PLGA-PEG-amine from PolySciTech used to generate brain-penetrating nanoparticles for treatment of neural diseases

A significant problem in treating disease which affect the brain is that getting medicine into the brain tissue is very difficult. This is due to the ‘blood-brain-barrier’ which prevents medicines in the bloodstream from crossing over into the brain tissue. This is a unique feature of the brain, as other organs (kidneys, liver, lungs, etc.) readily absorb medicines from the blood stream. A simple method to overcome this barrier is to simply dose the medicine so high that even if a small portion of the drug crosses into the brain it is effective. However, this strategy does not work with medicines that have side-effects at high doses. Another method of dealing with this problem is to generate medicine-loaded nanoparticles which are specifically modified in such a way as to allow them to penetrate across the blood-brain barrier so they can deliver medicine into the brain for treatment of neural diseases. Recently, researchers working jointly at University of Southern Denmark (Denmark) and Instituto de Investigacao e Inovacao em Saude (Portugal) utilized PLGA-PEG-NH2 from PolySciTech (www.polyscitech.com) (PolyVivo AI058) to generate transferrin decorated nanoparticles for blood-brain-barrier penetration. This research holds promise for improved delivery of medicine to brain tissue for improved treatment of cancer or neural disease such as alzeheimers. Read more: Gomes, Maria Joao, Patrick J. Kennedy, Susana Martins, and Bruno Sarmento. “Delivery of siRNA silencing P-gp in peptide-functionalized nanoparticles causes efflux modulation at the blood–brain barrier.” Nanomedicine 0 (2017). http://www.futuremedicine.com/doi/abs/10.2217/nnm-2017-0023

“Aim: Explore the use of transferrin-receptor peptide-functionalized nanoparticles (NPs) targeting blood–brain barrier (BBB) as siRNA carriers to silence P-glycoprotein (P-gp). Materials & methods: Permeability experiments were assessed through a developed BBB cell-based model; P-gp mRNA expression was evaluated in vitro; rhodamine 123 permeability was assessed after cell monolayer treatment with siRNA NPs. Results: Beyond their ability to improve siRNA permeability through the BBB by twofold, 96-h post-transfection, functionalized polymeric NPs successfully reduced P-gp mRNA expression up to 52%, compared with nonfunctionalized systems. Subsequently, the permeability of rhodamine 123 through the human BBB model increased up to 27%. Conclusion: Developed BBB-targeted NPs induced P-gp downregulation and consequent increase on P-gp substrate permeability, revealing their ability to modulate drug efflux at the BBB.”

PLGA from PolySciTech used as part of development of pH responsive nanoparticles for cancer treatment

One of the fundamental problems with treatment of cancer is that the disease itself is still “part” of the human body. Cancer is simply a portion of the tissue and cells which are growing/proliferating at the wrong rate or in a manner which is damaging other tissues. For most diseases caused by an external pathogen, designing a medicinal treatment is simply a matter of finding an agent which affects the pathogen and not the patient. For example, the antibiotic penicillin prevents synthesis of cell-walls, which are key components of bacteria but not found in human cells. For this reason, penicillin can be easily administered to patients at high systemic doses with minimal concern for side effects. Unfortunately, for cancer, the situation is not so simple. Most agents which act to kill or prevent growth of cancer cells also have similar action on healthy cells, due to the fact both that the disease and the patient are of the same cell-type. For this reason, the few differences between cancer cells and normal cells that do exist are ideal targets to improve the action of therapeutics against cancer while maintaining minimal activity against normal cells. One difference between normal tissues and cancer is that, due to differences in tumor metabolism, the tumor tissues become acidic with pH ~6.5-7 (typical cellular pH is 7.4). This has led to rumors that acidity causes the tumor to grow and that cancer can be prevented, or even cured, simply by consuming pH basic (or so-called “alkaline”) foods. If this was true, then cancer could be cured by simply eating Rolaids or TUMS, which is not the case. It is the growing cancer which generates the acidic environment, not the other way around. This pH variability is one difference between normal tissue and cancerous tissues which can be used for optimizing targeted drug strategies. Recently, researchers working jointly at Purdue University, Fudan University (China), Shenyang Pharmaceutical University (China), and Eli Lilly, utilized PLGA from PolySciTech (www.polyscitech.com) (PolyVivo AP081) to create drug-loaded nanoparticles. These were surface modified to render them pH sensitive for preferential release at low pH. Although they worked well during in-vitro testing, there were problems with components of blood interacting with the coating and altering it preventing the pH effect from being fully utilized during in-vivo research. This is an important aspect of real science. Often, during development, there are setbacks to overcome which are discovered over the course of the research. This research holds promise for development of improved chemotherapeutics. Read more: Han, Ning, Jun Xu, Liang Pang, Hyesun Hyun, Jinho Park, and Yoon Yeo. “Development of surface-variable polymeric nanoparticles for drug delivery to tumors.” Molecular Pharmaceutics (2017). http://pubs.acs.org/doi/abs/10.1021/acs.molpharmaceut.7b00050

