Detecting cancer from urine sample – Will the development of a nanowire based device open the gates for early diagnoses and timely medical checkups for cancer?

MicroRNAs (miRNAs) encapsulated by extracellular vesicles (EVs) are found in body fluids of patients with malignant diseases as well as with those having a better health. The difference in the EV-encapsulated miRNAs between these two groups of people could be used as a signature to identify various diseases. The miRNAs present in urine could serve as biomarkers for detecting cancer. Unfortunately, the concentration of EVs in urine is extremely low (<0.01 volume %) and hence the most commonly used method of extraction – ultracentrifugation is not capable of extracting nearly 90% of the miRNA species, due to their low abundance. Recently, a nanowire-based device anchored to a microfluidic substrate is fabricated for the efficient collection of EVs and in situ extraction of various miRNAs of different sequences, which is believed to open the gates for urine-based early diagnoses and medical checkups for cancer (Yasui et al., Sci. Adv. 2017;3: e1701133).

Si (100) served as the base substrate (Fig. 1(a)), which was coated with a positive photoresist followed by channel patterning using photo lithography (Fig. 1(b)). A 140 nm thick Cr layer was sputter deposited (Fig. 1(c)), followed by removal of the photoresist layer and thermal oxidation of the Cr layer at 400 °C for 2 h (Fig. 1(d)). The thermally oxidized Cr layer served as the seed layer for subsequent growth of ZnO nanowires using a solution mixture of 15 mM hexamethylenetetramine (HMTA) and 15 mM zinc nitrate hexahydrate at 95 °C for 3 h (Fig. 1(e)). PDMS was poured over the ZnO nanowire grown substrate and cured (Fig. 1(f)). Subsequently, the PDMS was removed from the Si substrate (Fig. 1(g)) and the ZnO nanowires in the PDMS were transferred to another PDMS substrate. The transferred nanowires were uniformly and deeply buried into PDMS while their slightly emerged heads served as growth points for the second nanowire growth (Fig. 1(h)), which was carried out by immersing the PDMS in a solution mixture of 15 mM HMTA and 15 mM zinc nitrate hexahydrate at 95 °C for 3 h (Fig. 1(i)). To enhance contact events between the ZnO nanowires and the EVs as well as to avoid any pressure drop, the ZnO nanowire embedded PDMS substrate was anchored to a herringbone-structured PDMS substrate (Fig. 1(j)).

Fig. 1 (a-j) Various stages involved in the fabrication of nanowire-based device; and (k) schematic of the collection and extraction of EV–encapsulated miRNAs.

The nanowire-based device is capable of detecting around 1000 types of species of miRNAs when compared to the conventional ultracentrifugation method. Using this device, it is possible to extract EV–encapsulated miRNAs within 40 min (collection, 20 min; extraction, 20 min) by introducing just 1 ml of urine sample followed by 1 ml of lysis buffer into the device (Fig. 1(k)). In contrast, the ultracentrifugation method requires 20 ml of urine sample and more than 5 h for collection and extraction. The device enables a four-fold increase in the miRNA expression level with a larger variety of extracted species of miRNAs. The ZnO nanowire-based device is found to be superior to the commonly used ultracentrifugation method in terms of treatment time and RNA extraction efficiency. This attribute is due to its large surface area of ZnO nanowires and their positively charged surface (isoelectric point of 9.50 at pH 6 to 8), which electrostatically attracts the negatively charged EVs in urine sample at pH 6-8. In addition, the mechanical stability of ZnO nanowires, which are firmly anchored to the PDMS substrate helps to retain their strength during buffer flow and enhances the extraction efficiency. The positively charged surface of ZnO nanowires offers benefit in collecting negatively charged objects in urine samples, including exosomes, microvesicles, and EV-free miRNAs.

The ZnO nanowire-based device is believed to help in the early diagnoses and timely medical checkups based on urine miRNA analysis. The method is capable of identifying urinary miRNAs that could potentially serve as biomarkers for detecting bladder, prostate lung, pancreas, and liver cancer.

T.S.N. Sankara Narayanan

Flexible sensors for real-time monitoring of temperature, pressure and flow

Development of strain/stress sensors for human-motion detection and implantable devices for human health monitoring assumed significance in recent years. Multi-functionality, poor interface stability and flexibility of such sensors still remain as a major challenge. Researchers at State Key Laboratory for Modification of Chemical Fibers and Polymer Materials, and Engineering Research Center of Advanced Glasses Manufacturing Technology, College of Materials Science and Engineering, Donghua University, China have reported fabrication of a mechanically strong and flexible vessel-like sensor that is capable of continuously monitoring applied temperature, strain, and the frequency, pressure, and temperature of the in-vessel flow.

