A mini review and hypothesis for coronavirus detection using photonics: surface enhanced Raman scattering and fluorescence resonance energy transfer

COVID-19 has devastated numerous nations around the world and has overburdened numerous healthcare systems,which has also caused the loss of livelihoods due to prolonged shutdowns and further led to a cascading effect on the global economy.COVID-19 infections have an incubation period of 2–7 days,but 40 to 45%of cases are asymptomatic or show mild to moderate respiratory symptoms after the period due to subclinical lung abnormalities,making it more likely to spread the pandemic disease.To restrict the spread of the virus,on-site diagnosis methods that are quicker,more precise,and easily accessible are required.Rapid Antigen Detection Tests and Polymerase Chain Reaction tests are currently the primary methods used to determine the presence of COVID-19 viruses.These tests are typically time-consuming,not accurate,and,more importantly,not available to everyone.Hence,in this review and hypothesis,we proposed equipment that employs the properties of photonics to improve the detection of COVID-19 viruses by taking the advantage of typical binding of coronavirus with angiotensin-converting enzyme 2(ACE2)receptors.This hypothetical model would combine Surface-Enhanced Raman Scattering(SERS)and Fluorescence Resonance Energy Transfer(FRET)to provide great flexibility,high sensitivities,and enhanced accessibility.

Screening and identification of bioactive compounds from citrus against non-structural protein 3 protease of hepatitis C virus genotype 3a by fluorescence resonance energy transfer assay and mass spectrometry

BACKGROUND Hepatitis C virus genotype 3a(HCV G3a)is highly prevalent in Pakistan.Due to the elevated cost of available Food and Drug Administration-approved drugs against HCV,medicinal natural products of potent antiviral activity should be screened for the cost-effective treatment of the disease.Furthermore,from natural products,active compounds against vital HCV proteins like non-structural protein 3(NS3)protease could be identified to prevent viral proliferation in the host.AIM To develop cost-effective HCV genotype 3a NS3 protease inhibitors from citrus fruit extracts.METHODS Full-length NS3 without co-factor non-structural protein 4A(NS4A)and codon optimized NS3 protease in fusion with NS4A were expressed in Escherichia coli.The expressed protein was purified by metal ion affinity chromatography and gel filtration.Citrus fruit extracts were screened using fluorescence resonance energy transfer(FRET)assay against the protease and polyphenols were identified as potential inhibitors using electrospray ionization-mass spectrometry(MS)/MS technique.Among different polyphenols,highly potent compounds were screened using molecular modeling approaches and consequently the most active compound was further evaluated against HCV NS4A-NS3 protease domain using FRET assay.RESULTS NS4A fused with NS3 protease domain gene was overexpressed and the purified protein yield was high in comparison to the lower yield of the full-length NS3 protein.Furthermore,in enzyme kinetic studies,NS4A fused with NS3 protease proved to be functionally active compared to full-length NS3.So it was concluded that co-factor NS4A fusion is essential for the purification of functionally active protease.FRET assay was developed and validated by the half maximal inhibitory concentration(IC50)values of commercially available inhibitors.Screening of citrus fruit extracts against the native purified fused NS4A-NS3 protease domain showed that the grapefruit mesocarp extract exhibits the highest percentage inhibition 91%of protease activity.Among the compounds identified by LCMS analysis,hesperidin showed strong binding affinity with the protease catalytic triad having S-score value of-10.98.CONCLUSION Fused NS4A-NS3 protease is functionally more active,which is effectively inhibited by hesperidin from the grapefruit mesocarp extract with an IC50 value of 23.32μmol/L.

Investigation of fluorescence resonance energy transfer ultrafast dynamics in electrostatically repulsed and attracted exciton-plasmon systems

Following the gradual maturation of synthetic techniques for nanomaterials,exciton-plasmon composites have become a research hot-spot due to their controllable energy transfer through electromagnetic fields on the nanoscale.However,most reports ignore fluorescence resonance energy transfer(FRET)under electrostatic repulsion conditions.In this study,the FRET process is investigated in both electrostatic attraction and electrostatic repulsion systems.By changing the Au:quantum dot ratio,local-field induced FRET can be observed with a lifetime of ns and a fast component of hundreds of ps.These results indicate that the intrinsic transfer process can only elucidated by considering both steady and transient state information.