“Abstract: To develop nanoparticle drug carriers that interact with cells specifically in the mildly acidic tumor microenvironment, we produced polymeric nanoparticles modified with amidated TAT peptide via a simple surface modification method. Two types of core poly(lactic-co-glycolic acid) nanoparticles (NL and NP) were prepared with a phospholipid shell as an optional feature and covered with polydopamine that enabled the conjugation of TAT peptide on the surface. Subsequent treatment with acid anhydrides such as cis-aconitic anhydride (CA) and succinic anhydride (SA) converted amines of lysine residues in TAT peptide to β-carboxylic amides, introducing carboxylic groups that undergo pH-dependent protonation and deprotonation. The nanoparticles modified with amidated TAT peptide (NLpT-CA and NPpT-CA) avoided interactions with LS174T colon cancer cells and J774A.1 macrophages at pH 7.4 but restored the ability to interact with LS174T cells at pH 6.5, delivering paclitaxel efficiently to the cells following a brief contact time. In LS174T tumor-bearing nude mice, NPpT-CA showed less accumulation in the lung than NPpT, reflecting the shielding effect of amidation, but tumor accumulation of NPpT and NPpT-CA was equally minimal. Comparison of particle stability and protein corona formation in media containing sera from different species suggests that NPpT-CA has been activated and opsonized in mouse blood to a greater extent than those in bovine serum-containing medium, thus losing the benefits of pH-sensitivity expected from in vitro experiments. Keywords: acid anhydrides; drug delivery; pH sensitive; PLGA nanoparticles; TAT peptide”

PLGA from PolySciTech used as rapamycin eluting coating on magnesium alloy stents for restenosis prevention as part of heart-disease research

A popular treatment for cardiac blockage is angioplasty. Under this treatment, a thin catheter is run up to the affection portion of the heart and then a balloon is expanded near the tip to remove the blockage. A drawback to this technique is that, over time, the affected blood vessel re-narrows unless something is left in place, such as a stent. Over a longer period of time, the tissues of the blood vessel will regrow over the stent and re-block the vessel by a process called restenosis. A wide variety of technologies have been applied to dealing with this issue so as to provide a long-term and effective angioplasty treatment for treating coronary artery diseases which can lead to heart-attacks if the vessel.  Recently, Researchers working jointly at Purdue University, Shanghai Jiao Tong University (China), and Microport Endovascular Co. utilized PLGA from PolySciTech (www.polyscitech.com) (PolyVivo AP122) to generate a drug-loaded coating on the stent which released anti-proliferative rapamycin to prevent restenosis. They tested this coating both on conventional stainless steel surfaces as well as novel magnesium alloys. They analyzed these samples for drug release, polymer degradation, cellular response, and other parameters. They found drug release was accelerated by the magnesium alloy underlayment and that these materials showed superior anti-proliferative capacity relative to stainless steel. This research holds promise to effectively treat coronary artery disease and prevent heart-attacks by maintaining good blood flow through the blood vessels of the heart. Read more:  Shi, Yongjuan, Jia Pei, Lei Zhang, Byung Kook Lee, Yeonhee Yun, Jian Zhang, Zhonghua Li, Song Gu, Kinam Park, and Guangyin Yuan. “Understanding the effect of magnesium degradation on drug release and anti-proliferation on smooth muscle cells for magnesium-based drug eluting stents.” Corrosion Science (2017). http://www.sciencedirect.com/science/article/pii/S0010938X16314433

