Multiplexing techniques for measurement of the immunomodulatory effects of particulate materials: Precautions when testing micro- and nano-particles.

Particulate materials at nano- and micro-scales have widespread pharmaceutical and medical applications. Understanding the interactions of these materials with biological systems is crucial for the design of clinically-viable biomaterials of high safety profiles. Immunomodulatory effects of particulate materials can be studied via multiplexing techniques that are capable of measuring up to 500 biomarkers in a few microliters of biological samples. However, there are several challenges towards the use of multiplexing techniques for testing the ability of nanomaterials to induce the release of various biomarkers. As one of the potential challenges, the adsorption of biomarkers on surfaces or within internal structures of nano- or micro-particles has been explored to a lesser extent, although it can lead to biased conclusions and data misinterpretation. Herein, we provide technical details on the use of multiplexing techniques for the evaluation of immunomodulatory effects of nanoparticulates. The same principles can also be applied for the assessment of microparticles. Importantly, precautions to avoid artifacts and data misinterpretation, due to interactions between particles and biomarkers, are provided.

Influence of novel design alteration of pedicle screw on pull-out strength: A finite element study.

BACKGROUND: We conducted a finite element study to assess the effectiveness of a novel pedicle screw design with two alterations in the distal and proximal portions. METHODS: Finite element (FE) models of 24 vertebrae were constructed using computed tomographic data. Pull-out strength of 4 different pedicle screws were compared. The basic screw design was a dual threaded one (PS0), in which the proximal portion is double-threaded (cortical thread), and the distal portion is single-threaded (cancellous thread). In PS1, the inter-thread double-core shape was added to PS0 in the distal portion. Compared to PS0, in PS2, the proximal portion was elongated by 5 mm. PS3 had both PS1 and PS2 features. In addition, the 24 vertebrae were classified into 3 groups based on volumetric bone mineral density (vBMD) of the vertebral body: low <120 mg/cm3, moderate 120-170 mg/cm3, and high >170 mg/cm3. RESULTS: The mean pull-out strengths (±SD) were 1137 ± 500 N, 1188 ± 520 N, 1191 ± 512 N, and 1242 ± 538 N for PS0, PS1, PS2, and PS3, respectively. In PS1, there was significant difference in the incremental ratio of pull-out strength to PS0 between the low and high vBMD groups (3.7 ± 1.6% vs. 5.0 ± 1.0%, p = 0.006). In PS2, there was a significant difference in the incremental ratio to PS0 between the moderate and high vBMD groups (7.6 ± 4.0% vs. 3.3 ± 1.8%, p < 0.001). In PS3, there was a significant difference in the incremental ratio to PS0 between the moderate and high vBMD groups (12.1 ± 4.8% vs. 8.5 ± 2.1%, p = 0.003). CONCLUSIONS: The two design alterations showed the combined additive effect in the PS3 design. The moderate vBMD group has a balanced bone property to reflect the combined effects of the PS1 and PS2 design alterations.

Evaluation of Magnesium-based Medical Devices in Preclinical Studies: Challenges and Points to Consider.

Absorbable metallic implants have been under investigation for more than a century. Animal and human studies have shown that magnesium (Mg) alloys can be safely used in bioresorbable scaffolds. Several cardiovascular and orthopedic biodegradable metallic devices have recently been approved for use in humans. Bioresorbable Mg implants present many advantages when compared to bioabsorbable polymer or nonabsorbable metallic implants, including similar strength and mechanical properties as existing implant-grade metals without the drawbacks of permanence or need for implant removal. Imaging visibility is also improved compared to polymeric devices. Additionally, with Mg-based cardiovascular stents, the risk of late stent thrombosis and need for long-term anti-platelet therapy may be reduced as the host tissue absorbs the Mg degradation products and the morphology of the vessel returns to a near-normal state. Absorbable Mg implants present challenges in the conduct of preclinical animal studies and interpretation of pathology data due to their particular degradation process associated with gas production and release of by-products. This article will review the different uses of Mg implants, the Mg alloys, the distinctive degradation features of Mg, and the challenges confronting pathologists at tissue collection, fixation, imaging, slide preparation, evaluation, and interpretation of Mg implants.

Pathology of Bioabsorbable Implants in Preclinical Studies.

