Microfluidics‐Enabled Multimaterial Maskless Stereolithographic Bioprinting

A stereolithography‐based bioprinting platform for multimaterial fabrication of heterogeneous hydrogel constructs is presented. Dynamic patterning by a digital micromirror device, synchronized by a moving stage and a microfluidic device containing four on/off pneumatic valves, is used to create 3D constructs. The novel microfluidic device is capable of fast switching between different (cell‐loaded) hydrogel bioinks, to achieve layer‐by‐layer multimaterial bioprinting. Compared to conventional stereolithography‐based bioprinters, the system provides the unique advantage of multimaterial fabrication capability at high spatial resolution. To demonstrate the multimaterial capacity of this system, a variety of hydrogel constructs are generated, including those based on poly(ethylene glycol) diacrylate (PEGDA) and gelatin methacryloyl (GelMA). The biocompatibility of this system is validated by introducing cell‐laden GelMA into the microfluidic device and fabricating cellularized constructs. A pattern of a PEGDA frame and three different concentrations of GelMA, loaded with vascular endothelial growth factor, are further assessed for its neovascularization potential in a rat model. The proposed system provides a robust platform for bioprinting of high‐fidelity multimaterial microstructures on demand for applications in tissue engineering, regenerative medicine, and biosensing, which are otherwise not readily achievable at high speed with conventional stereolithographic biofabrication platforms.     

Cell‐Instructive Microgels with Tailor‐Made Physicochemical Properties

A microfluidic in vitro cell encapsulation platform to systematically test the effects of microenvironmental parameters on cell fate in 3D is developed. Multiple cell types including fibroblasts, embryonic stem cells, and cancer cells are incorporated in enzymatically cross‐linked poly(ethylene glycol)‐based microgels having defined and tunable mechanical and biochemical properties. Furthermore, different approaches to prevent cell “escape” from the microcapsules are explored and shown to substantially enhance the potential of this technology. Finally, coencapsulation of microgels within nondegradable gels allows cell viability, proliferation, and morphology to be studied in different microenvironmental conditions up to two weeks in culture.     

Preparation and characterization of fenofibrate-loaded PLA–PEG microspheres

A series of biodegradable block copolymer of poly(lactide)(PLA)/poly(ethylene glycol) (PEG) were prepared by Ring-Opening polymerization of D, L-lactide, using stannous octoate as a catalyst. By nanoprecipitation method, the PLA-PEG can be made into microspheres containing fenofibrate, which is a kind of important cholesterol-lowering drugs. The purpose of this study is to investigate the effect of the copolymer composition on the size, the entrapment and the release behavior of the fenofibrate loaded microspheres. The microspheres can be achieved with small size below 100 nm, better encapsulation efficiencies of more than 55.3% and slower release rates. The release of fenofibrate from microsphere would reach the balance first, when the microsphere prepared by high proportion of hydrophilic PEG block. And the release property of fenofibrate/PLA-PEG microsphere was better than Lipanthyl (a commercial capsule of fenofibrate). It was observed that the composition of PLA-PEG copolymer played a major role in encapsulation efficiency of microspheres and release rates.     

Biodegradable,Drug‐Loaded Nanovectors via Direct Hydration as a New Platform for Cancer Therapeutics

The stabilization and transport of low‐solubility drugs, by encapsulation in nanoscopic delivery vectors (nanovectors), is a key paradigm in nanomedicine. However, the problems of carrier toxicity, specificity, and producibility create a bottleneck in the development of new nanomedical technologies. Copolymeric nanoparticles are an excellent platform for nanovector engineering due to their structural versatility; however, conventional fabrication processes rely upon harmful chemicals that necessitate purification. In engineering a more robust (copolymeric) nanovector platform, it is necessary to reconsider the entire process from copolymer synthesis through self‐assembly and functionalization. To this end, a process is developed whereby biodegradable copolymers of poly(ethylene glycol)‐block‐poly(trimethylene carbonate), synthesized via organocatalyzed ring‐opening polymerization, undergo assembly into highly uniform, drug‐loaded micelles without the use of harmful solvents or the need for purification. The direct hydration methodology, employing oligo(ethylene glycol) as a nontoxic dispersant, facilitates rapid preparation of pristine, drug‐loaded nanovectors that require no further processing. This method is robust, fast, and scalable. Utilizing parthenolide, an exciting candidate for treatment of acute lymphoblastic leukemia (ALL), discrete nanovectors are generated that show strikingly low carrier toxicity and high levels of specific therapeutic efficacy against primary ALL cells (as compared to normal hematopoietic cells).     