Retraction Note:Screening and identification of bioactive compounds from citrus against non-structural protein 3 protease of hepatitis C virus genotype 3a by fluorescence resonance energy transfer assay and mass spectrometry

Retraction note:Khan M,Rauf W,Habib F,Rahman M,Iqbal M.Screening and identification of bioactive compounds from citrus against non-structural protein 3 protease of hepatitis C virus genotype 3a by fluorescence resonance energy transfer assay and mass spectrometry.World J Hepatol 2020;12(11):976-992 PMID:33312423 DOI:10.4254/wjh.v12.i11.976.The online version of the original article can be found at https://www.wjgnet.com/1948-5182/full/v12/i11/976.htm.

A novel fluorescence sensor for milk clotting enzyme chymosin using peptide as substrate and covalent organic framework nanosheet as fluorescence quencher

Chymosin is one of the critical enzymes in cheese making.Herein,we proposed a novel fluorometric assay for chymosin determination.Firstly,covalent organic frameworks(COF)were synthesized and exfoliated to 2-dimensional COF nanosheets(COF NS)by ultrasound treatment.Gold nanoparticles(Au NPs)were loaded with COF NS to prepare AuNPs/COF NS(Au@COF NS).Secondly,rhodamine B(RhB)modified substrate peptide(Pep)for chymosin was linked with Au@COF NS to construct a Pep-Au@COF NS nanocomposite.For the sensing principle,fluorescence of RhB was quenched by Au@COF NS and the fluorescence intensity was weak due to the fluorescence resonance energy transfer between COF NS and RhB of Pep.However,in the presence of chymosin,the RhB was released by specific cleavage of the substrate peptide by chymosin and resulted in the recovery of fluorescence.The increased fluorescence intensity was proportional to the increase of chymosin concentration and thus a“turn on”fluorescent sensor for chymosin was constructed.The sensor showed a linear range in the concentration of 0.05-60.00μg/mL for the detection of chymosin with a detection limit of 20 ng/mL.The sensor was used to quantify chymosin in rennet product with good selectivity,which has the potential applications in cheese manufacturing.

Enhancing the Photosensitivity of Hypocrellin A by Perylene Diimide Metallacage‑Based Host–Guest Complexation for Photodynamic Therapy

The development of supramolecular hosts which can efficiently encapsulate photosensitizers to improve the photodynamic efficacy holds great promise for cancer therapy.Here,we report two perylene diimide-based metallacages that can form stable host–guest complexes with planar conjugated molecules including polycyclic aromatic hydrocarbons and photosensitizers(hypocrellin A).Such host–guest complexation not only prevents the aggregation of photosensitizers in aqueous environments,but also offers fluorescence resonance energy transfer(FRET)from the metallacage to the photosensitizers to further improve the singlet oxygen generation(Φ_(Δ)=0.66).The complexes are further assembled with amphiphilic polymers,forming nanoparticles with improved stability for anticancer study.Both in vitro and in vivo studies indicate that the nanoparticles display excellent anticancer activities upon light irradiation,showing great potential for cancer photodynamic therapy.This study provides a straightforward and effective approach for enhancing the photosensitivity of conventional photosensitizers via host–guest complexation-based FRET,which will open a new avenue for host–guest chemistry-based supramolecular theranostics.

Fluorescence resonance energy transfer-based nanomaterials for the sensing in biological systems

The applications of fluorescence resonance energy transfer(FRET)are coming to be one of the simplest and most accessible strategy with super-resolved optical measurements.Meanwhile,nanomaterials have become ideal for constructing FRET-based system,due to their unique advantages of tunable emission,broad absorption,and long fluorescence(FL)lifetime.The limitations of traditional FRET-based detections,such as the intrinsic FL,auto-FL,as well as the short FL lifetime,could be overcome with nanomaterials.Consequently,numbers of FRET-based nanomaterials have been constructed for precise,sensitive and selective detections in biological systems.They could act as both energy donors and/or acceptors in the optical energy transfer process for biological detections.Some other nanomaterials would not participate in the energy transfer process,but act as the excellent matrix for modifications.The review will be roughly classified into nanomaterial-involved and uninvolved ones.Different detection targets,such as nucleic acids,pathogenic microorganisms,proteins,heavy metal ions,and other applications will be reviewed.Finally,the other biological applications,including environmental evaluation and mechanism studies would also be summarized.

