Optofluidic Sensor Based on Polymer Optical Microresonators for the Specific, Sensitive and Fast Detection of Chemical and Biochemical Species.

The accurate, rapid, and specific detection of DNA strands in solution is becoming increasingly important, especially in biomedical applications such as the trace detection of COVID-19 or cancer diagnosis. In this work we present the design, elaboration and characterization of an optofluidic sensor based on a polymer-based microresonator which shows a quick response time, a low detection limit and good sensitivity. The device is composed of a micro-racetrack waveguide vertically coupled to a bus waveguide and embedded within a microfluidic circuit. The spectral response of the microresonator, in air or immersed in deionised water, shows quality factors up to 72,900 and contrasts up to 0.9. The concentration of DNA strands in water is related to the spectral shift of the microresonator transmission function, as measured at the inflection points of resonance peaks in order to optimize the signal-over-noise ratio. After functionalization by a DNA probe strand on the surface of the microresonator, a specific and real time measurement of the complementary DNA strands in the solution is realized. Additionally, we have inferred the dissociation constant value of the binding equilibrium of the two complementary DNA strands and evidenced a sensitivity of 16.0 pm/µM and a detection limit of 121 nM.

Antimicrobial Activity of Cationic Poly(3-hexylthiophene) Nanoparticles Coupled with Dual Fluorescent and Electrochemical Sensing: Theragnostic Prospect.

Designing therapeutic and sensor materials to diagnose and eliminate bacterial infections remains a significant challenge for active theragnostic nanoprobes. In the present work, fluorescent/electroactive poly(3-hexylthiophene) P3HT nanoparticles (NPs) stabilized with quaternary ammonium salts using cetyltrimethylammonium bromide (CTAB), (CTAB-P3HT NPs) were prepared using a simple mini-emulsion method. The morphology, spectroscopic properties and electronic properties of CTAB-P3HT NPs were characterized by DLS, zeta potential, SEM, TEM, UV-vis spectrophotometry, fluorescence spectroscopy and electrochemical impedance spectroscopy (EIS). In an aqueous solution, CTAB-P3HT NPs were revealed to be uniformly sized, highly fluorescent and present a highly positively charged NP surface with good electroactivity. Dual detection was demonstrated as the binding of the bacteria to NPs could be observed by fluorescence quenching as well as by the changes in EIS. Binding of E. coli to CTAB-P3HT NPs was demonstrated and LODs of 5 CFU/mL and 250 CFU/mL were obtained by relying on the fluorescence spectroscopy and EIS, respectively. The antimicrobial activity of CTAB-P3HT NPs on bacteria and fungi was also studied under dark and nutritive conditions. An MIC and an MBC of 2.5 µg/mL were obtained with E. coli and with S. aureus, and of 0.312 µg/mL with C. albicans. Additionally a good biocompatibility toward normal human cells (WI38) was observed, which opens the way to their possible use as a therapeutic agent.

Sensor based on redox conjugated poly(para-phenylene) for the simultaneous detection of dopamine, ascorbic acid, and uric acid in human serum sample.

An electrochemical sensor for the individual and the simultaneous detection of dopamine (DA), ascorbic acid (AA), and uric acid (UA) based on redox conjugated "poly(para-phenylene)" (Fc-ac-PPP) bearing ferrocene and carboxylic acid in lateral position has been developed. The electrochemical characterization of the sensor has been studied with cyclic voltammetry (CV), differential pulse voltammetry (DPV), and chronoamperometry (CA). We highlighted that the catalytic activity of the Fc-ac-PPP polymer provided by its redox electrochemical properties and chemical structure allows the electrochemical detection of DA, AA, and UA. We demonstrated that the sensor provides high sensitivity and selective signal in the coexistence of DA, AA, and UA within a short time. Low detection limits and wide linear ranges of detection have been demonstrated respectively for DA 3 × 10-10 M (1 nM-10 µM), AA 1.6 × 10-8 M (0.1 µM-1 mM), and UA 1 × 10-8 M (0.1 µM-1 mM). In addition, the sensor has been successfully applied to determine DA in urine and human serum samples even in the presence of high concentrations of AA and UA. This sensor could be a powerful device for the detection of other electroactive compounds thanks to its high catalytic properties and chemical structure. Graphical abstract.

Novel N-substituted 5-phosphate-d-arabinonamide derivatives as strong inhibitors of phosphoglucose isomerases: Synthesis, structure-activity relationship and crystallographic studies.

