilised and free ZEA, and c current signal of the binding event based on SWV technique.

Anti-VCAM-1 and Anti-IL4Rα Aptamer-Conjugated Super Paramagnetic Iron Oxide Nanoparticles for Enhanced Breast Cancer Diagnosis and Therapy.

The surface protein overexpressed on cancer cells can be used as biomarkers for early detection of specific diseases. Anti-VCAM-1 and anti-IL4Rα DNA aptamers specific to VCAM-1 and IL4Rα receptors that are overexpressed in 4T1 tumor-bearing mice could be used as potential biomarker for both diagnostic and therapeutic applications in cancer biology. Cell Viability and luciferase assay of 4T1-Luc2 cancer cells in the presence of anti-VCAM-1 ssDNA or anti-IL4Rα RNA aptamers was assessed by monitoring the changes in the absorbance and the fluorescence of Alamar blue dye. The aptamer-conjugated SPIO magnetic beads, used for the selective targeting to tumor sites, were monitored using noninvasive MRI and Bioluminescence imaging (BLI). Cell viability and luciferase assays showed that both anti-VCAM-1 and anti-IL4Rα aptamers favor the depletion of cancer cells and limit tumor progression. Microscopic analyses confirmed that the target specific aptamers significantly trigger tumor cell apoptosis and limit cancer cell growth in vitro. The intravenous injection of SPIO nanoparticle-conjugated aptamers were further confirmed using noninvasive MRI and Bioluminescence imaging. Anti-VCAM1 and anti-IL4Rα aptamers, specific to VCAM-1 and IL4Rα receptors overexpressed in 4T1-Luc2 tumor-bearing mice, were used as diagnostic and therapeutic tools.

An aptamer based fluorometric microcystin-LR assay using DNA strand-based competitive displacement.

The high-affinity region of a truncated aptamer was applied to the development of a sensitive method for the determination of microcystin-LR (MC-LR) using competitive displacement and molecular beacons. In this assay, the fluorophore and quencher labelled complementary sequences of the aptamer are hybridized with the truncated aptamer to form a fluorophore-quencher pair. In the presence of MC-LR, the aptamer duplex dissociates, and the fluorophore-quencher pair is separated. This turn leads to an increase in the yellow fluorescence which is best measured at excitation/emission wavelengths of 555/580 nm. One of the truncated aptamers showed a 50-fold increase in the affinity (0.93 nM) compared to the wild type aptamer (50 nM). The truncated sequence shows considerable cross-reactivity with L congeners but none with other congeners. The assay works in 0.5 to 200 nM MC-LR concentration range. It was applied to spiked tap water samples and gave recoveries around 95 ± 5%. Graphical abstract Schematic representation of a method for determination of microcystin-LR via fluorescence that is induced by competitive displacement of complementary DNA strands in a truncated dsDNA aptamer.

Fluorometric determination of okadaic acid using a truncated aptamer.

Okadaic acid (OKA), a marine toxin produced by dinoflagellates, is responsible for most human diarrhetic shellfish poisoning-associated health disorders. A competitive displacement assay for OKA is described here. An OKA-binding aptamer was truncated with two sequences, one labeled with 6-carboxyfluorescein (FAM), and one with a quencher. On addition of OKA, it will bind to the aptamer and green fluorescence pops up because label and quencher become spatially separated. One of the truncated aptamers exhibis an excellent binding capability (Kd 2.77 nM) for OKA compared to its full-length aptamer (526 nM). The selectivity of the assay was proven by the successful fluorometric determination of OKA in the presence of common diarrhoetic toxins and in shellfish extracts. The detection limit is as low as 39 pg·mL-1. Graphical abstract Schematic representation of the competitive displacement assay for okadaic acid (OKA). The OKA-binding aptamer is truncated with two parts, one labeled with 6-carboxyfluorescein (FAM), and one with a quencher. On addition of OKA, green fluorescence pops up because label and quencher become spatially separated.

Electrochemical selection of a DNA aptamer, and an impedimetric method for determination of the dedicator of cytokinesis 8 by self-assembly of a thiolated aptamer on a gold electrode.

