Microfluidic Platforms for the Isolation and Detection of Exosomes: A Brief Review.

Extracellular vesicles (EVs) are a group of communication organelles enclosed by a phospholipid bilayer, secreted by all types of cells. The size of these vesicles ranges from 30 to 1000 nm, and they contain a myriad of compounds such as RNA, DNA, proteins, and lipids from their origin cells, offering a good source of biomarkers. Exosomes (30 to 100 nm) are a subset of EVs, and their importance in future medicine is beyond any doubt. However, the lack of efficient isolation and detection techniques hinders their practical applications as biomarkers. Versatile and cutting-edge platforms are required to detect and isolate exosomes selectively for further clinical analysis. This review paper focuses on lab-on-chip devices for capturing, detecting, and isolating extracellular vesicles. The first part of the paper discusses the main characteristics of different cell-derived vesicles, EV functions, and their clinical applications. In the second part, various microfluidic platforms suitable for the isolation and detection of exosomes are described, and their performance in terms of yield, sensitivity, and time of analysis is discussed.

Gold Nano-Island Platforms for Localized Surface Plasmon Resonance Sensing: A Short Review.

Nano-islands are entities (droplets or other shapes) that are formed by spontaneous dewetting (agglomeration, in the early literature) of thin and very thin metallic (especially gold) films on a substrate, done by post-deposition heating or by using other sources of energy. In addition to thermally generated nano-islands, more recently, nanoparticle films have also been dewetted, in order to form nano-islands. The localized surface plasmon resonance (LSPR) band of gold nano-islands was found to be sensitive to changes in the surrounding environment, making it a suitable platform for sensing and biosensing applications. In this review, we revisit the development of the concept of nano-island(s), the thermodynamics of dewetting of thin metal films, and the effect of the substrate on the morphology and optical properties of nano-islands. A special emphasis is made on nanoparticle films and their applications to biosensing, with ample examples from the authors' work.

Lab-On-A-Chip for the Development of Pro-/Anti-Angiogenic Nanomedicines to Treat Brain Diseases.

There is a huge demand for pro-/anti-angiogenic nanomedicines to treat conditions such as ischemic strokes, brain tumors, and neurodegenerative diseases such as Alzheimer's and Parkinson's. Nanomedicines are therapeutic particles in the size range of 10-1000 nm, where the drug is encapsulated into nano-capsules or adsorbed onto nano-scaffolds. They have good blood-brain barrier permeability, stability and shelf life, and able to rapidly target different sites in the brain. However, the relationship between the nanomedicines' physical and chemical properties and its ability to travel across the brain remains incompletely understood. The main challenge is the lack of a reliable drug testing model for brain angiogenesis. Recently, microfluidic platforms (known as "lab-on-a-chip" or LOCs) have been developed to mimic the brain micro-vasculature related events, such as vasculogenesis, angiogenesis, inflammation, etc. The LOCs are able to closely replicate the dynamic conditions of the human brain and could be reliable platforms for drug screening applications. There are still many technical difficulties in establishing uniform and reproducible conditions, mainly due to the extreme complexity of the human brain. In this paper, we review the prospective of LOCs in the development of nanomedicines for brain angiogenesis-related conditions.

Enhanced Internalization of Indian Ayurvedic Swarna Bhasma (Gold Nanopowder) for Effective Interaction with Human Cells.

In the ancient traditional Indian Ayurvedic system of natural healing, gold nanoparticles (Swarna Bhasma, gold ash) have been used for its therapeutic benefits as far back as 2500 B.C. Ayurvedic medicinal preparations are complex mixtures that include many plant-derived products and metals. Bhasmas date as far back as the 8th century and are made by samskaras (processings), such as shodhana (purification and potentiation), jarana (roasting), and marana (incineration, trituration) in the presence of plant products, including juices and concoctions. Previous studies characterized the physical properties of gold ash, and the mechanisms of its entry into human cells, but only preliminary data exist on its toxicity. Before using nanoparticles for therapeutic application, it is extremely important to study their toxicity and cellular internalization. In the present study, various imaging techniques were used to investigate Swarna Bhasma's (gold nanopowder) toxicity in both cancerous and noncancerous cells (HeLa and HFF-1) and to characterize its spectral properties. The results showed that gold ash particles had no impact on the cellular viability of both HeLa and HFF-1 cells, even at high concentrations or long incubation times. Moreover, it was found that the internalization level of Swarna Bhasma to cells may be improved by mechanical breaking of the large aggregates into smaller agglomerates. Hyperspectral images revealed that after breaking, the small agglomerates have different spectral properties in cells, compared to the original aggregates, suggesting that size of particles is instrumental for the subcellular interaction with human cells.