“Abstract: To understand the possible influence of substrate degradation on the drug-loading system of magnesium alloy-based drug-eluting stents, a rapamycin drug-loading poly(lactic-co-glycolic acid) coating was prepared on Mg-Nd-Zn-Zr stents for a systematic investigation in a phosphate buffer system. Mg degradation accelerated the drug release kinetics prominently, which was mainly attributed to H2 evolution in the diffusion-controlled phase while thereafter to PLGA erosion. Although physiochemical stability of the released rapamycin was partially deteriorated by magnesium degradation, the drug-loading system on magnesium substrates exhibited a more potent long-term inhibition on smooth muscle cell proliferation in vitro as compared to drug-loaded stainless steel. Highlights: We firstly reported that the degradation of magnesium substrate would improve the in vitro rapamycin release from drug-loading PLGA/RAPA system on a Mg-Nd-Zn-Zr alloy. We quantitatively analyzed the factors enhancing the in vitro drug release kinetics from Mg-based drug-eluting system, distinguishing that it was mainly caused by H2 evolution, while pH only played a trivial role. We reported for the first time that the Mg-based PLGA/RAPA drug-loading system exhibited more pronounced long-term inhibition for the proliferation of smooth muscle cells, under conditions that PLGA with low degradation rate was used as the drug carrier. Keywords Magnesium; Organic coatings; Polymer; Erosion; Interfaces; Kinetic parameters.”

PLGA from PolySciTech used for generating dopamine-Mn coated theranostic nanoparticles for use in cancer treatment

Chemotherapy is the primary means of treating cancer however the currently available regimens suffer from significant side-effects and related toxicity due to the non-specific nature of this approach which damages both tumors as well as normal tissues. Combination therapies have been developed as a means for dealing with this by providing for a more targeted approach to cancer treatment in which the tumor is affected by the medicine to a greater degree than healthy tissues. Recently, PLGA from PolySciTech (www.polyscitech.com) (PolyVivo cat# AP040) was utilized to generate a doxorubicin loaded nanoparticle coated with dopamine and manganese. These particles serve both as magnetic resonance contrast agent and as a photothermal-triggered delivery system. This research holds promise for improved treatment of a wide array of cancers. Read more: Xi, Juqun, Lanyue Da, Changshui Yang, Rui Chen, Lizeng Gao, Lei Fan, and Jie Han. “Mn2+-coordinated PDa@ DOX/Plga nanoparticles as a smart theranostic agent for synergistic chemo-photothermal tumor therapy.” International Journal of Nanomedicine 12 (2017): 3331. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5411169/

“Abstract: Nanoparticle drug delivery carriers, which can implement high performances of multi-functions, are of great interest, especially for improving cancer therapy. Herein, we reported a new approach to construct Mn2+-coordinated doxorubicin (DOX)-loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles as a platform for synergistic chemo-photothermal tumor therapy. DOX-loaded PLGA (DOX/PLGA) nanoparticles were first synthesized through a double emulsion-solvent evaporation method, and then modified with polydopamine (PDA) through self-polymerization of dopamine, leading to the formation of PDA@DOX/PLGA nanoparticles. Mn2+ ions were then coordinated on the surfaces of PDA@DOX/PLGA to obtain Mn2+-PDA@DOX/PLGA nanoparticles. In our system, Mn2+-PDA@DOX/PLGA nanoparticles could destroy tumors in a mouse model directly, by thermal energy deposition, and could also simulate the chemotherapy by thermal-responsive delivery of DOX to enhance tumor therapy. Furthermore, the coordination of Mn2+ could afford the high magnetic resonance (MR) imaging capability with sensitivity to temperature and pH. The results demonstrated that Mn2+-PDA@ DOX/PLGA nanoparticles had a great potential as a smart theranostic agent due to their imaging and tumor-growth-inhibition properties. Keywords: PLGA nanoparticles, polydopamine, chemo-photothermal therapy, smart theranostic agent”