Bioabsorbable implants can be advantageous for certain surgical tissue bioengineering applications and implant-assisted tissue repair. They offer the obvious benefits of nonpermanence and eventual restoration of the native tissue's biomechanical and immunological properties, while providing a structural scaffold for healing and a route for additional therapies (i.e., drug elution). They present unique developmental, imaging, and histopathological challenges in the conduct of preclinical animal studies and in interpretation of pathology data. The bioabsorption process is typically associated with a gradual decline (over months to years) in structural strength and integrity and may also be associated with cellular responses such as phagocytosis that may confound interpretation of efficacy and safety end points. Additionally, as these implants bioabsorb, they become increasingly difficult to isolate histologically and thus imaging modalities such as microCT become very valuable to determine the original location of the implants and to assess the remodeling response in tandem with histopathology. In this article, we will review different types of bioabsorbable implants and commonly used bioabsorbable materials; additionally, we will address some of the most common challenges and pitfalls confronting histologists and pathologists in collecting, handling, imaging, preparing tissues through histology, evaluating, and interpreting study data associated with bioabsorbable implants.

Roles of the screw types, proximity and anterior band wiring in the surgical fixation of transverse patellar fractures: a finite element investigation.

BACKGROUND: Cannulated screws with an anterior wire are currently used for managing transverse patellar fracture. However, the addition of anterior wiring with various types of screws via open surgery to increase the mechanical stability is yet to be determined. Hence, this study aimed to compare the mechanical behaviors of a fractured patella fixed with various screws types and at various screw locations with and without the anterior wire. The present study hypothesized that using the anterior wire reduces the fracture gap formation. METHODS: A finite element (FE) model containing a fractured patella fixed with various types of cannulated screws and anterior wiring was created in this study. Three types of screws, namely partial thread, full thread, and headless compression screws, and two screw depths, namely 5 and 10 mm away from the anterior surface of the patella, were included. The effect of the anterior wire was clarified by comparing the results of surgical fixation with and without the wire. Two magnitudes and two loading directions were used to simulate and examine the mechanical responses of the fractured patella with various fixation conditions during knee flexion/extension. RESULTS: Compared with partial thread and headless compression screws, the full thread screw increased the stability of the fractured patella by reducing fragment displacement, fracture gap formation, and contact pressure while increasing the contact area at the fracture site. Under 400-N in the direction 45°, the full thread screw with 5-mm placement reduced the gap formation by 86.7% (from 2.71 to 0.36 mm) and 55.6% (from 0. 81 to 0. 36 mm) compared with the partial thread screw with 10-mm placement, respectively without and with the anterior wire. CONCLUSION: The anterior wire along with the full thread screw is preferentially recommended for maintaining the surgical fixation of the fractured patella. Without the use of anterior wiring, the full thread screw with 5-mm placement may be considered as a less invasive alternative; however, simple screw fixation at a deeper placement (10 mm) is least recommended for the fixation of transverse patellar fracture.

Fully integrated fused quartz acoustic horns for structural health monitoring.

Non-destructive acoustic structural sensing is an imperative technology, applicable to many different fields such as aerospace and civil engineering. To maintain a high sensitivity or to mitigate acoustic loss, it is important to increase the signal-to-noise ratio by improving coupling efficiency from acoustic sources to the object under test, such as an acoustic waveguide. Here, a fully integrated fused quartz horn design is combined with a fused quartz acoustic waveguide. The resulting system is intended to demonstrate a high accuracy low cost alternative to current sensing systems and the present article report on the viability of using a merged acoustic horn and waveguide.

Evaluating the combined effects of gravity and compression on the fluid handling properties of surgical dressings.

OBJECTIVE: A new test method is described which provides a convenient technique for assessing the combined effects of gravity and compression on the fluid handling characteristics of surgical dressing which can have important implications for their use in the treatment of venous ulcers. Simple ways of improving use of the fluid handling capacity of products currently used clinically are suggested, and the possibility of developing a dressing specifically designed to resist gravitational effects is discussed.

Transdermal Microneedles-A Materials Perspective.

Transdermal drug delivery is an emerging field in the pharmaceutical remit compared with conventional methods (oral and parenteral). Microneedle (MN)-based devices have gained significant interest as a strategy to overcome the skin's formidable barrier: the stratum corneum. This approach provides a less invasive, more efficient, patient friendly method of drug delivery with the ability to incorporate various therapeutic agents including macromolecules (proteins and peptides), anti-cancer agents and other hydrophilic and hydrophobic compounds. This short review attempts to assess the various materials involved in the fabrication of MNs as well as incorporation of other excipients to improve drug delivery for novel medical devices. The focus will be on polymers, metals and other inorganic materials utilised for MN drug delivery, as well as their application, limitations and future work to be carried out.