Comparative study on cellular entry of incinerated ancient gold particles (Swarna Bhasma) and chemically synthesized gold particles.

Gold nanoparticles (AuNPs) are used for a number of imaging and therapeutic applications in east and western part of the world. For thousands of years, the traditional Indian Ayurvedic approach to healing involves the use of incinerated gold ash, prepared with a variety of plant extracts and minerals depending on the region. Here, we describe the characterization of incinerated gold particles (IAuPs) in HeLa (human cells derived from cervical cancer) and HFF-1 (human foreskin fibroblast cells) in comparison to synthesized citrate-capped gold nanoparticles (AuNPs). We found that while individual IAuP crystallites are around 60 nm in size, they form large aggregates with a mean diameter of 4711.7 nm, some of which can enter cells. Fewer cells appeared to have IAuPs compared to AuNPs, although neither type of particle was toxic to cells. Imaging studies revealed that IAuPs were in vesicles, cytosol, or in the nucleus. We found that their nuclear accumulation likely occurred after nuclear envelope breakdown during cell division. We also found that larger IAuPs entered cells via macropinocytosis, while smaller particles entered via clathrin-dependent receptor-mediated endocytosis.

A brief review on microfluidic platforms for hormones detection.

Lab-on-chip technology is attracting great interest due to its potential as miniaturized devices that can automate and integrate many sample-handling steps, minimize consumption of reagent and samples, have short processing time and enable multiplexed analysis. Microfluidic devices have demonstrated their potential for a broad range of applications in life sciences, including point-of-care diagnostics and personalized medicine, based on the routine diagnosis of levels of hormones, cancer markers, and various metabolic products in blood, serum, etc. Microfluidics offers an adaptable platform that can facilitate cell culture as well as monitor their activity and control the cellular environment. Signaling molecules released from cells such as neurotransmitters and hormones are important in assessing the health of cells and the effect of drugs on their functions. In this review, we provide an insight into the state-of-art applications of microfluidics for monitoring of hormones released by cells. In our works, we have demonstrated efficient detection methods for bovine growth hormones using nano and microphotonics integrated microfluidics devices. The bovine growth hormone can be used as a growth promoter in dairy farming to enhance the milk and meat production. In the recent years, a few attempts have been reported on developing very sensitive, fast and low-cost methods of detection of bovine growth hormone using micro devices. This paper reviews the current state-of-art of detection and analysis of hormone using integrated optical micro and nanofluidics systems. In addition, the paper also focuses on various lab-on-a-chip technologies reported recently, and their benefits for screening growth hormones in milk.

Technical note: A portable on-chip assay system for absorbance and plasmonic detection of protein hormone in milk.

This paper reports a portable device and method to extract and detect protein hormone in milk samples. Recombinant protein hormone spiked into milk samples was extracted by solid-phase extraction, and detection was carried out using the plasmonic property of gold nanoislands deposited on a glass substrate. Trace levels of hormone spiked in milk were analyzed by their optical absorbance property using a microfluidic chip. We built a portable assay system using disposable lab-on-chip devices. The proposed method is able to detect spiked recombinant protein hormone in milk at concentrations as low as 5ng/mL.

Rapid microwave-induced synthesis of gold-polydimethylsiloxane nanocomposites for biosensing of proteins.