Amine-endcap PLGA from PolySciTech used in the development of nanoparticle based asthma treatment

Asthma is a very common disease affecting over 300 million people across the globe and is typified by severe inflammation of respiratory passages. Recently, overexpression of a Ca2+/calmodulin-dependent protein kinase (CaMKII) has been identified as one of the pathways which leads to this inflammation in asthma patients. A peptide which acts to inhibit CaMKII has been identified however delivering high doses of this peptide specifically to the lung-tissue requires a unique delivery system. Recently, Researchers working jointly at University of Iowa, Johns Hopkins University, and Mahidol University (Thailand) utilized amine-end capped PLGA from PolySciTech (www.polyscitech.com) (PolyVivo Cat# AI063) along with chitosan to develop inhalable cationic nanoparticle to deliver this peptide to the lung-tissue. They found this particle to be effective at cell penetration and to provide for asthma treatment with minimal side-effects in a mouse model. This research holds promise for improved asthma therapy. Read more: Morris, Angie S., Sara C. Sebag, John D. Paschke, Amaraporn Wongrakpanich, Kareem Ebeid, Mark E. Anderson, Isabella M. Grumbach, and Aliasger K. Salem. “Cationic CaMKII Inhibiting Nanoparticles Prevent Allergic Asthma.” Molecular Pharmaceutics (2017). http://pubs.acs.org/doi/abs/10.1021/acs.molpharmaceut.7b00114

“Abstract: Asthma is a common lung disease affecting over 300 million people worldwide and is associated with increased reactive oxygen species (ROS), eosinophilic airway inflammation, bronchoconstriction and mucus production. Targeting of novel therapeutic agents to the lungs of patients with asthma may improve efficacy of treatments and minimize side effects. We previously demonstrated that Ca2+/calmodulin-dependent protein kinase (CaMKII) is expressed and activated in the bronchial epithelium of asthmatic patients. CaMKII inhibition in murine models of allergic asthma reduces key disease phenotypes, providing the rationale for targeted CaMKII inhibition as a potential therapeutic approach for asthma. Herein we developed a novel cationic nanoparticle (NP)-based system for delivery of the potent and specific CaMKII inhibitor peptide, CaMKIIN, to airways. CaMKIIN-loaded NPs abrogated the severity of allergic asthma in a murine model. These findings provide the basis for development of innovative, site-specific drug delivery therapies, particularly for treatment of pulmonary diseases such as asthma. Keywords: Polylactide-co-glycolide, PLGA, Nanoparticle, Chitosan, Asthma, CaMKIIN”

mPEG-PLGA from PolySciTech utilized in optimization and fine-tuning of microfluidic nanoparticle formation techniques

Polymeric nanoparticles are widely used to improve solubility of poorly soluble medicines and blood-circulation times of rapidly cleared medicines. In this way, these are often utilized to improve the efficacy of medicines by ensuring more of the medication actually reaches the location of usage rather than be cleared out of the blood-stream. There are a wide variety of ways to make nanoparticles, most of which are based around mixing the polymer from a solvent that dissolves the polymer well in with a solvent that doesn’t dissolve it at all. This is typically done in the presence of a surfactant so that the polymer solidifies into tiny spheres. The easiest of these techniques is a simple emulsion. Anyone could do this, even in a kitchen. Simply dissolve a Styrofoam cup in a small amount of acetone (paint thinner), load a household blender with soapy water and slowly drip the polystyrene cup solution into the blender full of sudsy water while it is stirring at maximum speed. After a few minutes of stirring, pass the white slurry through a coffee filter to remove any big particles and you now have a milky-looking slurry of polystyrene nanoparticles. I do not actually suggest doing this because: 1) acetone is flammable, 2) there is no practical application for generating nanoparticles in your kitchen, 3) the next time you go to make milk-shakes, they may taste terrible, and 4) it will certainly void the warranty on your house-hold blender (just because you can, doesn’t mean you should try this at home). Nanoparticles made by this type of emulsion technique come out in a wide range of different sizes, because the processes which drive their formation are random. However, microfluidics is a newer technique in which the mixing is precisely controlled so that all the nanoparticles are generated at the same size and in a highly controlled manner. Defining exactly how the blending of the polymer solution with the non-solvent occurs is a process which requires a great deal of experimentation and fluid mechanics to elucidate the precise parameters (concentrations, mixing speeds, etc.) that allow for predetermined sizes of polymer nanoparticles to be made. Recently, Researchers at The University of Queensland (Australia) utilized mPEG-PLGA from PolySciTech (www.polyscitech.com) of two different block sizes (PolyVivo AK026 (5k-55K) and AK037 (5K-20K)) to investigate microfluidic mechanisms for producing monodisperse nanoparticles with extremely well controlled sizes.  For this, they dissolved the polymers into acetonitrile and then processed them through an advanced microfluidics system to generate precisely sized nanoparticles. This research holds promise for the generation of well-controlled nanoparticles to encapsulate medicines and improve their efficacy. Read more: Baby, Thejus, Yun Liu, Anton PJ Middelberg, and Chun-Xia Zhao. “Fundamental studies on throughput capacities of hydrodynamic flow-focusing microfluidics for producing monodisperse polymer nanoparticles.” Chemical Engineering Science (2017). http://www.sciencedirect.com/science/article/pii/S0009250917302993