Multimodal Measurements of Single-Molecule Dynamics Using FluoRBT.

Single-molecule methods provide direct measurements of macromolecular dynamics, but are limited by the number of degrees of freedom that can be followed at one time. High-resolution rotor bead tracking (RBT) measures DNA torque, twist, and extension, and can be used to characterize the structural dynamics of DNA and diverse nucleoprotein complexes. Here, we extend RBT to enable simultaneous monitoring of additional degrees of freedom. Fluorescence-RBT (FluoRBT) combines magnetic tweezers, infrared evanescent scattering, and single-molecule FRET imaging, providing real-time multiparameter measurements of complex molecular processes. We demonstrate the capabilities of FluoRBT by conducting simultaneous measurements of extension and FRET during opening and closing of a DNA hairpin under tension, and by observing simultaneous changes in FRET and torque during a transition between right-handed B-form and left-handed Z-form DNA under controlled supercoiling. We discover unanticipated continuous changes in FRET with applied torque, and also show how FluoRBT can facilitate high-resolution FRET measurements of molecular states, by using a mechanical signal as an independent temporal reference for aligning and averaging noisy fluorescence data. By combining mechanical measurements of global DNA deformations with FRET measurements of local conformational changes, FluoRBT will enable multidimensional investigations of systems ranging from DNA structures to large macromolecular machines.

Tensile testing of the mechanical behavior of the human periodontal ligament.

BACKGROUND: The periodontal ligament (PDL) plays a key role in alveolar bone remodeling and resorption during tooth movements. The prediction of tooth mobility under functional dental loads requires a deep understanding of the mechanical behavior of the PDL, which is a critical issue in dental biomechanics. This study was aimed to examine the mechanical behavior of the PDL of the maxillary central and lateral incisors from human. The experimental results can contribute to developing an accurate constitutive model of the human PDL in orthodontics. METHODS: The samples of human incisors were cut into three slices. Uniaxial tensile tests were conducted under different loading rates. The transverse sections (cervical, middle and apex) normal to the longitudinal axis of the root of the tooth were used in the uniaxial tensile tests. Based on a bilinear simplification of the stress-strain relations, the elastic modulus of the PDL was calculated. The values of the elastic modulus in different regions were compared to explore the factors that influence the mechanical behavior of the periodontal ligament. RESULTS: The obtained stress-strain curves of the human PDL were characterized by a bilinear model with two moduli (E1 and E2) for quantifying the elastic behavior of the PDL from the central and lateral incisors. Statistically significant differences of the elastic modulus were observed in the cases of 1, 3, and 5 N loading levels for the different teeth (central and lateral incisors). The results showed that the mechanical property of the human incisors' PDLs is dependent on the location of PDL (ANOVA, P = 0.022, P < 0.05). The elastic moduli at the middle planes were greater than at the cervical and apical planes. However, at the cervical, middle, and apical planes, the elastic moduli of the mesial and distal site were not significantly different (ANOVA, P = 0.804, P > 0.05). CONCLUSIONS: The values of elastic modulus were determined in the range between 0.607 and 4.274 MPa under loads ranging from 1 to 5 N. The elastic behavior of the PDL is influenced by the loading rate, tooth type, root level, and individual variation.

Structural Model for Viscoelastic Properties of Pericardial Bioprosthetic Valves.

The benefit of bioprosthetic aortic valve over mechanical valve replacements is the release of thromboembolism and digression of long-term anticoagulation treatment. The function of bioprostheses and their efficiency is known to depend on the mechanical properties of the leaflet tissue. So it is necessary to select a suitable tissue for the bioprosthesis. The purpose of the present study is to clarify the viscoelastic behavior of bovine, equine, and porcine pericardium. In this study, pericardiums were compared mechanically from the viscoelastic aspect. After fixation of the tissues in glutaraldehyde, first uniaxial tests with different extension rates in the fiber direction were performed. Then, the stress relaxation tests in the fiber direction were done on these pericardial tissues by exerting 20, 30,40, and 50% strains. After evaluation of viscoelastic linearity, the Prony series, quasilinear viscoelastic (QLV) and modified superposition theory were applied to the stress relaxation data. Finally, the parameters of these constitutive models were extracted for each pericardium tissue. All three tissues exhibited a decrease in relaxation rate with elevating strain, indicating the nonlinear viscoelastic behavior of these tissues. The three-term Prony model was selected for describing the linear viscoelasticity. Among different models, the QLV model was best able to capture the relaxation behavior of the pericardium tissues. More stiffness of porcine pericardium was observed in comparison to the two other pericardium tissues. The relaxation percentage of porcine pericardium was less than the two others. It can be concluded that porcine pericardium behaves more as an elastic and less like a viscous tissue in comparison to the bovine and equine pericardium.