In this paper a novel in-situ microwave-induced synthesis of the gold-polydimethylsiloxane nanocomposite is presented. Microwave-induced synthesis has the advantages of a very short reaction time, small particle size and narrow size distribution of the particles. The ethanol solution of gold chloroauric acid is used as the precursor solution. The mechanism of formation and growth of nanoparticles are discussed in detail. UV/Vis spectroscopy and SEM imaging were used to characterize the optical properties and the size distribution of the particles. To improve the sensing properties of the nanocomposite, an annealing process were used. The results show that the annealed samples have the high sensitivity of 102 nm/RIU toward the surrounding medium which makes the nanocomposite suitable for biosensing applications. In addition, the elasticity of the platform in the presence of gold nanoparticles was found to be enhanced up to 20%. Finally, the immunosensing of the bovine growth hormone was performed by using the localized surface plasmon resonance (LSPR) band of gold nanoparticles. The results demonstrate suitability of the nanocomposite platform for biosensing applications. The results are highly relevant for microfluidic-based biosensors.

Integration of gold nanoparticles in PDMS microfluidics for lab-on-a-chip plasmonic biosensing of growth hormones.

Gold nanoparticles were synthesized in a poly(dimethylsiloxane) (PDMS) microfluidic chip by using an in-situ method, on the basis of reductive properties of the cross-linking agent of PDMS. The proposed integrated device was further used as a sensitive and low-cost LSPR-based biosensor for the detection of polypeptides. Synthesis of nanoparticles in the microfluidic environment resulted in improvement of size distribution with only 8% variation, compared with the macro-environment that yields about 67% variation in size. The chemical kinetics of the in-situ reaction in the microfluidic environment was studied in detail and compared with the reaction carried out at the macro-scale. The effect of temperature and gold precursor concentration on the kinetics of the reaction was investigated and the apparent activation energy was estimated to be Ea*=30 kJ/mol. The sensitivity test revealed that the proposed sensor has a high sensitivity of 74 nm/RIU to the surrounding medium. The sensing of bovine growth hormone also known as bovine somatotropin (bST) shows that the proposed biosensor can reach a detection limit of as low as 3.7 ng/ml (185 pM). The results demonstrate the successful integration of microfluidics and nanoparticles which provides a potential alternative for protein detection in clinical diagnostics.

PDMS-gold nanocomposite platforms with enhanced sensing properties.

Gold-poly(dimethyl siloxoxane) (Au-PDMS) nanocomposite films with a high elasticity were fabricated for sensing experiments. The nanocomposite was prepared by a novel in-situ method by using the ethanol solution of the chloroauric acid. The high rate of permeation of ethanol in the polymer film, compared to an aqueous solution, allows the introduction of the gold precursor into the polymer network with a higher rate and, thus the reduction reaction is accelerated. The strong hydrophobicity of the as-prepared films precludes the diffusion of aqueous solutions of biomolecules in the polymer network, essential for sensing purposes. In order to modify the morphology and the surface properties of the samples, they have been heat-treated and the polymer network has been expanded mechanically by repeated swellings and shrinkages. As a result, the free volume of the polymer is increased substantially and thus, the biosensing capability of the material is improved. The effect of gold nanoparticles on the porosity and the mechanical properties of the material has been studied. The highest value of the sensitivity (around 70 nm/RIU) has been obtained for the samples that were annealed and, subsequently swollen in toluene. Biosensing experiments involving antigen-antibody interactions showed a high sensitivity. The results of this work are relevant for sensing in a microfluidic environment.

Gold nanoisland structures integrated in a lab-on-a-chip for plasmonic detection of bovine growth hormone.

Three-dimensional gold nanostructures fabricated through a novel convective assembly method are treated thermally to obtain a nanoisland morphology. The new structure is proved to be adequate for the detection of bovine growth hormone, by using an immunoassay method based on the localized surface plasmon resonance band of gold. The nanoisland structures are integrated into a microfluidic device and the spectral measurements are carried out by introducing the device directly in the light beam of a ultraviolet-visible spectrophotometer. The principal motivation for this work is the need for a simple and rapid method of detection of hormone levels in milk and milk products.