“Abstract: Microfluidics enables the manipulation of liquids at the picoliter (or less) scale and proves to be superior over conventional bulk methods for mixing and reaction. The ability of microfluidic systems to rapidly mix reagents to provide homogeneous reaction environments, to vary the reaction conditions continuously, and to even allow reagent addition during the progress of a reaction, makes it attractive for nanoparticle synthesis. However, the low production rate limits its practical applications. Different approaches have been developed to achieve higher yield but most of them rely on the design of complex devices. Herein, we investigated fundamentally the throughput capacities of hydrodynamic flow-focusing microfluidics for producing poly (lactide-co-glycolide)-b-polyethylene glycol (PLGA-PEG) nanoparticles with uniform size ranging from 50-150 nm. The effects of different factors of microfluidic design, including channel width, channel depth, channel structure and flow rate ratios, on particle size, size distribution, and production throughput were studied and compared. In contrast to the widely used microfluidic device which has a production rate of 1.8 mg/h, our simple approach is capable of increasing the production rate of nanoparticles by more than two orders of magnitude up to 288 mg/h using a single simple device. This study demonstrated the potentials of using simple 2D microfluidic devices for a large scale production of polymeric nanoparticles that could eliminate the need for designing and fabricating complex microfluidic devices. Keywords: Microfluidics; 2D hydrodynamic flow focusing; PLGA-PEG NPs; mixing; nanoprecipitation. Highlights: A single hydrodynamic flow focusing (HFF) microfluidic device for production of polymeric nanoparticles at hundred milligram per hour scale. Tunable properties of the synthesized nanoparticles. Precise control over the size and size distribution of the synthesized nanoparticles. A library of polymer nanoparticles with systematically varied size.”

mPEG-PCL, mPEG-PLA, and mPEG-PLGA from PolySciTech used in design of theranostic stealth nanocarriers as part of drug-delivery research

One promising area of research in cancer therapy is the development of theranostics. This area of research focuses on simultaneous application of both a therapeutic agent (typically a chemotherapeutic agent such as paclitaxel) and a diagnostic agent (typically a contrast agent or fluorescent dye which renders the tumor ‘visible’). This research requires highly advanced delivery systems which can ensure that the tumor receives a suitable quantity of both agents such that it becomes visible to a surgeon as well as receives an effective dose of the therapeutic agent. In a fundamental sense, this requires well-designed nanocarriers with high loading efficiency (large doses of each agent) and which are highly stable in the bloodstream. Recently, researchers at Wroclaw University (Poland) utilized a series of PolySciTech (www.polyscitech.com) polymers including mPEG-PCL (PolyVivo Cat# AK128), mPEG-PLA (PolyVivo Cat# AK056), and mPEG-PLGA (PolyVivo Cat# AK037) to systematically generate a series of test-loaded nanoparticles containing model DNA and fluorescent dye Thiazole Orange. The researchers systematically investigated all steps involved in nanoparticle formation and tested the particles for their stability, loading capacity, and other parameters relevant to their clinical usage. This research holds promise for the development of highly advanced nanocarriers to assist in theranostic treatments of a wide variety of cancers. Read more:  Bazylińska, Urszula. “Rationally designed double emulsion process for co-encapsulation of hybrid cargo in stealth nanocarriers.” Colloids and Surfaces A: Physicochemical and Engineering Aspects (2017). http://www.sciencedirect.com/science/article/pii/S092777571730359X

