Amplified visual immunosensor integrated with nanozyme for ultrasensitive detection of avian influenza virus 

Syed Rahin Ahmed1, Juan C. Corredor2, Éva Nagy2, Suresh Neethirajan1* 

Nanotheranostics Journal 

Nanomaterial-based artificial enzymes or nanozymes exhibit superior properties such as stability, cost effectiveness and ease of preparation in comparison to conventional enzymes. However, the lower catalytic activity of nanozymes limits their sensitivity and thereby practical applications in the bioanalytical field. To overcome this drawback, herein we propose a very simple but highly sensitive, specific and low-cost dual enhanced colorimetric immunoassay for avian influenza A (H5N1) virus. 3,3´,5,5´- Tetramethylbenzidine (TMBZ) was used as a reducing agent to produce gold nanoparticles (Au NPs) with blue colored solution from a viral target-specific antibody-gold ion mixture at first step. The developed blue color from the sensing design was further amplified through catalytic activity of Au NPs in presence of TMBZ–hydrogen peroxide (H2O2) solution in second step. Hence, the developed dual enhanced colorimetric immunosensor enables the detection of avian influenza virus A (H5N1) with a limit of detection (LOD) of 1.11 pg/mL. Our results confirmed that the developed assay has superior sensitivity than the conventional ELISA method, plasmonic-based bioassay and commercial flu diagnostic kits. Proposed sensing method further showed its capability to detect real viruses, avian influenza A (H4N6) and A (H9N2) virus, in blood samples with limit of detection of 0.0269 HAU and 0.0331 HAU respectively. 


In situ self-assembly of gold nanoparticles on hydrophilic and hydrophobic substrates for influenza virus-sensing platform 

Syed Rahin Ahmed, Jeonghyo Kim, Van Tan Tran, , Tetsuro Suzuki, , Suresh Neethirajan, Jaebeom Lee, & Enoch Y. Park

Scientific Reports


Nanomaterials without chemical linkers or physical interactions that reside on a two-dimensional surface are attractive because of their electronic, optical and catalytic properties. An in situ method has been developed to fabricate gold nanoparticle (Au NP) films on different substrates, regardless of whether they are hydrophilic or hydrophobic surfaces, including glass, 96-well polystyrene plates, and polydimethylsiloxane (PDMS). A mixture of sodium formate (HCOONa) and chloroauric acid (HAuCl4) solution was used to prepare Au NP films at room temperature. An experimental study of the mechanism revealed that film formation is dependent on surface wettability and inter particle attraction. The as-fabricated Au NP films were further applied to the colorimetric detection of influenza virus. The response to the commercial target, New Caledonia/H1N1/1999 influenza virus, was linear in the range from 10 pg/ml to 10 μg/ml and limit of detection was 50.5 pg/ml. In the presence of clinically isolated influenza A virus (H3N2), the optical density of developed color was dependent on the virus concentration (10–50,000 PFU/ml). The limit of detection of this study was 24.3 PFU/ml, a limit 116 times lower than that of conventional ELISA (2824.3 PFU/ml). The sensitivity was also 500 times greater than that of commercial immunochromatography kits.








Electrochemical latent redox ratiometric probes for real-time tracking and quantification of endogenous hydrogen sulfide production in living cells

Biosensors and Bioelectronics -

Hydrogen sulfide (H2S) was discovered as a third gasotransmitter in biological systems and recent years have seen a growing interest to understand its physiological and pathological functions. However, one major limiting factor is the lack of robust sensors to quantitatively track its production in real-time. We described a facile electrochemical assay based on latent redox probe approach for highly specific and sensitive quantification in living cells. Two chemical probes, Azido Benzyl ferrocene carbamate (ABFC) and N-alkyl Azido Benzyl ferrocene carbamate (NABFC) composed of azide trigger group were designed. H2S molecules specifically triggered the release of reporters from probes and the current response was monitored using graphene oxide film modified electrode as transducer. The detection limits are 0.32 µM (ABFC) and 0.076 µM (NABFC) which are comparable to those of current sensitive methods. The probes are successful in the determination of H2S spiked in whole human blood, fetal bovine serum, and E. coli. The continuous monitoring and quantification of endogenous H2S production in E. coli were successfully accomplished. This work lays first step stone towards real-time electrochemical quantification of endogenous H2S in living cells, thus hold great promise in the analytical aspects of H2S.


GryphSens: A Smartphone-Based Portable Diagnostic Reader for the Rapid Detection of Progesterone in Milk

Hyunwook Jang, Syed Rahin Ahmed and Suresh Neethirajan

Enzyme-linked immunosorbent assay (ELISA) is a popular assay technique for the detection and quantification of various biological substances due its high sensitivity and specificity. More often, it requires large and expensive laboratory instruments, which makes it difficult to conduct when the tests must be performed quickly at the point-of-care (POC). To increase portability and ease of use, we propose a portable diagnostic system based on a Raspberry Pi imaging sensor for the rapid detection of progesterone in milk samples. We designed, assembled, and tested a standalone portable diagnostic reader and validated it for progesterone detection against a standard ELISA assay using a commercial plate reader. The portable POC device yielded consistent results, regardless of differences in the cameras and flashlights between various smartphone devices. An Android application was built to provide front-end access to users, control the diagnostic reader, and display and store the progesterone measurement on the smartphone. The diagnostic reader takes images of the samples, reads the pixel values, processes the results, and presents the results on the handheld device. The proposed POC reader can perform to superior levels of performance as a plate reader, while adding the desirable qualities of portability and ease of use.  


The anti-listerial properties of biodegradable polylactide coatings modified with titanium dioxide have been studied. Free standing films were prepared by casting solutions prepared from titanium dioxide and previously extruded polylactide. It was demonstrated that polylactide alone could support 2.84 ± 0.10 log CFU reduction of Listeria monocytogenes when incubated at 23 °C for 2 h. However, the log reduction for Listeriacould be increased to >4 log CFU with titanium dioxide:polylactide composites illuminated with UV-A. The inactivation kinetics of L. monocytogenes followed a diphasic die-off with an initial 30 min lag period then a progressive decline in bacterial levels over a further 90 min period. The anti-listeria effect of polylactide:titanium dioxide films was dependent on illumination with UV-A but independent on the concentration of TiO2incorporated in the film within the range of 1–5% w/w. The mode of L. monocytogenesinactivation was via direct contact of the pathogen with the polylactide, in addition to the generation of oxygen radicals produced by excitation of the titanium dioxide. The composite film illuminated with UV-A was equally effective against SalmonellaTyphimurium and Shiga toxin producing Escherichia coli. The coating was stable to 5 repeated sanitation cycles consisting of detergent and sodium hypochlorite rinses. The polylactide-titanium dioxide coating shows potential as an antimicrobial coating although further work is required to assess if the protective film can function under commercial conditions. 


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