Toward Personalized Nanomedicine: The Critical Evaluation of Micro and Nanodevices for Cancer Biomarker Analysis in Liquid Biopsy.

Liquid biopsy for the analysis of circulating cancer biomarkers (CBs) is a major advancement toward the early detection of cancer. In comparison to tissue biopsy techniques, liquid biopsy is relatively painless, offering multiple sampling opportunities across easily accessible bodily fluids such as blood, urine, and saliva. Liquid biopsy is also relatively inexpensive and simple, avoiding the requirement for specialized laboratory equipment or trained medical staff. Major advances in the field of liquid biopsy are attributed largely to developments in nanotechnology and microfabrication that enables the creation of highly precise chip-based platforms. These devices can overcome detection limitations of an individual biomarker by detecting multiple markers simultaneously on the same chip, or by featuring integrated and combined target separation techniques. In this review, the major advances in the field of portable and semi-portable micro, nano, and multiplexed platforms for CB detection for the early diagnosis of cancer are highlighted. A comparative discussion is also provided, noting merits and drawbacks of the platforms, especially in terms of portability. Finally, key challenges toward device portability and possible solutions, as well as discussing the future direction of the field are highlighted.

Exosomal microRNAs array sensor with a bioconjugate composed of p53 protein and hydrazine for the specific lung cancer detection.

For the early diagnosis of lung cancer, a novel strategy to detect microRNAs encapsulated in exosomes with immunomagnetic isolation was demonstrated for the selective extraction of exo-miRNAs from patient serum. Here, miRNA was captured from lysed exosomes in specially designed capture probe modified magnetic beads, followed by T4 DNA polymerase-mediated in situ formation of chimeric 5'-miRNA-DNA-3' (Target). The poly-(2,2':5',2''-terthiophene-3'-(p-benzoic acid)) (pTBA)-modified electrode harbors Probe-1 DNA that hybridizes to the 5' end of the chimera, followed by hybridization of Probe-2 DNA to the 3' end of the chimera, resulting in the formation of a 20-nucleotide-long dsDNA consensus sequence for p53 protein binding. A bioconjugate composed of p53 and hydrazine assembled on AuNPs (p53-AuNPs-Hyd) recruits the p53 protein to recognize a specific sequence, forming the final sensor probe (pTBA-Probe-1:Target/Probe-2:bioconjugate), where hydrazine functions as an electrocatalyst to generate amperometric signal from the reduction of H2O2. This sensor has double specificity via selective capture of the target in Probe-1 and p53 recognition, which shows excellent analytical performance, revealing a dynamic range between 100 aM and 10 pM with a detection limit of 92 (±0.1) aM. For practical applications, we prepared a multiplexed array sensor to simultaneously detect four exo-miRNAs (miRNA-21, miRNA-155, miRNA-205, and miRNA-let-7b) up to femtomolar levels from 1.0 mL to 125 µL of cell culture (A549, MCF-7 and BEAS-2B) media and lung cancer patient serum samples, respectively.

A novel DNA binding protein-based platform for electrochemical detection of miRNA.

We present a novel amplification-free sandwich type platform assay for electrochemical detection of miRNA. The assay is based on T4 DNA polymerase mediated synthesis of the p53 binding DNA sequence at the 3' end of target miRNA. The resulting miRNA-DNA chimera is detected via an electrochemical sandwich hybridization assay where HRP-labelled p53 binds to its recognition sequence and an amperometric signal is generated by hydroquinone-mediated enzymatic reduction of H2O2. The limit of detection of our assay was estimated to be 22 fM with a linear dynamic range of 100 fM-1 nM. This new platform method of detecting miRNA shows superior performance to conventional electrochemical miRNA biosensors and has the potential for amplification-free analysis of miRNA with high specificity and sensitivity.

Nanozyme-based electrochemical biosensors for disease biomarker detection.

In recent years, a new group of nanomaterials named nanozymes that exhibit enzyme-mimicking catalytic activity has emerged as a promising alternative to natural enzymes. Nanozymes can address some of the intrinsic limitations of natural enzymes such as high cost, low stability, difficulty in storage, and specific working conditions (i.e., narrow substrate, temperature and pH ranges). Thus, synthesis and applications of hybrid and stimuli-responsive advanced nanozymes could revolutionize the current practice in life sciences and biosensor applications. On the other hand, electrochemical biosensors have long been used as an efficient way for quantitative detection of analytes (biomarkers) of interest. As such, the use of nanozymes in electrochemical biosensors is particularly important to achieve low cost and stable biosensors for prognostics, diagnostics, and therapeutic monitoring of diseases. Herein, we summarize the recent advances in the synthesis and classification of common nanozymes and their application in electrochemical biosensor development. After briefly overviewing the applications of nanozymes in non-electrochemical-based biomolecular sensing systems, we thoroughly discuss the state-of-the-art advances in nanozyme-based electrochemical biosensors, including genosensors, immunosensors, cytosensors and aptasensors. The applications of nanozymes in microfluidic-based assays are also discussed separately. We also highlight the challenges of nanozyme-based electrochemical biosensors and provide some possible strategies to address these limitations. Finally, future perspectives on the development of nanozyme-based electrochemical biosensors for disease biomarker detection are presented. We envisage that standardization of nanozymes and their fabrication process may bring a paradigm shift in biomolecular sensing by fabricating highly specific, multi-enzyme mimicking nanozymes for highly sensitive, selective, and low-biofouling electrochemical biosensors.

MicroRNAs in ovarian cancer and recent advances in the development of microRNA-based biosensors.

Ovarian cancer is the most aggressive of all gynaecological malignancies and is the leading cause of cancer-associated mortality worldwide. Over the recent years, there has been a sharp increase in this mortality rate, mostly due to late diagnosis, which can be attributed to the lack of an early and specific biomarker. Under this scenario, recent interest has shifted towards ovarian cancer associated miRNAs which play strong regulatory roles in various cellular processes. miRNAs have emerged as promising non/minimally invasive cancer biomarkers for improved diagnostic, prognostic and streamlined therapeutic applications. A large number of miRNA assays have been reported that are based on nucleic acid detection-based techniques such as RT-qPCR, microarrays and RNA sequencing methods. Despite demonstrating commendable analytical performances, these laboratory-based techniques are expensive and hence not ideally suited for routine use in resource-limited settings. In recent years, considerable attention has been dedicated to the development of relatively simple, rapid and inexpensive miRNA biosensor strategies. Among these, electrochemical sensors have shown a great promise towards point-of-care diagnostics, due to their inherent advantages such as simplicity, sensitivity, amenability to high levels of multiplexing as well as low cost. In this paper, we provide an overview of the potential role of miRNAs in ovarian cancer, as well as recent advances in the development of nanotechnology-based, optical, and electrochemical biosensing-strategies for miRNA detection.