Validation of a Custom Instrumented Retainer Form Factor for Measuring Linear and Angular Head Impact Kinematics.

Head impact exposure in popular contact sports is not well understood, especially in the youth population, despite recent advances in impact-sensing technology which has allowed widespread collection of real-time head impact data. Previous studies indicate that a custom-instrumented mouthpiece is a superior method for collecting accurate head acceleration data. The objective of this study was to evaluate the efficacy of mounting a sensor device inside an acrylic retainer form factor to measure six-degrees-of-freedom (6DOF) head kinematic response. This study compares 6DOF mouthpiece kinematics at the head center of gravity (CG) to kinematics measured by an anthropomorphic test device (ATD). This study found that when instrumentation is mounted in the rigid retainer form factor, there is good coupling with the upper dentition and highly accurate kinematic results compared to the ATD. Peak head kinematics were correlated with r2 > 0.98 for both rotational velocity and linear acceleration and r2 = 0.93 for rotational acceleration. These results indicate that a rigid retainer-based form factor is an accurate and promising method of collecting head impact data. This device can be used to study head impacts in helmeted contact sports such as football, hockey, and lacrosse as well as nonhelmeted sports such as soccer and basketball. Understanding the magnitude and frequency of impacts sustained in various sports using an accurate head impact sensor, such as the one presented in this study, will improve our understanding of head impact exposure and sports-related concussion.

An Ergonomic Testing System for the First Metatarsophalangeal Joint Stiffness.

Osteoarthritis sufferers commonly have first metatarsophalangeal joint (MTPJ) problems in which articular surfaces are changed permanently due to fatigue. Therefore, medical devices for early diagnosis would increase the opportunity for prevention of disease progression. In previous studies on stiffness of the first MTPJ many details, although functionally of great importance, have not been fully considered including: design and size of the device, tribology consideration, and errors from device. Therefore, the motivation of our research was to enhance the device design by reducing the size of the device, and device design was enhanced by minimizing measurement errors through development of a new ergonomic left and right foot instrument located medial to the first MTPJ (instead of beneath the foot). The first MTPJ stiffness (N mm/kg radian) measurement was taken on 28 subjects with two replicates per subject by the same tester. The first MTPJ stiffness ranged from 3.49 to 14.42 N mm/kg radian with the mean (SD) value of 8.28 (3.15) N mm/kg radian for the left feet and 3.91 to 11.90 N mm/kg radian with the mean (SD) value of 7.65 (2.07) N mm/kg radian for the right feet. Reliability evaluation was measured using intraclass correlation coefficient and described an excellent reliability between two tests.

A Novel Small-Specimen Planar Biaxial Testing System With Full In-Plane Deformation Control.

Simulations of soft tissues require accurate and robust constitutive models, whose form is derived from carefully designed experimental studies. For such investigations of membranes or thin specimens, planar biaxial systems have been used extensively. Yet, all such systems remain limited in their ability to: (1) fully prescribe in-plane deformation gradient tensor F2D, (2) ensure homogeneity of the applied deformation, and (3) be able to accommodate sufficiently small specimens to ensure a reasonable degree of material homogeneity. To address these issues, we have developed a novel planar biaxial testing device that overcomes these difficulties and is capable of full control of the in-plane deformation gradient tensor F2D and of testing specimens as small as ∼4 mm × âˆ¼4 mm. Individual actuation of the specimen attachment points, combined with a robust real-time feedback control, enabled the device to enforce any arbitrary F2D with a high degree of accuracy and homogeneity. Results from extensive device validation trials and example tissues illustrated the ability of the device to perform as designed and gather data needed for developing and validating constitutive models. Examples included the murine aortic tissues, allowing for investigators to take advantage of the genetic manipulation of murine disease models. These capabilities highlight the potential of the device to serve as a platform for informing and verifying the results of inverse models and for conducting robust, controlled investigation into the biomechanics of very local behaviors of soft tissues and membrane biomaterials.

