Toward a nanopaper-based and solid phase immunoassay using FRET for the rapid detection of bacteria

Abstract

in this study, we propose a novel sensitive solid-based immunosensor developed on a plasmonic nanopaper platform for the detection of Escherichia coli (E. coli) bacteria. this plasmonic nanopaper that comprises of carboxylated bacterial cellulose (cBc) impregnated with gold nanoparticles (Aunp-cBc), employed as a quencher and a sustainable functionalized platform to be conjugated with protein A. thus, the conjugated protein A allows the aligned linkage of eAb-QD (anti-E. coli conjugated quantum dot) and eAb-Af (anti-E. coli conjugated Alexa Fluor 488). Interestingly, once E. coli was captured by the Aunp-cBc/eAb-QD or Aunp-cBc/eAb-Af, the energy transfer from the QD or Alexa Fluor fluorophores is triggered due to the conformational change in the antibody structure and this, in turn, causes a decrease in the distance between fluorophores and the quencher nanopaper and, therefore diminishing their photoluminescence. the immunosensors performed successfully to recognize E. coli at concentrations as low as 50 CFU mL −1 in the standard buffer. The examined functionality of the immunosensors in a real matrix such as chicken extract and lettuce juice demonstrated a highly efficient response while QD is the main fluorophore with a limit of detection around 100 CFU mL −1. During the last decades, the outbreak of foodborne diseases has seriously threatened people lives worldwide and led to a considerable number of hospitalizations and even deaths 1. Indeed, the incidence of foodborne infection is a result of ingesting food contaminated by foodborne pathogens such as bacteria, viruses, and parasites. To prevent its subsequence, precise monitoring and early recognition of pathogen would be the primary step to be ensured about the safety of our food 2. Upon comparing various categories of biosensors developed for pathogen detection, immunosensors have attracted plenty of attention due to their flexibility and compatibility in numerous transducer techniques such as electrochemical (voltammetry, impedancemetry, potentiometry), optical (fluorescence, SPR) and mass-based 3-5. The excellence of an immunosensor is directly dominated by the affinity binding of the antigen-antibody that this makes it uniquely sensitive and selective toward a specific target 6. Being fast, selective and sensitive, low-priced, in-situ and user-friendly, immunosensors can fully address the challenges associated with traditional bacteria detection approaches, such as; costly analysis, long processing time, and needs in scientific expertise 7. Given these benefits, its applications have been widely explored in the environment, medical and, food industries that designed either on a solid-based platform or solution-phase method 8,9. Assuredly, the integration of nanomaterials with immunosensing techniques has caused a huge improvement in its functionality and sensitivity 10-12. Nanoscale materials, with a size less than 100 nm are endowed with unique physical and chemical properties such as; high surface area, optical, and electrical characteristics, which ...

Sandwich-Based Immunosensor for Dual-Mode Detection of Pathogenic F17–Positive Escherichia coli Strains

“…Although no electrochemical biosensor was reported for detection of F17–positive E. coli bacteria, our biosensor showed satisfactory analytical performances compared to other biosensors of pathogenic E. coli ( Table 1 ). For instance, the developed immunosensor showed better LOD than that of E. coli O157:H7 impedimetric biosensor (LOD = 48 CFU·mL −1 ), based on the use of self-assembled gold nanoparticles and protein G [ 39 ], or E. coli O157:H7 biosensor combining aptamer−induced catalytic hairpin assembly circle amplification with microchip electrophoresis (LOD = 75 CFU·mL −1 ) [ 40 ]. Moreover, the elaboration of the sandwich biosensor and the detection of F17–positive E. coli had lower assay time.…”

In Situ Grown Gold Nanoparticles Enable Plasmonic Staining of Bacteria for High-Performance On-Site Microbial Analysis in Drinking Water

“…Those strategies exhibited high sensitivity, but the long-term storage of Au nanoprobes weakened the reproducibility of analytical methods. Moreover, the coexisting substances, such as salt, small molecules, and proteins in real samples, might either cause the nonspecific aggregation or influence the surface groups of Au nanoprobes, and consequently, false results were identified. − Second, the presynthetic Au nanoparticles were fixed on paper substrates, rather than dispersed in homogeneous solutions, and the bacterium measurements were conducted using the lateral flow assay (LFA). − The strategies exhibited merits of comparable speediness, simplicity, and anti-interference, but in those protocols, Au nanomaterials were used as the visual probes, and low sensitivity was exposed due to the lack of signal amplification routes. If a novel method is integrated with those attractive merits of solution-based tests and LFA, the on-site bacterium analysis approach could be remarkably developed.…”

Development of gold nanoparticles-aptamer nanocomposite for multiplexed analysis of antibiotics and design of molecular logic gates

“…FRET, a non-radiative energy transfer process, depends on the distance between energy donors and acceptors [18]. Actually, the FRET mechanism has been extensively-used in the screening and detection of various analytes [19][20][21][22][23][24][25]. For example, Deka et al [19] developed a FRET-based method to determine the helicase activity using the optical properties of DNA-conjugated AuNPs.…”