Phosphoglucose isomerase (PGI) is a cytosolic enzyme that catalyzes the reversible interconversion of d-glucose 6-phosphate and d-fructose 6-phosphate in glycolysis. Outside the cell, PGI is also known as autocrine motility factor (AMF), a cytokine secreted by a large variety of tumor cells that stimulates motility of cancer cells in vitro and metastases development in vivo. Human PGI and AMF are strictly identical proteins both in terms of sequence and 3D structure, and AMF activity is known to involve, at least in part, the enzymatic active site. Hence, with the purpose of finding new strong AMF-PGI inhibitors that could be potentially used as anticancer agents and/or as bioreceptors for carbohydrate-based electrochemical biosensors, we report in this study the synthesis and kinetic evaluation of several new human PGI inhibitors derived from the synthon 5-phospho-d-arabinono-1,4-lactone. Although not designed as high-energy intermediate analogue inhibitors of the enzyme catalyzed isomerization reaction, several of these N-substituted 5-phosphate-d-arabinonamide derivatives appears as new strong PGI inhibitors. For one of them, we report its crystal structure in complex with human PGI at 2.38 Å. Detailed analysis of its interactions at the active site reveals a new binding mode and shows that human PGI is relatively tolerant for modified inhibitors at the "head" C-1 part, offering promising perspectives for the future design of carbohydrate-based biosensors.

Nanocomposite based on Poly (para-phenylene)/Chemical Reduced Graphene Oxide as a Platform for Simultaneous Detection of Ascorbic Acid, Dopamine and Uric Acid.

In this study, an efficient and simple designed nanohybrid created for individual and simultaneous detection of ascorbic acid (AA), dopamine (DA) and uric acid (UA). This nanohybrid is a combination of reduced graphene oxide (CRGO) and redox poly(para-phenylene) (Fc-ac-PP) modified in a lateral position with ferrrocenyl group CRGO/Fc-ac-PPP. The CRGO/Fc-ac-PPP nanohybrid demonstrated a synergistic effect resulting in a large conductivity, surface area and catalytic properties provided by the redox attached ferrocene. Moreover, this nanocomposite is able to detect individually as well as simultaneously AA, DA and UA in a co-existence system with defined and separated redox peaks oxidation. The linear response ranges for AA, DA and UA, when detected simultaneously, are 0.1-10000 µM, 0.0001-1000 µM and 0.1-10000 µM, respectively, and the detection limits (S/N = 3) are 0.046 µM, 0.2 nM and 0.013 µM, respectively. The proposed sensor shown satisfactory results when applied to real spiked urine samples for measuring the abnormal high or lowconcentration of AA, DA and UA in vivo.

E-DNA sensor of Mycobacterium tuberculosis based on electrochemical assembly of nanomaterials (MWCNTs/PPy/PAMAM).

Two-step electrochemical patterning methods have been employed to elaborate composite nanomaterials formed with multiwalled carbon nanotubes (MWCNTs) coated with polypyrrole (PPy) and redox PAMAM dendrimers. The nanomaterial has been demonstrated as a molecular transducer for electrochemical DNA detection. The nanocomposite MWCNTs-PPy has been formed by wrapping the PPy film on MWCNTs during electrochemical polymerization of pyrrole on the gold electrode. The MWCNTs-PPy layer was modified with PAMAM dendrimers of fourth generation (PAMAM G4) with covalent bonding by electro-oxidation method. Ferrocenyl groups were then attached to the surface as a redox marker. The electrochemical properties of the nanomaterial (MWCNTs-PPy-PAMAM-Fc) were studied using both square wave voltammetry and cyclic voltammetry to demonstrate efficient electron transfer. The nanomaterial shows high performance in the electrochemical detection of DNA hybridization leading to a variation in the electrochemical signal of ferrocene with a detection limit of 0.3 fM. Furthermore, the biosensor demonstrates ability for sensing DNA of rpoB gene of Mycobacterium tuberculosis in real PCR samples. Developed biosensor was suitable for detection of sequences with a single nucleotide polymorphism (SNP) T (TCG/TTG), responsible for resistance of M. tuberculosis to rifampicin drug, and discriminating them from wild-type samples without such mutation. This shows potential of such systems for further application in pathogens diagnostic and therapeutic purpose.

Electrochemical detection of the oligomerization of PB1-F2 influenza A virus protein in infected cells.