Determination of Elastic Modulus in Mouse Bones Using a Nondestructive Micro-Indentation Technique Using Reference Point Indentation.

The determination of the elastic modulus of bone is important in studying the response of bone to loading and is determined using a destructive three-point bending method. Reference point indentation (RPI), with one cycle of indentation, offers a nondestructive alternative to determine the elastic modulus. While the elastic modulus could be determined using a nondestructive procedure for ex vivo experiments, for in vivo testing, the three-point bending technique may not be practical and hence RPI is viewed as a potential alternative and explored in this study. Using the RPI measurements, total indentation distance (TID), creep indentation distance, indentation force, and the unloading slope, we have developed a numerical analysis procedure using the Oliver-Pharr (O/P) method to estimate the indentation elastic modulus. Two methods were used to determine the area function: (1) Oliver-Pharr (O/P-based on a numerical procedure) and (2) geometric (based on the calculation of the projected area of indentation). The indentation moduli of polymethyl methacrylate (PMMA) calculated by the O/P (3.49-3.68 GPa) and geometric (3.33-3.49 GPa) methods were similar to values in literature (3.5-4 GPa). In a study using femurs from C57Bl/6 mice of different ages and genders, the three-point bending modulus was lower than the indentation modulus. In femurs from 4 to 5 months old TOPGAL mice, we found that the indentation modulus from the geometric (5.61 ± 1.25 GPa) and O/P (5.53 ± 1.27 GPa) methods was higher than the three-point bending modulus (5.28 ± 0.34 GPa). In females, the indentation modulus from the geometric (7.45 ± 0.86 GPa) and O/P (7.46 ± 0.92 GPa) methods was also higher than the three-point bending modulus (7.33 ± 1.13 GPa). We can conclude from this study that the RPI determined values are relatively close to three-point bending values.

The effect of abrading and cutting instruments on machinability of dental ceramics.

The aim was to investigate the effect of machining instruments on machinability of dental ceramics. Four dental ceramics, including two zirconia ceramics were machined by three types (SiC, diamond vitrified, and diamond sintered) of wheels with a hand-piece engine and two types (diamond and carbide) of burs with a high-speed air turbine. The machining conditions used were abrading speeds of 10,000 and 15,000 r.p.m. with abrading force of 100 gf for the hand-piece engine, and a pressure of 200 kPa and a cutting force of 80 gf for the air-turbine hand-piece. The machinability efficiency was evaluated by volume losses after machining the ceramics. A high-abrading speed had high-abrading efficiency (high-volume loss) compared to low-abrading speed in all abrading instruments used. The diamond vitrified wheels demonstrated higher volume loss for two zirconia ceramics than those of SiC and diamond sintered wheels. When the high-speed air-turbine instruments were used, the diamond points showed higher volume losses compared to the carbide burs for one ceramic and two zirconia ceramics with high-mechanical properties. The results of this study indicated that the machinability of dental ceramics depends on the mechanical and physical properties of dental ceramics and machining instruments. The abrading wheels show autogenous action of abrasive grains, in which ground abrasive grains drop out from the binder during abrasion, then the binder follow to wear out, subsequently new abrasive grains come out onto the instrument surface (autogenous action) and increase the grinding amount (volume loss) of grinding materials.

Monitoring and Assessing the Degradation Rate of Magnesium-Based Artificial Bone In Vitro Using a Wireless Magnetoelastic Sensor.

A magnetoelastic-based (MB) sensor was employed as a novel method to monitor and assess the degradation rate of magnesium-based artificial bone (MBAB) in vitro, which can be used as an implant to repair a bone defect, providing a quantitative method to depict the degradation rate of MBAB. MBABs were fabricated by the Pro/Engineering software and a precision machine tool using high-purity (HP) magnesium. The MB sensor was embedded in the neutral surface of MBAB by an unharmful quick adhesive, forming the MB sensor-embedded MBAB (EMBAB). The modified simulated body fluid (MSBF) media (PH = 7.4), mimicking the human internal environment, and the NaOH media (PH = 12), accelerating EMBAB's degradation, were used to immerse the EMBAB for 15 days at 37 °C. The EMBAB was then tested daily on a self-developed experimental platform to monitor the relative output power under a 100 N external force. The results showed that the relative output power of the sensing coil gradually increased with the EMBAB's degradation. The degradation rate of the EMBAB could be calculated on the basis of the changes of the relative output power caused by the MB sensor and of the degradation time. With the EMBAB's degradation, an increasing strain directly worked on the MB sensor, significantly changing the value of the relative output power, which means that the EMBAB was characterized by a quick degradation rate. During the 15 days of the experiment, the degradation rates on the 7th and 15th days were 0.005 dbm/day and 0.02 dbm/day, and 0.02 dbm/day and 0.04 dbm/day in MSBF and alkaline media, respectively. Therefore, the MB sensor provides a wireless and passive method to monitor and assess the degradation rate of bone implants in vitro.