“Abstract: Double emulsion process has become highly promising for development of PEG-ylated nanocarriers (NCs) with co-encapsulated hybrid model agents, i.e, hydrophilic deoxyribonucleic acid (DNA) and hydrophobic Thiazole Orange (TO) dye, in the double compartment structure to protect them from the environmental conditions and to investigate different parameters affecting the size, charge and morphology as well as colloidal and biological stability of the final theranostic nanosystems. Different stabilizing agents including surfactants: Cremophor A25, Cremophor RH 40, Poloxamer 407, di-C12DMAB as well as polymers: PEG-PDLLA, PEG-PLGA, PEG-PCL, were screened to choose suitable ones for this approach. The average size of the synthesized NCs measured by dynamic light scattering (DLS) remained < 200 nm. The encapsulation efficiency of the hybrid cargo was confirmed by UV-Vis spectroscopy. Morphology and shape of the loaded nanocontainers were investigated by transmission electron microscopy (TEM) and atomic force microscopy (AFM). Time-depended colloidal stability studies with DLS and ζ-potential followed by turbidimetric technique allow to select only the long-term nanosystems to final investigation the “stealth” properties of the fabricated PEGylated NCs. Highlights: Double emulsion process has become easy-scalable synthetic approach to develop “stealth” nanocarriers (NCs) successful in DNA and TO co-encapsulation. PEG-PDLLA, PEG-PLGA, PEG-PCL acted as pre-approved biocompatible components of the NCs polymer shell.The optimized encapsulation process resulted in NCs with diameter < 200 nm, narrow size distribution and nearly neutral surface. DLS, ζ-potential and backscattering studies confirmed a long-term NCs stability, indicating their potential as theranostic biocompatible agents. The biological stability exposed the PEG-ylated NCs ability to overcome various specific barriers to efficient drug and gene delivery. Keywords: w/o/w emulsions; PEG-ylated polyesters; DNA; Thiazole Orange; colloidal stability. Fabrication method: Polymeric nanocarriers stabilized by PEG-PLGA, PEG-PCL, PEG-PDLLA and non-ionic or cationic surfactants for co-encapsulation of therapeutic (model DNA in the initial concentration of 0.1 mg/ml) and diagnostic agent (TO in the initial concentration of 0.2 mg/ml) were prepared by modified double emulsion (w/o/w) evaporation process without any pH adjustment [8]. Generally, aqueous internal phase (with DNA) was emulsified for 5 min in dichloromethane (containing TO, PEG-ylated polymer in concentration of 5 mg/ml and di-C12DMAB) in the ratio 1:4 using a homogenizer with 25,000 rpm. This primary nanoemulsion was poured into 1% hydrophilic surfactant solution (Cremophor A25, Cremophor RH 40 or Poloxamer 407) aqueous solution stirring in a homogenizer for 10 min (25,000 rpm) and immersed in an ice water bath to create the water-in-oil-in-water (w/o/w) emulsion. The organic solvent was then evaporated under reduced pressure in a rotary evaporator (Ika RV 10 digital) and polymeric nanocarriers loaded by the hybrid cargo were collected overnight.”

mPEG-PLA from PolySciTech used by Yale University in development of a novel blood-circulation assay method