Dual-crosslinked homogeneous alginate microspheres for mesenchymal stem cell encapsulation

A smart hydrogel material was used in combination with custom microfluidic devices (MFDs) to create microspheres for human mesenchymal stem cell (MSC) encapsulation. Methods for fabricating homogeneous stimuli-responsive microspheres for MSC encapsulation and cell delivery have gained interest to increase viability and manipulate microencapsulation within microspheres 10–1000?µm in diameter. Herein, MFDs were combined with non-toxic smart hydrogel materials to tune both the size and mechanics of the microspheres. Traditional hydrogels have a single input/stimulus for crosslinking, utilize potentially toxic ultraviolet radiation, and fail to mimic surrounding musculoskeletal tissue mechanics. Thus, it is highly beneficial to encapsulate MSCs inside a mechanically-stable microsphere made from naturally-derived materials. The objectives of this research were to optimize microsphere fabrication techniques using custom microfluidic devices (MFDs), and to encapsulate viable MSCs within visible-light crosslinked smart-alginate microspheres, with tunable mechanical properties. Microsphere production was characterized optically, and MSC viability, post-encapsulation, was verified using a standard florescence assay. Cell viability was maintained in chemically-modified alginate homogenous microspheres post encapsulation, and after subsequent crosslinking via green light exposure.

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Development and characterization of wheat gluten microspheres for use in a controlled release system

The purpose of this study was the development and characterization of wheat gluten microspheres for use as controlled release devices, and the evaluation of the effect of the addition of poly (ethylene glycol) (PEG). Diltiazem hydrochloride was used as the model drug in the in vitro release essay. The physical–chemical and morphological properties of the microspheres were evaluated, as well as their encapsulation efficiency. Porosity varied with the presence or absence of PEG. The diltiazem encapsulation efficiency was 72.8% and 96.7% for wheat gluten and gluten/PEG 95/05 microspheres, respectively. The DSC and FTIR results indicated interactions between the microparticles and additives used. In the in vitro release tests it was observed that, for all the studied systems, the burst effect occurred in the first 2 h of release and the microspheres prepared with PEG had a faster release rate. In the attempt to elucidate the release mechanism, the systems were treated based on two well known mathematical models: the Higuchi and the power law. It was found that the microsphere release mechanism is not exclusively diffusion-controlled and, probably, the release occurs through a combination of partial diffusion through the swelling matrix and hydrophilic pores.     

Mesenchymal Stem Cell Capping on ECM‐Anchored Caspase Inhibitor–Loaded PLGA Microspheres for Intraperitoneal Injection in DSS‐Induced Murine Colitis

Mesenchymal stem cells (MSCs) are considered as a promising alternative for the treatment of various inflammatory disorders. However, poor viability and engraftment of MSCs after transplantation are major hurdles in mesenchymal stem cell therapy. Extracellular matrix (ECM)‐coated scaffolds provide better cell attachment and mechanical support for MSCs after transplantation. A single‐step method for ECM functionalization on poly(lactic‐co‐glycolic acid) (PLGA) microspheres using a novel compound, dopamine‐conjugated poly(ethylene‐alt‐maleic acid), as a stabilizer during the preparation of microspheres is reported. The dopamine molecules on the surface of microspheres provide active sites for the conjugation of ECM in an aqueous solution. The results reveal that the viability of MSCs improves when they are coated over the ECM‐functionalized PLGA microspheres (eMs). In addition, the incorporation of a broad‐spectrum caspase inhibitor (IDN6556) into the eMs synergistically increases the viability of MSCs under in vitro conditions. Intraperitoneal injection of the MSC–microsphere hybrid alleviates experimental colitis in a murine model via inhibiting Th1 and Th17 differentiation of CD4⁺ T cells in colon‐draining mesenteric lymph nodes. Therefore, drug‐loaded ECM‐coated surfaces may be considered as attractive tools for improving viability, proliferation, and functionality of MSCs following transplantation.     