Exploring ratiometric endolysosomal pH nanosensors with hydrophobic indicators responding at the nanoscale interface and multiple fluorescence resonance energy transfers

Compared with conventional water-soluble fluorescence probes,pH-sensitive fluorescent nanosensors based on hydrophobic indicators remain largely unexplored.We report here the unique pH response of the nanosensors with a hydrophobic indicator(Ch3,a Nile Blue derivative)in polymeric nanoparticles(NPs).At the aqueous-organic interface of the NPs,spectral overlap and dye accumulation caused significant Förster resonance energy transfer(FRET)not only between the protonated and deprotonated Ch3(hetero-FRET),but also between the protonated and deprotonated Ch3 themselves(homo-FRET).The pH response was simulated according to an interfacial response mechanism and the dynamic range was found to depend on the size of the NPs and dye distribution(Kp).Therefore,adjusting the size of the NPs and the local dye concentration gave rise to a series of dynamic sensing ranges with apparent pKa values from 2.7 to 9.6 based on a single indicator.The nanosensors were successfully delivered to HeLa cells to monitor subcellular pH values in the endosomes and lysosomes.Based on cellular calibrations,the average pH in the organelles were determined to be ca.4.7.Moreover,the pH neutralization process during lysosome membrane permeabilization(LMP)induced by hydrogen peroxide stimulation was also successfully visualized with the nanosensors.

A Highly Selective and Sensitive Ratiometric Fluorescent Probe for pH Measurement Based on Fluorescence Resonance Energy Transfer

Fluorescein-rhodamine 6G（Flu-Rh） was synthesized and used as the fluorescence probe for pH measurement based on fluorescence resonance energy transfer（FRET）. In the probe, fluorescein fluorophore and pH-sensitive rhodamine 6G hydrazide were used as FRET donor and acceptor, respectively. The values of acidity constant（pKa） and fluorescence quantum yield（Ф） of Flu-Rh were 3.71 and 0.72, respectively. The fluorescence efficiency of Flu-Rh remains almost constant when the pH value of the sample solution changed 10 times in a range of 4.78-7.03 continuously. The present probe is simple and easy-to-use for the pH measurement in acidic media.

Construction of triblock copolymer-gold nanorod composites for fluorescence resonance energy transfer via pH-sensitive allosteric

To explore the effects of microenvironmental adjustments on fluorescence,a pH-sensitive nanocomposite system based on fluorescence resonance energy transfer(FRET)was constructed.The model system included a modified triblock copolymer(polyhistidine-b-polyethylene glycol-b-polycaprolactone)and gold nanoparticles.A near-infrared dye was used as the donor,and spectrally matched gold nanorods,attached after C-terminus modification with α-lipoic acid,were used as the receptor to realize control of the FRET effect over the fluorescence intensity for two polymer configurational changes(i.e.,"folded"and"stretched"states)in response to pH.After synthesis and characterization,we investigated the self-assembly behavior of the system.Analysis by quartz crystal microbalance revealed the pH sensitivity of the polymer,which exhibited"folding"and"stretching"states with changes in pH,providing a structural basis for the FRET effect.Fluorescence spectrophotometry investigations also revealed the regulatory impact of the assembled system on fluorescence.

Interaction of Surface-active Fluorescence Probes with Bovine Serum Albumin

The binding between three surface-active substituted 3H-indole fluorescence probes and bovine serum albumin (BSA) in aqueous solution was studied using fluorescence quenching. The binding constants of 3H-indole molecules with BSA were obtained. According to the F?rster resonance energy transfer theory, the distances between 3H-indole molecules and tryptophan of BSA were calculated. The results show that the oligoethyloxyethylene chain of 3H-indole molecules is longer, the binding between them is stronger, the energy transfer efficiency is higher, and the distance between tryptophan and 3H-indole is nearer.

Lineal DNA logic gate for microRNA diagnostics with strand displacement and fluorescence resonance energy transfer

Designing molecular logic gates to operate programmably for molecular diagnostics in molecular computing still remains challenging.Here,we designed a novel linear DNA logic gates for microRNA analysis based on strand displacement and fluorescence resonance energy transfer（FRET）.Two labeled strands closed each other produce to FRET through hybridization with a complementary strand to form a basic work unit of logic gate.Two indicators of heart failure（microRNA-195 and microRNA-21） were selected as the logic inputs and the fluorescence mode was used as the logic output.We have demonstrated that the molecular logic gate mechanism worked well with the construction of YES and AND gates.

Observation of amyloid precursor protein cleavage and Aβ generation in living cells by using multiphoton laser scanning microscopy