The autosomal recessive-hyper immunoglobulin E syndromes (AR-HIES) are inherited inborn primary immunodeficiency disorders caused mainly by mutations in the dedicator of cytokinesis 8 (DOCK8) gene. A method is described for the selection of DNA aptamers against DOCK8 protein. The selection was performed by using a gold electrode as the solid matrix for immobilization of DOCK8. This enables voltammetric monitoring of the bound DNA after each selection cycle. After eight rounds of selection, high affinity DNA aptamers for DOCK8 were identified with dissociation constants (Kds) ranging from 3.3 to 66 nM. The aptamer which a Kd of 8.8 nM was used in an aptasensor. A gold electrode was modified by self-assembly of the thiolated aptamer, and the response to the DOCK8 protein was detected by monitoring the change in the electron transfer resistance using the ferro/ferricyanide system as a redox probe. The aptasensor works in the 100 pg.mL-1 to 100 ng.mL-1 DOCK8 concentration range, has a detection limit of 81 pg.mL-1 and good selectivity over other proteins in the serum. Graphical abstractSchematic representation of an electrochemical screening protocol for the selection of DNA aptamer against dedicator of cytokinesis 8 protein using electrode as solid support for target immobilization.

A rapid colorimetric immunoassay for the detection of pathogenic bacteria on poultry processing plants using cotton swabs and nanobeads.

The work describes a simple cotton swab-based colorimetric immunoassay as a rapid screening tool for pathogenic bacteria on poultry processing plants. This immunosensing platform can be used for the detection of pathogens present on surfaces such as glass, stainless steel and chicken meat. Unlike the reported assays, here, cotton swab plays dual function: as a sample collector from the solid surfaces and as detection platform. The immunoassay was tested for the detection of 4 different bacteria; Salmonella typhimurium, Salmonella enteritidis, Staphylococcus aureus and Campylobacter jejuni. The immunoassays were fabricated by immobilizing specific antibody for each bacterium on a cotton swab that is used to recover the cells from contaminated surfaces. Then, a sandwich immunoassay was developed by immersing the cotton swab in different colored nanobead-conjugated antibody solutions which corresponds to different bacteria. The immunoassays response was detected colorimetrically by following the change in the color intensity produced by the nanobeads due to the specific binding on the cotton swab. This simple colorimetric assay is very sensitive with a detection limit of 10 cfu.mL-1. Furthermore, no significant cross reactivity of the immunoassays with non specific bacteria was observed indicating good selectivity of the immunoassays. This simple, disposable and easy-to- use colorimetric platform shows great promise as rapid qualitative and semi quantitative detection tool for microorganisms on food processing plants and other surfaces. Graphical abstract Schematic of the sandwich colorimetric immunosensor for the detection of pathogenic bacteria on poultry processing plants using cotton swabs and nanobeads.

High affinity truncated DNA aptamers for the development of fluorescence based progesterone biosensors.

Aptamers have shown a number of potential applications in sensing and therapeutic due to the high affinity and specificity towards their target molecules. Not all the nucleotides in the full length aptamers are involved in the binding with their targets. The non-binding domain of the aptamer may affect the binding affinity of the aptamer-target complex. Mapping the aptamer binding region could increase the affinity and the specificity. In this paper, we designed aptamer-based fluorescence sensors from a truncated progesterone (P4) aptamer. Then, fluorescein and quencher labelled aptamer complementary oligonucleotide sequences were hybridized to the truncated aptamer at different sites to form duplex structures. We used fluorescence-quencher pair displacement assay upon progesterone binding for the determination of P4. One of the truncated sequences has shown high binding affinity with 16 fold increase in the dissociation constant, Kd (2.1 nM) compared to the original aptamer. The aptasensor was highly selective for P4 against similar compounds such as 17-ß estradiol, bisphenol-A and vitamin D. The sensor has been applied for the detection of P4 in spiked tap water and in urine samples showing good recovery. This new developed aptamer-based fluorescence biosensor can be applied in food, pharmaceutical, and clinical industries.

Development of magnetic nanoparticle based calorimetric assay for the detection of bovine mastitis in cow milk.

Mastitis in dairy cattle is an inflammatory reaction of the udder tissue. Mastitis increases plasmin levels, leading to an increased proteolysis of milk proteins such as casein, resulting in a significant decrease in milk quality and related dairy products. Due to its key-role in mastitis, we used plasmin proteolytic activity as a biomarker for the detection of mastitis in bovine mastitic milk. Inspired by earlier studies on protease activity using mastitic milk samples, we developed a simple colorimetric assay to distinguish mastitic milk from milk derived from healthy animals. The plasmin substrate coupled to magnetic nanoparticles form a black self-assembled monolayer on a gold sensor surface. In the presence of increased levels of plasmin, the substrate is cleaved and the peptide fragment attached to the magnetic beads, will be attracted by the magnet which is present under the sensor strips revealing the golden surface. We found the area of the golden color surface proportional to plasmin activity. The sensitivity of this method was determined to be 1 ng/ml of plasmin in vitro. Next, we tested the biosensor using mastitis positive milk of which infection is confirmed by bacterial cultures. This newly developed colorimetric biosensor has high potential in applications for the diagnosis of mastitis with potential spin offs to health, food and environmental sectors.