PB1-F2 is a nonstructural accessory protein of Influenza A virus described to enhance the mortality and the morbidity of the virus in a host-dependent manner. In this work, an electrochemical biosensor based on an immunodetection system was developed to follow the oligomerization of PB1-F2 during the viral cycle. The immunosensor was based on conductive polypyrrole modified with ferrocenyl groups as a redox marker for enhancing signal detection. Antibodies specific for monomeric or oligomeric PB1-F2 forms were immobilized on polypyrrole matrix via biotin/streptavidin layer. We demonstrated that this electrochemical biosensor sensitively detects PB1-F2 in both conformational forms. The linear range extends from 5 nM to 1.5 µM and from 5 nM to 0.5 µM for monomeric and oligomeric PB1-F2, respectively. The calculated limit of detection was 0.42 nM for monomeric PB1-F2 and 16 nM for oligomers. The biosensor platform allows the detection and quantification of PB1-F2 in lysates of infected cells during viral cycle. We show that at early stages of viral cycle, PB1-F2 is mainly monomeric but switched to amyloid-like structures at a later stage of infection. The quantification of two protein structural forms points out that PB1-F2 expression profiles and kinetics of oligomerization are cell-type-dependent.

Impedimetric Detection of Cancer Markers Based on Nanofiber Copolymers.

The sensitive determination of folate receptors (FRs) in the early stages of cancer is of great significance for controlling the progression of cancerous cells. Many folic acid (FA)-based electrochemical biosensors have been utilized to detect FRs with promising performances, but most were complicated, non-reproducible, non-biocompatible, and time and cost consuming. Here, we developed an environmentally friendly and sensitive biosensor for FR detection. We proposed an electrochemical impedimetric biosensor formed by nanofibers (NFs) of bio-copolymers prepared by electrospinning. The biosensor combines the advantages of bio-friendly polymers, such as sodium alginate (SA) and polyethylene oxide (PEO) as an antifouling polymer, with FA as a biorecognition element. The NF nanocomposites were characterized using various techniques, including SEM, FTIR, zeta potential (ZP), cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS). We evaluated the performance of the NF biosensor using EIS and demonstrated FR detection in plasma with a limit of detection of 3 pM. Furthermore, the biosensor showed high selectivity, reliability, and good stability when stored for two months. This biosensor was constructed from 'green credentials' holding polymers that are highly needed in the new paradigm shift in the medical industry.

Electrochemical aptasensor of human cellular prion based on multiwalled carbon nanotubes modified with dendrimers: a platform for connecting redox markers and aptamers.

The present work aims to develop an electrochemical biosensor based on aptamer able to detect human cellular prions PrP(C) as a model biomarker of prion disease with high sensitivity. We designed the biosensor using multiwalled carbon nanotubes (MWCNTs) modified with polyamidoamine dendrimers of fourth generation (PAMAM G4) which in turn were coupled to DNA aptamers used as bioreceptors. Electrochemical signal was detected by a ferrocenyl redox marker incorporated between the dendrimers and aptamers interlayer. MWCNTs, thanks to their nanostructure organization and electrical properties, allow the distribution of aptamers and redox markers over the electrode surface. We demonstrated that the interaction between aptamers and prion proteins leads to variation in the electrochemical signal of the ferrocenyl group. High sensitivity with a detection limit of 0.5 pM and a wide linear range of detection from 1 pM to 10 µM has been demonstrated. Detection of PrP(C) in spiked blood plasma has been achieved in the same range of concentrations as for detection of PrP(C) in buffer. The sensor demonstrated a recovery of minimum 85% corresponding to 1 nM PrP(C) and a maximum of 127% corresponding to 1 pM PrP(C).

π-Conjugated Polymer Nanoparticles from Design, Synthesis to Biomedical Applications: Sensing, Imaging, and Therapy.

In the past decade, π-conjugated polymer nanoparticles (CPNs) have been considered as promising nanomaterials for biomedical applications, and are widely used as probe materials for bioimaging and drug delivery. Due to their distinctive photophysical and physicochemical characteristics, good compatibility, and ease of functionalization, CPNs are gaining popularity and being used in more and more cutting-edge biomedical sectors. Common synthetic techniques can be used to synthesize CPNs with adjustable particle size and dispersion. More importantly, the recent development of CPNs for sensing and imaging applications has rendered them as a promising device for use in healthcare. This review provides a synopsis of the preparation and functionalization of CPNs and summarizes the recent advancements of CPNs for biomedical applications. In particular, we discuss their major role in bioimaging, therapeutics, fluorescence, and electrochemical sensing. As a conclusion, we highlight the challenges and future perspectives of biomedical applications of CPNs.