Noncontact Strain Monitoring of Osseointegrated Prostheses.

The objective of this study was to develop a noncontact, noninvasive, imaging system for monitoring the strain and deformation states of osseointegrated prostheses. The proposed sensing methodology comprised of two parts. First, a passive thin film was designed such that its electrical permittivity increases in tandem with applied tensile loading and decreases while unloading. It was found that patterning the thin films could enhance their dielectric property's sensitivity to strain. The film can be deposited onto prosthesis surfaces as an external coating prior to implant. Second, an electrical capacitance tomography (ECT) measurement technique and reconstruction algorithm were implemented to capture strain-induced changes in the dielectric property of nanocomposite-coated prosthesis phantoms when subjected to different loading scenarios. The preliminary results showed that ECT, when coupled with strain-sensitive nanocomposites, could quantify the strain-induced changes in the dielectric property of thin film-coated prosthesis phantoms. The results suggested that ECT coupled with embedded thin films could serve as a new noncontact strain sensing method for scenarios when tethered strain sensors cannot be used or instrumented, especially in the case of osseointegrated prostheses.

Accuracy and reliability of a hand-held in vivo skin indentation device to assess skin elasticity.

OBJECTIVE: The aim of this study was to evaluate the performance of a hand-held indentation device for fast and reliable determination of skin stiffness. METHODS: Device accuracy to indentation depths of 0.6 and 1.3 mm was first evaluated on plastic foam materials with mechanical properties verified by a laboratory material testing device. Subsequently, the device's sensitivity to detect age-related changes in skin stiffness was evaluated among 46 healthy women (18-79 years). Finally, the reproducibility of the method was tested with six healthy subjects. RESULTS: High correlation was detected between indentation stiffness of reference material and Young's modulus determined with mechanical testing device (0.6 mm indenter: r = 0.97, P = 0.05; 1.3 mm indenter: r = 0.98, P = 0.04). Age-related decrease of 38% in skin stiffness was observed in healthy volunteers (P < 0.05). The coefficient of variation for 0.6 and 1.3 mm indenters was 7.4% and 8.5%, respectively. No trend related to hysteresis effect was observed from repeated measurements. CONCLUSIONS: The presented indentation technique was accurate against the laboratory material testing device. Furthermore, skin changes related to ageing could be detected with the indentation technique. The new device was found to be feasible for monitoring skin stiffness in cosmetics and clinical conditions.

Immunoaffinity based methods are superior to kits for purification of prostate derived extracellular vesicles from plasma samples.

BACKGROUND: The ability to isolate extracellular vesicles (EVs) such as exosomes or microparticles is an important method that is currently not standardized. While commercially available kits offer purification of EVs from biofluids, such purified EV samples will also contain non-EV entities such as soluble protein and nucleic acids that could confound subsequent experimentation. Ideally, only EVs would be isolated and no soluble protein would be present in the final EV preparation. METHODS: We compared commercially available EV isolation kits with immunoaffinity purification techniques and evaluated our final EV preparations using atomic force microscopy (AFM) and nanoscale flow cytometry (NFC). AFM is the only modality capable of detecting distinguishing soluble protein from EVs which is important for downstream proteomics approaches. NFC is the only technique capable of quantitating the proportion of target EVs to non-target EVs in the final EV preparation. RESULTS: To determine enrichment of prostate derived EVs relative to non-target MPs, anti-PSMA (Prostate Specific Membrane Antigen) antibodies were used in NFC. Antibody-based immunoaffinity purification generated the highest quality of prostate derived EV preparations due to the lack of protein and RNA present in the samples. All kits produced poor purity EV preparations that failed to deplete the sample of plasma protein. CONCLUSIONS: While attractive due to their ease of use, EV purification kits do not provide substantial improvements in isolation of EVs from biofluids such as plasma. Immunoaffinity approaches are more efficient and economical and will also eliminate a significant portion of plasma proteins which is necessary for downstream approaches.