Fluorometric graphene oxide-based detection of Salmonella enteritis using a truncated DNA aptamer.

The work describes a fluorescence-based study for mapping the highest affinity truncated aptamer from the full length sequence and its integration in a graphene oxide platform for the detection of Salmonella enteriditis. To identify the best truncated sequence, molecular beacons and a displacement assay design are applied. In the fluorescence displacement assay, the truncated aptamer was hybridized with fluorescein and quencher-labeled complementary sequences to form a fluorescence/quencher pair. In the presence of S. enteritidis, the aptamer dissociates from the complementary labeled oligonucleotides and thus, the fluorescein/quencher pair becomes physically separated. This leads to an increase in fluorescence intensity. One of the truncated aptamers identified has a 2-fold lower dissociation constant (3.2 nM) compared to its full length aptamer (6.3 nM). The truncated aptamer selected in this process was used to develop a fluorometric graphene oxide (GO) based assay. If fluorescein-labeled aptamer is adsorbed on GO via π stacking interaction, fluorescence is quenched. However, in the presence of target (S. enteriditis), the labeled aptamers is released from surface to form a stable complex with the bacteria and fluorescence is restored, depending on the quantity of bacteria being present. The resulting assay has an unsurpassed detection limit of 25 cfu·mL-1 in the best case. The cross reactivity to Salmonella typhimurium, Staphylococcus aureus and Escherichia coli is negligible. The assay was applied to analyze doped milk samples for and gave good recovery. Thus, we believe that the truncated aptamer/graphene oxide platform is a potential tool for the detection of S. Enteritidis. Graphical abstract Fluorescently labelled aptamer against Salmonella enteritidis was adsorbed on the surface of graphene oxide by π-stacking interaction. This results in quenching of the fluorescence of the label. Addition of Salmonella enteritidis restores fluorescence, and this effect is used for quantification of this food-borne pathogen.

Fluorescence monitoring of riboswitch transcription regulation using a dual molecular beacon assay.

Riboswitches are mRNA elements that specifically bind cellular metabolites and control gene expression by modifying their structure. As riboswitches often control essential genes in pathogenic bacteria, riboswitches have been proposed as new targets for antibiotics. High-throughput screening provides a powerful approach to identify riboswitch ligand analogs that could act as powerful antibacterial drugs. Biochemical assays have already been used to find riboswitch-binding analogs, but those methods do take into account the transcriptional context for riboswitch regulation. As the importance of co-transcriptional ligand binding has been shown for several riboswitches, it is vital to develop an assay that screens riboswitch-binding analogs during the transcriptional process. Here, we describe the development of a dual molecular beacon system monitoring the transcriptional regulation activity of the Bacillus subtilis pbuE adenine riboswitch. This system relies on two molecular beacons that enable the monitoring of transcription efficiency, as well as the regulatory activity of the riboswitch. Different analogs were tested using our system, and a good correlation was observed between riboswitch activity and reported metabolite affinities. This method is specific, reliable and could be applied at the high-throughput level for the identification of new potential antibiotics targeting any riboswitch-regulating gene expression at the mRNA level.

Rapid Colorimetric Quantita2tive Portable Platform for Detection of Brucella melitensis Based on a Fluorescence Resonance Energy Transfer Assay and Nanomagnetic Particles.