A fundamental difficulty with medicinal applications to humans is that the circulatory systems of most living organisms are designed specifically to screen out any perceived toxins or ‘non-self’ components. Typically, the kidneys and the liver work together along with macrophages (white blood cells) to remove any chemicals or particulates from the bloodstream. Although this system provides protection to the human body from toxic ingestion, it creates great difficulty for applying medicines as it greatly reduces the blood circulation time of medicinal molecules. For general medicinal applications, the loss of drug from the bloodstream is calculated as the circulation half-life and dosing schedules are calculated to match. One method to improve blood-circulation is to encapsulate the drug molecule inside of PEG-PLA so-called ‘stealth’ nanoparticles. For these particles, the PEG external coating prevents attacks by macrophages while the size alone reduces uptake and clearance by kidneys or liver. These particles enhance the blood circulation time of medicines, but a key question is by exactly how much is the blood circulation time enhanced and what is the new circulation half-life. This question critical for practical applications as it would define the dosing schedule of the encapsulated drug as it must be dosed often enough to maintain effect but not too often so as to potentially have toxic side-effects. Recently, researchers at Yale University utilized mPEG-PLA from PolySciTech (www.polyscitech.com) (PolyVivo Cat# AK054) to generate stealth-nanoparticles as test substrates for their novel fluorescence microscopy-based technique for determining half-life of particles using as little as 2 uL of blood. This research holds promise for rapid and routine determination of half-life using very small samples of blood. Read more: Tietjen, Gregory T., Jenna DiRito, Jordan S. Pober, and W. Mark Saltzman. “Quantitative microscopy-based measurements of circulating nanoparticle concentration using microliter blood volumes.” Nanomedicine: Nanotechnology, Biology and Medicine (2017). http://www.sciencedirect.com/science/article/pii/S1549963417300643

“Abstract: Nanoparticles (NPs) are potential drug delivery vehicles for treatment of a broad range of diseases. Intravenous (IV) administration, the most common form of delivery, is relatively non-invasive and provides (in theory) access throughout the circulatory system. However, in practice, many IV injected NPs are quickly eliminated by specialized phagocytes in the liver and spleen. Consequently, new materials have been developed with the capacity to significantly extend the circulating half-life of IV administered NPs. Unfortunately, current procedures for measuring circulation half-lives are often expensive, time consuming, and can require large blood volumes that are not compatible with mouse models of disease. Here we describe a simple and reliable procedure for measuring circulation half-life utilizing quantitative microscopy. This method requires only 2 μL of blood and minimal sample preparation, yet provides robust quantitative results. Graphical Abstract: Quantitative microscopy can be used to measure circulating concentrations of nanoparticles with as little as 2 μL of blood. However, when using such small volumes, the path length within and between samples can vary significantly as the high viscosity of blood can yield differences in think layer thickness as the blood spreads following application of a coverslip. This yields variability in the measured mean fluorescence intensity. Addition of a reference nanoparticle of a different color can correct the mean fluorescence intensity variance. Thus, quantitative microscopy can serve as a robust method for measuring nanoparticle half-life using μL volumes of blood. NP formulation: NP were prepared by a standard nanoprecipitation procedure. PLA–PEG (PolyVivo AK054) was dissolved at an initial concentration of 100 mg/mL in DMSO and then diluted to the desired concentration for NP formulation (typically ~55 mg/mL for the ~165 nm NPs used in this study) along with addition of either DiI or DiO dye also dissolved in DMSO. NPs were loaded with DiI or DiO dye at a final wt dye/wt polymer ratio of 0.5%. The dye/polymer solution in DMSO was added drop wise to vigorously stirring sterile diH2O in batches of 200 μL polymer/dye solution added to 1.3 mL of diH2O with identical repetitions performed to generate a full NP batch. NP were subsequently filtered through a 1.2 μm cellulose acetate membrane (GE Healthcare Life Sciences – Whatman) filter to remove any free dye or polymer aggregates and then pooled. Typically, 8 small batches of ~11 mg polymer each were combined for a total pooled batch size of ~88 mg initial polymer weight. The pooled NP solutions were then transferred to a 12 mL volume 10,000 MWCO dialysis cassettes (Thermo Scientific – Slide-A-Lyzer) and dialyzed against 2× exchanges of ~2.2 L of diH2O at room temperature to remove excess DMSO. Following dialysis NPs were aliquoted and snap frozen in liquid N2. One aliquot from each NP batch was lyophilized in a pre-weighed tube in order to determine the NP concentration. Standard NP concentration was typically ~5 mg/mL. NP batches were diluted to ~0.1 mg/mL and analyzed via dynamic light scattering (DLS) to confirm NP size and homogeneity.”