Label free electrochemical sensors for Pb(II) detection based on hemicellulose extracted from Opuntia Ficus Indica cactus.

We aim to develop an electrochemical sensor for a divalent metal ion (lead II), a highly toxic water contaminant. We explore a sensor formed with a hemicellulose polysaccharide extracted from the Opuntia Ficus Indica cactus associated with agarose as a sensitive layer deposited on a gold electrode. This sensor combines the functional groups of hemicellulose that could form a complex with metal ions and agarose with gelling properties to form a stable membrane. The sensor demonstrated a loading ability of Pb2+, with higher affinity compared to other metal ions such as Hg2+, Ni2+, and Cu2+, thanks to the chemical structure of hemicellulose. The detection was measured by square wave voltammetry based on a well-defined redox peak of the metal ions. The sensor shows high sensitivity towards Pb2+ with a detection limit of 1.3 fM. The application in river and sea water using the standard addition method for lead detection was studied.

Characterization of structure and activity of garlic peroxidase (POX(1B)).

Structural characterization and study of the activity of new POX(1B) protein from garlic which has a high peroxidase activity and can be used as a biosensor for the detection of hydrogen peroxide and phenolic compounds were performed and compared with the findings for other heme peroxidases. The structure-function relationship was investigated by analysis of the spectroscopic properties and correlated to the structure determined by a new generation of high-performance hybrid mass spectrometers. The reactivity of the enzyme was analyzed by studies of the redox activity toward various ligands and the reactivity with various substrates. We demonstrated that, in the case of garlic peroxidase, the heme group is pentacoordinated, and has an histidine as a proximal ligand. POX(1B) exhibited a high affinity for hydrogen peroxide as well as various reducing cosubstrates. In addition, high enzyme specificity was demonstrated. The k(cat) and K(M) values were 411 and 400 mM(-1) s(-1) for 3,3',5,5'-tetramethylbenzidine and 2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid), respectively. Furthermore, the reduction of nitro compounds in the presence of POX(1B) was demonstrated by iron(II) nitrosoalkane complex assay. In addition, POX(1B) showed a great potential for application for drug metabolism since its ability to react with 1-nitrohexane in the presence of sodium dithionite was demonstrated by the appearance of a characteristic Soret band at 411 nm. The high catalytic efficiency obtained in the case of the new garlic peroxidase (POX(1B)) is suitable for the monitoring of different analytes and biocatalysis.

MoS2/PPy Nanocomposite as a Transducer for Electrochemical Aptasensor of Ampicillin in River Water.

We report the design of an electrochemical aptasensor for ampicillin detection, which is an antibiotic widely used in agriculture and considered to be a water contaminant. We studied the transducing potential of nanostructure composed of MoS2 nanosheets and conductive polypyrrole nanoparticles (PPyNPs) cast on a screen-printed electrode. Fine chemistry is developed to build the biosensors entirely based on robust covalent immobilizations of naphthoquinone as a redox marker and the aptamer. The structural and morphological properties of the nanocomposite were studied by SEM, AFM, and FT-IR. High-resolution XPS measurements demonstrated the formation of a binding between the two nanomaterials and energy transfer affording the formation of heterostructure. Cyclic voltammetry and electrochemical impedance spectroscopy were used to analyze their electrocatalytic properties. We demonstrated that the nanocomposite formed with PPyNPs and MoS2 nanosheets has electro-catalytic properties and conductivity leading to a synergetic effect on the electrochemical redox process of the redox marker. Thus, a highly sensitive redox process was obtained that could follow the recognition process between the apatamer and the target. An amperometric variation of the naphthoquinone response was obtained regarding the ampicillin concentration with a limit of detection (LOD) of 10 pg/L (0.28 pM). A high selectivity towards other contaminants was demonstrated with this biosensor and the analysis of real river water samples without any treatment showed good recovery results thanks to the antifouling properties. This biosensor can be considered a promising device for the detection of antibiotics in the environment as a point-of-use system.

E-DNA biosensors of M. tuberculosis based on nanostructured polypyrrole.