Brucellosis is a bacterial zoonotic disease that requires major attention for both health and financial facilities in many parts of the world including the Mediterranean and the Middle East. The existing gold standard diagnosis relies on the culturing technique, which is costly and time-consuming with a duration of up to 45 days. The Brucella protease biosensor represents a new detection approach that will lead to low-cost point-of-care devices for sensitive Brucella detection. In addition, the described diagnostic device is portable and simple to operate by a nurse or non-skilled clinician making it appropriate for the low-resource setting. In this study, we rely on the total extracellular protease proteolytic activity on specific peptide sequences identified using the FRET assay by high-throughput screening from the library of peptide (96 short peptides such as dipeptides and tripeptides) substrates for Brucella melitensis (B. melitensis). The B. melitensis-specific protease substrate was utilized in the development of the paper-based colorimetric assay. Two specific and highly active dipeptide substrates were identified (FITC-Ahx-K-r-K-Ahx-DABCYL and FITC-Ahx-R-r-K-Ahx-DABCYL). The peptide-magnetic bead nanoprobe sensors developed based on these substrates were able to detect the Brucella with LOD as low as 1.5 × 102 and 1.5 × 103 CFU/mL using K-r dipeptide and R-r dipeptide substrates, respectively, as the recognition element. The samples were tested using this sensor in few minutes. Cross-reactivity studies confirmed that the other proteases extracted from closely related pathogens have no significant effect on the sensor. To the best of our knowledge, this assay is the first assay that can be used with low-cost, rapid, direct, and visual detection of B. melitensis.

Bioengineered Organoids Offer New Possibilities for Liver Cancer Studies: A Review of Key Milestones and Challenges.

Hepatic cancer is widely regarded as the leading cause of cancer-related mortality worldwide. Despite recent advances in treatment options, the prognosis of liver cancer remains poor. Therefore, there is an urgent need to develop more representative in vitro models of liver cancer for pathophysiology and drug screening studies. Fortunately, an exciting new development for generating liver models in recent years has been the advent of organoid technology. Organoid models hold huge potential as an in vitro research tool because they can recapitulate the spatial architecture of primary liver cancers and maintain the molecular and functional variations of the native tissue counterparts during long-term culture in vitro. This review provides a comprehensive overview and discussion of the establishment and application of liver organoid models in vitro. Bioengineering strategies used to construct organoid models are also discussed. In addition, the clinical potential and other relevant applications of liver organoid models in different functional states are explored. In the end, this review discusses current limitations and future prospects to encourage further development.

Recent Advances in Biosensor Technology for Early-Stage Detection of Hepatocellular Carcinoma-Specific Biomarkers: An Overview.

Hepatocellular carcinoma is currently the most common malignancy of the liver. It typically occurs due to a series of oncogenic mutations that lead to aberrant cell replication. Most commonly, hepatocellular carcinoma (HCC) occurs as a result of pre-occurring liver diseases, such as hepatitis and cirrhosis. Given its aggressive nature and poor prognosis, the early screening and diagnosis of HCC are crucial. However, due to its plethora of underlying risk factors and pathophysiologies, patient presentation often varies in the early stages, with many patients presenting with few, if any, specific symptoms in the early stages. Conventionally, screening and diagnosis are performed through radiological examination, with diagnosis confirmed by biopsy. Imaging modalities tend to be limited by their requirement of large, expensive equipment; time-consuming operation; and a lack of accurate diagnosis, whereas a biopsy's invasive nature makes it unappealing for repetitive use. Recently, biosensors have gained attention for their potential to detect numerous conditions rapidly, cheaply, accurately, and without complex equipment and training. Through their sensing platforms, they aim to detect various biomarkers, such as nucleic acids, proteins, and even whole cells extracted by a liquid biopsy. Numerous biosensors have been developed that may detect HCC in its early stages. We discuss the recent updates in biosensing technology, highlighting its competitive potential compared to conventional methodology and its prospects as a tool for screening and diagnosis.

Emerging role of exosomal microRNA in liver cancer in the era of precision medicine; potential and challenges.

Exosomal microRNAs (miRNAs) have great potential in the fight against hepatocellular carcinoma (HCC), the fourth most common cause of cancer-related death worldwide. In this study, we explored the various applications of these small molecules while analyzing their complex roles in tumor development, metastasis, and changes in the tumor microenvironment. We also discussed the complex interactions that exist between exosomal miRNAs and other non-coding RNAs such as circular RNAs, and show how these interactions coordinate important biochemical pathways that propel the development of HCC. The possibility of targeting exosomal miRNAs for therapeutic intervention is paramount, even beyond their mechanistic significance. We also highlighted their growing potential as cutting-edge biomarkers that could lead to tailored treatment plans by enabling early identification, precise prognosis, and real-time treatment response monitoring. This thorough analysis revealed an intricate network of exosomal miRNAs lead to HCC progression. Finally, strategies for purification and isolation of exosomes and advanced biosensing techniques for detection of exosomal miRNAs are also discussed. Overall, this comprehensive review sheds light on the complex web of exosomal miRNAs in HCC, offering valuable insights for future advancements in diagnosis, prognosis, and ultimately, improved outcomes for patients battling this deadly disease.