Microfluidic Surface Plasmon Resonance Sensors: From Principles to Point-of-Care Applications.

Surface plasmon resonance (SPR) is a label-free, highly-sensitive, and real-time sensing technique. Conventional SPR sensors, which involve a planar thin gold film, have been widely exploited in biosensing; various miniaturized formats have been devised for portability purposes. Another type of SPR sensor which utilizes localized SPR (LSPR), is based on metal nanostructures with surface plasmon modes at the structural interface. The resonance condition is sensitive to the refractive index change of the local medium. The principles of these two types of SPR sensors are reviewed and their integration with microfluidic platforms is described. Further applications of microfluidic SPR sensors to point-of-care (POC) diagnostics are discussed.

Opto-Microfluidic Immunosensors: From Colorimetric to Plasmonic.

Optical detection has long been the most popular technique in immunosensing. Recent developments in the synthesis of luminescent probes and the fabrication of novel nanostructures enable more sensitive and efficient optical detection, which can be miniaturized and integrated with microfluidics to realize compact lab-on-a-chip immunosensors. These immunosensors are portable, economical and automated, but their sensitivity is not compromised. This review focuses on the incorporation and implementation of optical detection and microfluidics in immunosensors; it introduces the working principles of each optical detection technique and how it can be exploited in immunosensing. The recent progress in various opto-microfluidic immunosensor designs is described. Instead of being comprehensive to include all opto-microfluidic platforms, the report centers on the designs that are promising for point-of-care immunosensing diagnostics, in which ease of use, stability and cost-effective fabrication are emphasized.

Tip-enhanced fluorescence with radially polarized illumination for monitoring loop-mediated isothermal amplification on hepatitis C virus cDNA.

A tip nanobiosensor for monitoring DNA replication was presented. The effects of excitation power and polarization on tip-enhanced fluorescence (TEF) were assessed with the tip immersed in fluorescein isothiocyanate solution first. The photon count rose on average fivefold with radially polarized illumination at 50 mW. We then used polymerase-functionalized tips for monitoring loop-mediated isothermal amplification on Hepatitis C virus cDNA. The amplicon-SYBR® Green I complex was detected and compared to real-time loop-mediated isothermal amplification. The signals of the reaction using 4 and 0.004 ng∕µl templates were detected 10 and 30 min earlier, respectively. The results showed the potential of TEF in developing a nanobiosensor for real-time DNA amplification.

Gold nanorods as probes in two-photon fluorescence correlation spectroscopy: emerging applications and potential artifacts.

Owing to the highly efficient two-photon fluorescence of gold nanorods and very short fluorescence lifetime compared with the rotational correlation time, the rotation and diffusion of a single gold nanorod can be easily observed by two-photon fluorescence correlation spectroscopy (TP-FCS). This property, along with the previous successful use as a contrast agent in two-photon fluorescence imaging, suggests a potential application in TP-FCS as well. Although the FCS measurement becomes highly efficient with gold nanorods as probes, the amplitude and temporal decay of the measured correlation functions depend critically on excitation power. Here, we investigate various photophysical processes of gold nanorods to determine the cause of such a sensitive power dependency. This understanding provides a basis for choosing appropriate FCS models to recover reasonable physical parameters. Although the correlation function amplitude G(0) is 32 times lower when the excitation power increases from 20 µW to 1.12 mW, the application of a saturation-modified FCS model yields very good fit to each data set and the fitted concentration of 0.64 nM is comparable to the 0.7 nM given by the inductively coupled plasma mass spectrometry measurement. The FCS assay appears to be an efficient method for the quantification of gold nanorods when correctly interpreted. However, even with the saturation considered in the fitting model, the fitted rotational and translational diffusion rates are getting faster as the power increases. This indicates that other effects such as photothermal effects may raise the local temperature, and thus increasing the rotational and translational diffusion rate.

Detection of tip-enhanced fluorescence from loop-mediated isothermal amplification of hepatitis B virus by two-photon microscopy.

Tip-enhanced fluorescence of localized DNA replication by loop-mediated isothermal amplification (LAMP) is a potential way to observe real-time biological reaction confined in nanometer scale. We successfully coated Bst polymerase on the apex (~100 nm) of an atomic force microscope (AFM) tip and performed localized LAMP reaction of hepatitis B virus (HBV). By using this tip-based reaction, the replicated HBV DNA can be directly imaged to be 400~500 nm spots by using two-photon excitation fluorescence microscopy.

Surface plasmon effects on two photon luminescence of gold nanorods.

Gold nanorods emit strong photoluminescence under two photon excitation; the efficient two photon lumininescence (TPL) arises from the local field enhancement assisted by surface plasmons. The surface plasmon effects on the TPL efficiency and spectrum are investigated by measuring the TPL of gold nanorods with various aspect ratios. A large TPL efficiency is found when incident light wavelength coincides with the longitudinal surface plasmon mode of the gold nanorods. However, the emission spectra of nanorods with various aspect ratios look similar and exhibit modest surface plasmon features, which implies a major non-radiative decay of excited surface plasmons.

Density-dependent optical response of gold nanoparticle monolayers on silicon substrates.

We experimentally and theoretically study the density-dependent optical properties of 20 nm gold nanoparticle monolayers on silicon at 530 nm. Both results indicate a density-dependent resonance behavior. The density-dependent resonance suggests that the optical properties of the gold nanoparticle monolayers can be drastically modulated by changing the volume ratio V(r). It also implies, in this composite system, that the total surface plasmon resonance is maximized at a certain V(r).

Peripheral binding mode and penetration depth of cobra cardiotoxin on phospholipid membranes as studied by a combined FTIR and computer simulation approach.

Cobra cardiotoxin, a cytotoxic beta-sheet basic polypeptide, is known to cause membrane leakage in many cells including human erythrocytes. Herein, we demonstrate that the major cobra cardiotoxin from Naja atra, CTX A3, can cause leakage of vesicle contents in phosphatidylglycerol (PG) and phosphatidylserine containing, but not in pure phosphatidylcholine (PC), membrane bilayers. By the combined polarized attenuated total reflection infrared spectroscopy and computer simulation studies, CTX A3 is shown to peripherally bind to both zwitterionic and anionic monolayers in a similar edgewise manner with a tilted angle of approximately 48 +/- 20 degrees between the beta-sheet plane of the CTX molecule and the normal of the membrane surface. The average surface area expansion induced by CTX A3 binding to the PG monolayer, however, is two times larger than that of the PC monolayer as determined by the Langmuir minitrough method. Interaction energy considerations of CTX A3 on neutral and negatively charged membrane surfaces suggests that the electrostatic interaction between anionic lipid and cationic CTXs plays a role in modulating the penetration depth of CTX molecules on the initial peripheral binding mode and reveals a pathway leading to the formation of an inserted mode in negatively charged membrane bilayers.