The objective of this paper is to demonstrate the potential of nanostructured polypyrrole formed by template free as platform for amperometric detection of DNA. The nanowires of polypyrrole (nw-PPy) are formed through electrochemical polymerization and chemically modified by electrochemical oxidation of ethylene diamine or dendrimers PAMAM to obtain aminated surface. The DNA probe and ferrocenyl group, as redox reporter, were covalently linked to the surface of nw-PPy. The chemical structure of nanostructured platform was characterized through SEM, FT-IR and XPS and the electrochemical properties through cyclic voltammetry and electrochemical impedance spectroscopy (EIS). We show that the properties of nw-PPy such as, hydrophilic character and large surface area have large effect on the electronic properties. Thus, the electrochemical performance is increased compared to others nanomaterials considering the obtained value of the rate of electron transfer of 18 s-1. These properties allow enhanced DNA sensing where detection limit of 0.36 atomolar without any amplification step. The biosensor can be applied in detection of genomic DNA of Mycobacterium tuberculosis and the mutated one which present the resistance to rifampicin and large selectivity was demonstrated. We believe that nw-PPy modified with redox marker is a promising platform for electrochemical biosensors and can be applied for various diagnosis prospects.

Electrochemical Biosensors for Detection of MicroRNA as a Cancer Biomarker: Pros and Cons.

Cancer is the second most fatal disease in the world and an early diagnosis is important for a successful treatment. Thus, it is necessary to develop fast, sensitive, simple, and inexpensive analytical tools for cancer biomarker detection. MicroRNA (miRNA) is an RNA cancer biomarker where the expression level in body fluid is strongly correlated to cancer. Various biosensors involving the detection of miRNA for cancer diagnosis were developed. The present review offers a comprehensive overview of the recent developments in electrochemical biosensor for miRNA cancer marker detection from 2015 to 2020. The review focuses on the approaches to direct miRNA detection based on the electrochemical signal. It includes a RedOx-labeled probe with different designs, RedOx DNA-intercalating agents, various kinds of RedOx catalysts used to produce a signal response, and finally a free RedOx indicator. Furthermore, the advantages and drawbacks of these approaches are highlighted.

Dispersible Conjugated Polymer Nanoparticles as Biointerface Materials for Label-Free Bacteria Detection.

Fast and efficient identification of bacterial pathogens in water and biological fluids is an important issue in medical, food safety, and public health concerns that requires low-cost and efficient sensing strategies. Impedimetric sensors are promising tools for monitoring bacteria detection because of their reliability and ease-of-use. We herein report a study on new biointerface-based amphiphilic poly(3-hexylthiophene)-b-poly(3-triethylene-glycol-thiophene), P3HT-b-P3TEGT, for label-free impedimetric detection of Escherichia coli (E. coli). This biointerface is fabricated by the self-assembly of P3HT-b-P3TEGT into core-shell nanoparticles, which was further decorated with mannose, leading to an easy-to-use solution-processable nanoparticle material for biosensing. The hydrophilic block P3TEGT promotes antifouling and prevents nonspecific interactions, while improving the ionic and electronic transport properties, thus enhancing the electrochemical-sensing capability in aqueous solution. Self-assembly and micelle formation of P3HT-b-P3TEGT were analyzed by 2D-NMR, Fourier transform infrared, dynamic light scattering, contact angle, and microscopy characterizations. Detection of E. coli was characterized and evaluated using electrochemical impedance spectroscopy and optical and scanning electron microscopy techniques. The sensing layer based on the mannose-functionalized P3HT-b-P3TEGT nanoparticles demonstrates targeting ability toward E. coli pili protein with a detection range from 103 to 107 cfu/mL, and its selectivity was studied with Gram(+) bacteria. Application to real samples was performed by detection of bacteria in tap and the Nile water. The approach developed here shows that water/alcohol-processable-functionalized conjugated polymer nanoparticles are suitable for use as electrode materials, which have potential application in fabrication of a low-cost, label-free impedimetric biosensor for the detection of bacteria in water.

E-DNA detection of rpoB gene resistance in Mycobacterium tuberculosis in real samples using Fe3O4/polypyrrole nanocomposite.

In this work, we achieved the selective detection of wild and mutated rpoB gene in M. tuberculosis using an electrochemical DNA (E-DNA) sensor based on polypyrrole/Fe3O4 nanocomposite bearing redox naphthoquinone tag on PAMAM (spaNQ/PAMAM/PPy/Fe3O4). The hybridization between a given probe and the complementary DNA target induced a large decrease in the naphthoquinone redox signal as measured by SWV and no cross-hybridization with single nucleotide mismatch DNA target occurred. Thanks to the catalytic properties of iron oxide nanoparticles combined with conducting properties of polypyrrole platform, we demonstrated that the transducing system allowed the detection of 1 fM of DNA target in a 50-µL drop corresponding to 3 × 104 copies of DNA. The sensor was able to detect the rpoB gene in PCR-amplified samples of genomic DNA and could also discriminate between the wild type rpoB gene and a single nucleotide mutated rpoB gene that provides resistance to rifampicin. Furthermore, the sensor could selectively detect the wild and mutant DNA in genomic samples without PCR amplification.

