paper-based-devices-for-isolation-characterization-extracellular

Paper-based Devices for Isolation and Characterization of Extracellular Vesicles

Extracellular vesicles (EVs), membranous particles released from various types of cells, hold a great potential for clinical applications. They contain ...

Fabricating Cotton Analytical Devices

A robust, low-cost analytical device should be user-friendly, rapid, and affordable. Such devices should also be able to operate with scarce samples and ...

Taichung Veterans General Hospital

Polyethylene glycol (PEG) at various molecular weights (MWs) has been regarded as a wonder molecule in biomedical applications. For instance, PEG serves as a unique moiety for pegylation of "biobetter" drug development, PEG provides controlled-release and preserved activity of biologics, and PEG modified surface works as an antibiofouling surface. The primary characteristics of PEG molecules used in relevant applications have been attributed mainly to the hydration behavior in aqueous solutions. However, the effects on the solvation of solutes in solution caused by presenting PEG molecules as a cosolvent, as well as the thermodynamics aspect of the hydration behavior of PEG in solution, have not been well documented. The solvation behavior of solutes, such as protein, with PEG as a cosolvent, indicates the success of PEG applications, such as biofouling and controlled release. In this investigation, we examined the effects of a buffer solution containing PEG molecules on the solution behavior of solute and the interactions between solid surfaces with solutes. We adapted the study by selecting a lysozyme as a solute in a buffer solution with either ammonium sulfate (kosmotrope) or sodium chloride (chaotrope) and anionic resin (SP-Sepharose) as solid surfaces. The experiments primarily involved binding equilibrium measurements and thermodynamics analysis. The results revealed that, in both saline buffers, adding PEG increases the binding affinity between the lysozyme and the resin, similar to kosmotropic salt in the examined salt concentrations. The thermodynamics analyses involving microcalorimetric measurements show that the bindings are mainly driven by enthalpy, indicating that electrostatic interaction was the primary binding force under these experimental conditions. The variations of the enthalpy and entropy of the binding thermodynamics when adding PEG to different salt types in the buffer solution showed opposite behavior, and the results support the concept of kosmotrope-like behavior of PEG. The equilibrium and thermodynamics data demonstrate that PEG has a kosmotrope-like hydration behavior, and the extent of kosmotrope-like behavior depends on the molecular weight of PEG with the outcomes of various molecular weights of PEG being added to the binding solution. The results of this study provide essential knowledge for PEG as an additive (or cosolvent) in various research applications.

Kosmotrope-like hydration behavior of polyethylene glycol from microcalorimetry and binding isotherm measurements.

The vascular endothelial growth factor (VEGF) level in aqueous humor has been used as an indicator to monitor specific diseases in the retinal ischemic condition. For clinical diagnosis, only about 200 μL of aqueous humor can be collected from the anterior chamber before the threat of anterior chamber collapse. It is necessary to develop an inexpensive diagnostic approach with the characteristics of highly sensitive, short operation duration, and requires small clinical sample quantities. To achieve the main objective of this study, we first prepared bevacizumab to be conjugated with HRP. We then deposited 2 μL aqueous humor from patients with different diseases onto each test zone of paper-based 96-well plates. After the colorimetric results were performed via ELISA protocol, the output signals were recorded using a commercial desktop scanner for analysis. In this study, only 2 μL from the aqueous humor of each patient was required for paper-based ELISA. The mean aqueous VEGF level was 14.4 pg/mL from thirteen patients (N = 13) with senile cataract as the control. However, the mean aqueous VEGF level from other patients with proliferative diabetic retinopathy (N = 14), age-related macular degeneration (N = 17), and retinal vein occlusion (N = 10) showed VEGF increases to 740.1 pg/mL, 383 pg/mL, and 219.4 pg/mL, respectively.

Monitoring the VEGF level in aqueous humor of patients with ophthalmologically relevant diseases via ultrahigh sensitive paper-based ELISA.

The aim of this study was to investigate the relationship between pterygium and a decrease in the corneal endothelial cell density (ECD) in patients with unilateral primary pterygium.

Pterygium is related to a decrease in corneal endothelial cell density.

Caspofungin exhibits potent antifungal activities against Candida and Aspergillus species. The elimination rate and retinal toxicity of caspofungin were determined in this study to assess its pharmacokinetics and safety in the treatment of fungal endophthalmitis. Intravitreal injections of 50 μg/0.1 ml of caspofungin were administered to rabbits. Levels of caspofungin in the vitreous and aqueous humors were determined using high-performance liquid chromatography (HPLC) at selected time intervals (10 min and 1, 2, 4, 8, 16, 24, and 48 h), and the half-lives were calculated. Eyes were intravitreally injected with caspofungin to obtain concentrations of 10 μg/ml, 50 μg/ml, 100 μg/ml, and 200 μg/ml. Electroretinograms were recorded 4 weeks after injections, and the injected eyes were examined histologically. The concentrations of intravitreal caspofungin at various time points exhibited an exponential decay with a half-life of 6.28 h. The mean vitreous concentration was 6.06 ± 1.76 μg/ml 1 h after intravitreal injection, and this declined to 0.47 ± 0.15 μg/ml at 24 h. The mean aqueous concentration showed undetectable levels at all time points. There were no statistical differences in scotopic a-wave and b-wave responses between control eyes and caspofungin-injected eyes. No focal necrosis or other abnormality in retinal histology was observed. Intravitreal caspofungin injection may be considered to be an alternative treatment for fungal endophthalmitis based on its antifungal activity, lower retinal toxicity, and lower elimination rate in the vitreous. More clinical data are needed to determine its potential role as primary therapy for fungal endophthalmitis.