Braided cotton yarn served as the base and it was coated with single-walled carbon nanotubes (SWCNTs) by dip-coating (cotton@SWNTs) to function as a current collector/electrode. ZnO nanoseeds were nucleated on the surface of cotton@SWNTs to promote the growth of ZnO nanorods, which could serve as temperature sensing arrays. A polyvinylidene fluoride (PVDF) film was formed over the cotton@SWCNTs/ZnO through dip-coating. A PVDF fibrous membrane was further prepared on the PVDF surface through electrospinning. The PVDF fibrous membrane was immersed in ethyl acetate for 12 h to obtain a compact film. The PVDF membrane could serve as piezoelectric nanofibers. Finally, silver paste was uniformly coated on the surface of the PVDF membrane. The various stages involved in the fabrication are shown in Fig. 1.

Fig. 1 various stages involved in the fabrication of flexible sensor

The morphological features of cotton@SWNTs indicate a uniform distribution of SWCNTs on the surface of cotton yarn (Fig. 2(a)). Similarly, the ZnO nanorods arrays@PVDF are also uniformly developed on the surface of cotton@SWCNTs (Figs. 2(b) and 2(c)).

Fig. 2 Morphological features of (a) SWCNTs coated cotton yarn; (b, c) cotton@SWCNTs/ZnO@PVDF; (c) magnified image of ZnO nanorods

Structural representation of the tubular sensor used for monitoring temperature, motions and liquid flow is depicted in Fig 3(a). The thermosensitivity of ZnO grown between the fibers of SWCNTs helps monitoring the temperature variation, which is detected through resistance measurement of the tubular inner electrode (Fig. 3(a)). The I-V curves measured using the functionalized inner electrode of the sensor at different temperatures indicate the change in resistance with variation in temperature (Fig. 3(b)).

Fig. 3 (a) Schematic representation of the sensor used for monitoring temperature, motions and liquid flow; (b) Linear I-V curves of the sensor measured under different fluid temperature.

The sensor is also capable of monitoring human body motions by sensing both bending and pressing. When a force is applied on the surface of the PVDF nanofibers, they get polarized. The equivalent bound charges accumulated on the surface of PVDF nanofibers are measured as electrical signals with the help of bonding wires between the two electrodes – cotton@SWCNTs/ZnO layer and the silver paste layer. The output voltage is increased from 0.038V to 0.1V with an increase in bending angle from 1.4 rad to 2.1 rad (Figs. 4(a) and 4(c)). Similarly, an increase in output voltage is observed with an increase in frequency during compression (Figs. 4(b) and 4(d)). These inferences suggest that human motions such as bending and compression can be directly monitored without any external power supply.

Fig. 4 (a, b) Demonstration of the bending and pressing motion; (c) output voltage generated by the sensor under different bending angles; and (d) output voltage generated by the sensor under compression at different frequencies.

The ability of the sensor to monitor blood temperature and blood pressure is also evaluated. Fig. 5(a) shows the experimental set-up used to monitor liquid flow. The temperature of the test fluid (deionized water to mimic blood flow) that flows into the sensor is accurately controlled using microfluidic channels heated by sample heater. The variation in the temperature of the fluid under flow from 35 to 40 °C is clearly reflected by the change in resistance measured using the sensor (Fig. 5(b)). The sensor could also detect the variation in the input pressure and frequency of the pulsed flow. The increase in the output voltage with an increase in applied pressure indicates the ability of the sensor to measure the variation in pressure under flow conditions (Fig. 5(c)). Similarly, the variation in the output current is sufficient enough to qualify the sensor for its ability to measure the pulse frequency of fluid (Fig. 5(d)).

Fig. 5 (a) Experimental set-up used to monitor liquid flow using the sensor; (b) I-V curves of the sensor measured under a narrow fluid temperature range; (c) output voltage generated by the sensor under different fluid input pressure; and (d) output current generated by the sensor under different pulse frequencies.

The cotton@SWCNTs/ZnO@PVDF sensor is flexible and durable. The output signal of the sensor remains steady during 1920 circles of the bending circle test. The sensor facilities detection and continuous monitoring of the applied temperature, strain, and the frequency, pressure, and temperature of pulsed fluids. It can be used for implantable physical sensing applications and monitoring of human health.

T.S.N. Sankara Narayanan

For more information, the reader may kindly refer: Wei Zhang et al., A strong and flexible electronic vessel for real-time monitoring temperature, motions and flow, Nanoscale, 2017, DOI: 10.1039/C7NR05575G