Objective To investigate the proteolytic mechanism of amyloid precursor protein （APP） and to explore amyloidbeta （Aβ） generation in living neurons. Methods DNA fragments were amplified by PCR or synthesized. The four fragments, CFP, 54bp, YFP and C99 were ligated into pcDNA3.0 vector to construct the recombinant plasmids pcDNA3.0-CFP-54bp- YFP and pcDNA3.0-CFP-54bp-YFP-C99. The SH-SY5Y cells were transiently transfected with pcDNA3.0-CFP-54bp-YFP or pcDNA3.0-CFP-54bp-YFP-C99. The expression of fusion gene was examined under a multiphoton laser scanning microscope. Fluorescence resonance energy transfer （FRET） was used to measure the β cleavage and γ cleavage of APE Aβ generation was confirmed by immunocytochemistry and multiphoton laser scanning microscopy. Cell viability was tested by MTT assay at different time points. Results （1） The double restriction endonuclease digestion and sequencing analysis confirmed the authenticity of the recombinant plasmids pcDNA3.0-CFP-54bp-YFP and pcDNA3.0-CFP-54bp- YFP-C99. （2） Blue and yellow fluorescences were detected in the transfected cells. （3） FRET occurred in pcDNA3.0-CFP- 54bp-YFP-transfected cells but not in pcDNA3.0-CFP-54bp-YFP-C99-transfected cells. （4） Aβ was produced in the pcDNA3.0- CFP-54bp-YFP-C99 transfected cells. （5） Aβ-deposition was widespread in the cell. （6） Cell viability decreased along with the intracellular Aβ deposition. Conclusion C99 is important for the APP β cleavage. Aβ may be generated and deposited in cells at the early stage of Alzheimer＇s disease. Intracellular Aβ accumulation brings deleterious effects on cells.

Preassembly and ligand-induced restructuring of the chains of the IFN-γ receptor complex： the roles of Jak kinases, Statl and the receptor chains

We previously demonstrated using noninvasive technologies that the interferon-gamma （IFN-γ） receptor complex is preassembled [ 1 ]. In this report we determined how the receptor complex is preassembled and how the ligand-mediated conformational changes occur. The interaction of Statl with IFN-γR1 results in a conformational change localized to IFN- γR1. Jakl but not Jak2 is required for the two chains of the IFN-γ receptor complex （IFN-γR1 and IFN-γR2） to interact; however, the presence of both Jakl and Jak2 is required to see any ligand-dependant conformational change. Two IFN- γR2 chains interact through species-specific determinants in their extracellular domains. Finally, these determinants also participate in the interaction of IFN-γR2 with IFN-γR1. These results agree with a detailed model of the IFN-γ receptor that requires the receptor chains to be pre-associated constitutively for the receptor to be active.

Spectroscopic analysis on the binding interaction of biologically active pyrimidine derivative with bovine serum albumin

A biologically active antibacterial reagent, 2-amino-6-hydroxy-4-（4-N, N-dimethylaminophenyl）-pyr- imidine-5-carbonitrile （AHDMAPPC）, was synthesized. It was employed to investigate the binding in- teraction with the bovine serum albumin （BSA） in detail using different spectroscopic methods. It ex- hibited antibacterial activity against Escherichia cali and Staphylococcus aureus which are common food poisoning bacteria. The experimental results showed that the fluorescence quenching of model carrier protein BSA by AHDMAPPC was due to static quenching. The site binding constants and number of binding sites （n ≈ 1） were determined at three different temperatures based on fluorescence quenching results. The thermodynamic parameters, enthalpy change （AH）, free energy （AG） and entropy change （AS） for the reaction were calculated to be 15.15 kJ/mol, -36.11 kJ/mol and 51.26J/mol K according to van＇t Hoff equation, respectively. The results indicated that the reaction was an endothermic and spontaneous process, and hydrophobic interactions played a major role in the binding between drug and BSA. The distance between donor and acceptor is 2.79 nm according to Forster＇s theory. The alterations of the BSA secondary structure in the presence of AHDMAPPC were confirmed by UV-visible, synchronous fluorescence, circular dichroism （CD） and three-dimensional fluorescence spectra. All these results in- dicated that AHDMAPPC can bind to BSA and be effectively transported and eliminated in the body. It can be a useful guideline for further drug design.

Adaption of Au Nanoparticles and CdTe Quantum Dots in DNA Detection

A DNA fluorescence probe system based on fluorescence resonance energy transfer （FRET） from CdTe quantum dot （QD） donors to Au nanoparticle （AuNP） acceptors is presented. CdTe QDs, 2.5nm in diameter, as energy donors, were prepared in water. Au nanoparticles, 16nm in diameter, as energy acceptors, were prepared from gold chloride by reduction. CdTe QDs were linked to 5＇-NH2-DNA through 1-ethyl-3-（dimethylaminopropyl）car- bodiimide hydrochloride （EDC） as a linker, and the 3＇-SH-DNA was self-assembled onto the surface of AuNPs. The hybridization of complementary double stranded DNA （dsDNA） bound to the QDs and AuNPs （CdTe-dsDNA-Au） determined the FRET distance of CdTe QDs and Au nanoparticles. Compared to the fluorescence of CdTe-DNA, the fluorescence of CdTe-DNA-Au conjugates decreased extremely, which indicated that the FRET occurred between CdTe QDs and Au nanoparticles. The fluorescence change of this conjugate depended on the ratio of Au-DNA to CdTe-DNA. When the AuNPs-DNA to QD-DNA ratio was 10：1, the FRET efficiency reached a maximum. The probe system would have a certain degree of fluorescence recovery when a complementary single stranded DNA was introduced into this system, which showed that the distance between CdTe QDs and Au nanoparticles was increased.