Pharmacokinetics and safety of intravitreal caspofungin.

A good diagnostic procedure avoids wasting medical resources, is easy to use, resists contamination, and provides accurate information quickly to allow for rapid follow-up therapies. We developed a novel diagnostic procedure using a "cotton-based diagnostic device" capable of real-time detection, i.e., in vitro diagnostics (IVD), which avoids reagent contamination problems common to existing biomedical devices and achieves the abovementioned goals of economy, efficiency, ease of use, and speed. Our research reinforces the advantages of an easy-to-use, highly accurate diagnostic device created from an inexpensive and readily available U.S. FDA-approved material (i.e., cotton as flow channel and chromatography paper as reaction zone) that adopts a standard calibration curve method in a buffer system (i.e., nitrite, BSA, urobilinogen and uric acid assays) to accurately obtain semi-quantitative information and limit the cross-contamination common to multiple-use tools. Our system, which specifically targets urinalysis diagnostics and employs a multiple biomarker approach, requires no electricity, no professional training, and is exceptionally portable for use in remote or home settings. This could be particularly useful in less industrialized areas.

Cotton-based diagnostic devices.

The purpose of this article is to demonstrate the capacity of paper-based ELISA (P-ELISA) to monitor VEGF in patients requiring treatment for vision-threatening diseases. The most commonly encountered vision-threatening diseases are age-related macular degeneration (AMD) and diabetic retinopathy (DR), both of which may require short-term or life-long anti-VEGF injection treatment therapy. Accurate measurement of VEGF concentration in aqueous humor can provide significant and timely information to diagnose the disease state. Adequate and precise therapy may consequently be provided. At odds with conventional diagnostic approaches is the fact that a maximum of only 200 microliters of aqueous humor can be safely removed from the eye for testing. Fortunately, new diagnostic platforms, such as P-ELISA, require only minute volumes, i.e., approximately 2 microliters per test "well" and approximately 40 microliters total to quantify VEGF levels, and the testing process takes less than an hour. Thus, point-of-care (POC) diagnostics, such as P-ELISA, should be examined and improved upon as needed in order to develop an efficient tool for outpatient clinics and others to obtain semi-quantitative results that might facilitate accurate dosing of anti-VEGF treatment and delay or prevent the progression of AMD and DR.

Monitoring VEGF levels with low-volume sampling in major vision-threatening diseases: age-related macular degeneration and diabetic retinopathy.

This manuscript describes the development and clinical testing of a paper-based, metabolic assay designed for rapid, semi-quantitative measurement of organophosphate poisoning. Paper-based platforms, including point-of-care devices and 96-well plates, provided semi-quantitative information regarding the concentration of AchE (a biomarker for organophosphate poisoning). The paper-based 96-well-plate developed and implemented in this study was used to measure the level of organophosphate poisoning in three different clinical patients. Results were comparable to those obtained using conventional hospital methods currently considered the "gold standard". This diagnostic device offers several advantages over conventional methods, including short operating time (twice as fast as conventional methods), procedure simplicity, and reduced fabrication cost. With further commercialization efforts, the methods described in this manuscript could be applied to a wide range of potential diagnostic applications in the field.

Evaluating organophosphate poisoning in human serum with paper.

Escherichia coli is a generic indicator of fecal contamination, and certain serotypes cause food- and water-borne illness such as O157:H7. In the clinic, detection of bacteriuria, which is often due to E. coli, is critical before certain surgical procedures or in cases of nosocomial infection to prevent further adverse events such as postoperative infection or sepsis. In low- and middle-income countries, where insufficient equipment and facilities preclude modern methods of detection, a simple, low-cost diagnostic device to detect E. coli in water and in the clinic will have significant impact. We have developed a simple paper-based colorimetric platform to detect E. coli contamination in 5h. On this platform, the mean color intensity for samples with 10(5)cells/mL is 0.118±0.002 (n=4), and 0.0145±0.003 (P<0.01⁎⁎) for uncontaminated samples. This technique is less time-consuming, easier to perform, and less expensive than conventional methods. Thus, paper-based ELISA is an innovative point-of-care diagnostic tool to rapidly detect E. coli, and possibly other pathogens when customized as appropriate, especially in areas that lack advanced clinical equipment.

Min-Yen Hsu

Department of Ophthalmology

Taichung Veterans General Hospital

Paper-based Devices for Isolation and Characterization of Extracellular Vesicles

Fabricating Cotton Analytical Devices