Impedimetric array in polymer microfluidic cartridge for low cost point-of-care diagnostics.

Deep Vein Thrombosis and pulmonary embolism (DVT/PE) is one of the most common causes of unexpected death for hospital in-patients. D-dimer is used as a biomarker within blood for the diagnosis of DVT/PE. We report a low-cost microfluidic device with a conveniently biofunctionalised interdigitated electrode (IDE) array and a portable impedimetric reader as a point-of-care (POC) device for the detection of D-dimer to aid diagnosis of DVT/PE. The IDE array elements, fabricated on a polyethylenenaphtalate (PEN) substrate, are biofunctionalised in situ after assembly of the microfluidic device by electropolymerisation of a copolymer of polypyrrole to which is immobilised a histidine tag anti-D-Dimer antibody. The most consistent copolymer films were produced using chronopotentiometry with an applied current of 5µA for a period of 50 s using a two-electrode system. The quality of the biofunctionalisation was monitored using optical microscopy, chronopotentiometry curves and impedimetric analysis. Measurement of clinical plasma sample with a D-dimer at concentration of 437 ng/mL with 15 biofunctionalised IDE array electrodes gave a ratiometric percentage of sample reading against the blank with an average value of 124 ±â€¯15 at 95% confidence. We have demonstrated the concept of a low cost disposable microfluidic device with a receptor functionalised on the IDE array for impedimetric detection towards POC diagnostics. Changing the receptor on the IDE array would allow this approach to be used for the direct detection of a wide range of analytes in a low cost manner.

Direct E-DNA sensor of Mycobacterium tuberculosis mutant strain based on new nanocomposite transducer (Fc-ac-OMPA/MWCNTs).

Direct DNA sensor based on new nanocomposite materials (Fc-ac-OMPA/MWCNTs) has been investigated. This nanocomposite was formed by combining the redox oligomer "oligo-methoxy-phenyl-acetonitrile" (Fc-ac-OMPA) and the MWCNTs via a simple π-π stacking interaction in the aim to ameliorate the biosensor performance. The redox indicator and the functional groups of the redox oligomer have been used for monitoring the electrochemical behavior and the flexibility for direct covalent attachment of Hepatitis C DNA probe. This nanocomposite shows high performance of DNA hybridization with a detection limit of 0.08 fmol L-1. Moreover, the biosensor was applied for the detection of pathogenic bacterium such as DNA from Mycobacterium tuberculosis strand. Developed biosensor has been able to detect a single nucleotide polymorphism (SNP)T (TCG/ TTG) which confers resistance of M. tuberculosis to rifampicin drug.

Carbohydrate-based electrochemical biosensor for detection of a cancer biomarker in human plasma.

Autocrine motility factor (AMF) is a tumor-secreted cytokine that stimulates tumor cell motility in vitro and metastasis in vivo. AMF could be detected in serum or urine of cancer patients with worse prognosis. Reported as a cancer biomarker, AMF secretion into body fluids might be closely related to metastases formation. In this study, a sensitive and specific carbohydrate-based electrochemical biosensor was designed for the detection and quantification of a protein model of AMF, namely phosphoglucose isomerase from rabbit muscle (RmPGI). Indeed, RmPGI displays high homology with AMF and has been shown to have AMF activity. The biosensor was constructed by covalent binding of the enzyme substrate d-fructose 6-phosphate (F6P). Immobilization was achieved on a gold surface electrode following a bottom-up approach through an aminated surface obtained by electrochemical patterning of ethylene diamine and terminal amine polyethylene glycol chain to prevent non-specific interactions. Carbohydrate-protein interactions were quantified in a range of 10 fM to 100nM. Complex formation was analyzed through monitoring of the redox couple Fe2+/Fe3+ by electrochemical impedance spectroscopy and square wave voltammetry. The F6P-biosensor demonstrates a detection limit of 6.6 fM and high selectivity when compared to other non-specific glycolytic proteins such as d-glucose-6-phosphate dehydrogenase. Detection of protein in spiked plasma was demonstrated and accuracy of 95% is obtained compared to result obtained in PBS (phosphate buffered saline). F6P-biosensor is a very promising proof of concept required for the design of a carbohydrate-based electrochemical biosensor using the enzyme substrate as bioreceptor. Such biosensor could be generalized to detect other protein biomarkers of interest.