Von Willebrand Factor Antigen and ADAMTS13 Activity Assay in Pregnant Women and Severe Preeclamptic Patients

The present study examined von Willebrand factor (vWF) levels and ADAMTS13 activity in pregnant and severe preeclamptic women in order to shed light on the prothrombotic state in severe preeclampsia.Thirty healthy women of childbearing age,22 second trimester pregnant women,30 third trimester pregnant women and 10 severe preeclamptic patients were recruited in this study.ADAMTS13 activity was determined by the FRETS-vWF73 assay and vWF antigen (vWF:Ag) levels by an enzyme-linked immunosorbent assay.The results showed that there were statistically significant differences in plasma vWF antigen levels between the severe preeclamptic and third trimester pregnant women,between third and second trimester pregnant women (P【0.05).The third trimester pregnant women had significantly lower plasma ADAMTS13 activity than second trimester pregnant women (P【0.05).Nevertheless,no significant differences in plasma ADAMTS13 activity were found between severe preeclamptic patients and the third trimester pregnant women (P】0.05).In conclusion,plasma ADAMTS13 activity is normal in severe preeclampsia despite the increased vWF:Ag levels.Prothrombotic state is involved in the pathogenesis of severe preeclampsia,as a result of endothelial injury.

Implementation and application of FRET-FLIM technology

With the development of the new detection methods and the function of fluorescent molecule,researchers hope to further explore the internal mechanisms of organisms,monitor changes in the intracellular microenvironment,and dynamic processes of molecular interactions in cells.Fluo-rescence resonance energy transfer(FRET)describes the energy transfer process between donor fluorescent molecules and acceptor fluorescent molecules.It is an important means to detect protein-protein interactions and protein conformation changes in cells.Fluorescence lifetime imaging microscopy(FLIM)enables noninvasive measurement of the fAuorescence lifetime of fluorescent particles in vivo.The FRET-FLIM technology,which is use FLIM to quantify and analyze FRET,enables real-time monitoring of dynamic changes of proteins in biological cells and analysis of protein interaction mechanisms.The distance between donor and acceptor and their respective fAuorescent lifetime,which are of great importance for studying the mechanism of intracellular activity can be obtained by data analysis and algorithm ftting.

Dynamic photonic barcodes for molecular detection based on cavity-enhanced energy transfer

Optical barcodes have demonstrated a great potential in multiplexed bioassays and cell tracking for their distinctive spectral fingerprints.The vast majority of optical barcodes were designed to identify a specific target by fluorescence emission spectra,without being able to characterize dynamic changes in response to analytes through time.To overcome these limitations,the concept of the bioresponsive dynamic photonic barcode was proposed by exploiting interfacial energy transfer between a microdroplet cavity and binding molecules.Whispering-gallery modes resulting from cavity-enhanced energy transfer were therefore converted into photonic barcodes to identify binding activities,in which more than trillions of distinctive barcodes could be generated by a single droplet.Dynamic spectral barcoding was achieved by a significant improvement in terms of signal-to-noise ratio upon binding to target molecules.Theoretical studies and experiments were conducted to elucidate the effect of different cavity sizes and analyte concentrations.Timeresolved fluorescence lifetime was implemented to investigate the role of radiative and non-radiative energy transfer.Finally,microdroplet photonic barcodes were employed in biodetection to exhibit great potential in fulfilling biomedical applications.

Switchable 2,3-dithienylmaleimide bonded to different fluorophores: synthesis and photochromic properties

A series of photochromic diarylethene derivatives containing different fluorophores was synthesized by the Sonogashira coupling reaction and characterized by 1H nuclear magnetic resonance (NMR), 13C NMR and mass spectroscopy. Photochromic conversion of Cz-MS was observed and its structural change was confirmed by proton NMR. Switchable absorption and emission phenomena were observed for these molecules. These phenomena were attributed to the reversible photochromic fluorescence resonance energy transfer (pcFRET) process. The design of this novel photochromic system of Py-4MS enables information to be processed in a non-destructive manner. This overcomes the problem of the destructive nature of tracking photochromism using ultraviolet (UV)-vis spectroscopy.