Soybean Lecithin‐Mediated Nanoporous PLGA Microspheres with Highly Entrapped and Controlled Released BMP‐2 as a Stem Cell Platform

Injectable polymer microsphere‐based stem cell delivery systems have a severe problem that they do not offer a desirable environment for stem cell adhesion, proliferation, and differentiation because it is difficult to entrap a large number of hydrophilic functional protein molecules into the core of hydrophobic polymer microspheres. In this work, soybean lecithin (SL) is applied to entrap hydrophilic bone morphogenic protein‐2 (BMP‐2) into nanoporous poly(lactide‐co‐glycolide) (PLGA)‐based microspheres by a two‐step method: SL/BMP‐2 complexes preparation and PLGA/SL/BMP‐2 microsphere preparation. The measurements of their physicochemical properties show that PLGA/SL/BMP‐2 microspheres had significantly higher BMP‐2 entrapment efficiency and controlled triphasic BMP‐2 release behavior compared with PLGA/BMP‐2 microspheres. Furthermore, the in vitro and in vivo stem cell behaviors on PLGA/SL/BMP‐2 microspheres are analyzed. Compared with PLGA/BMP‐2 microspheres, PLGA/SL/BMP‐2 microspheres have significantly higher in vitro and in vivo stem cell attachment, proliferation, differentiation, and matrix mineralization abilities. Therefore, injectable nanoporous PLGA/SL/BMP‐2 microspheres can be potentially used as a stem cell platform for bone tissue regeneration. In addition, SL can be potentially used to prepare hydrophilic protein‐loaded hydrophobic polymer microspheres with highly entrapped and controlled release of proteins.     

Microfluidic Generation of All‐Aqueous Double and Triple Emulsions

Higher order emulsions are used in a variety of different applications in biomedicine, biological studies, cosmetics, and the food industry. Conventional droplet generation platforms for making higher order emulsions use organic solvents as the continuous phase, which is not biocompatible and as a result, further washing steps are required to remove the toxic continuous phase. Recently, droplet generation based on aqueous two‐phase systems (ATPS) has emerged in the field of droplet microfluidics due to their intrinsic biocompatibility. Here, a platform to generate all‐aqueous double and triple emulsions by introducing pressure‐driven flows inside a microfluidic hybrid device is presented. This system uses a conventional microfluidic flow‐focusing geometry coupled with a coaxial microneedle and a glass capillary embedded in flow‐focusing junctions. The configuration of the hybrid device enables the focusing of two coaxial two‐phase streams, which helps to avoid commonly observed channel‐wetting problems. It is shown that this approach achieves the fabrication of higher‐order emulsions in a poly(dimethylsiloxane)‐based microfluidic device, and controls the structure of the all‐aqueous emulsions. This hybrid microfluidic approach allows for facile higher‐order biocompatible emulsion formation, and it is anticipated that this platform will find utility for generating biocompatible materials for various biotechnological applications.     

Multiplexed spectral signature detection for microfluidic color-coded bioparticle flow

Here, we report a high-speed photospectral detection technique capable of discriminating subtle variations of spectral signature among fluorescently labeled cells and microspheres flowing in a microfluidic channel. The key component used in our study is a strain-tunable nanoimprinted grating microdevice coupled with a photomultiplier tube (PMT). The microdevice permits acquisition of the continuous spectral profiles of multiple fluorescent emission sources at 1 kHz. Optically connected to a microfluidic flow chamber via a multimode optical fiber, our multiwavelength detection platform allows for cytometric measurement of cell groups emitting nearly identical fluorescence signals with a maximum emission wavelength difference as small as 5 nm. The same platform also allows us to demonstrate microfluidic flow cytometry of four different microsphere types in a wavelength bandwidth as narrow as 40 nm at a high (>85%) confidence level. Our study shows that detection of fluorescent spectral signatures at high speed and high resolution can expand specificity of multicolor flow cytometry. The enhanced capability enables multiplexed analysis of color-coded bioparticles based on single-laser excitation and single-detector spectroscopy in a microfluidic setting. The fluorescence signal discrimination power achieved by the optofluidic technology holds great promise to enable quantification of cellular parameters with higher accuracy as well as enumeration of a larger number of cell types than conventional flow cytometric methods.     

Evaluation of BSA protein release from hollow hydroxyapatite microspheres into PEG hydrogel

Implants that simultaneously function as an osteoconductive matrix and as a device for local drug or growth factor delivery could provide an attractive system for bone regeneration. In our previous work, we prepared hollow hydroxyapatite (abbreviated HA) microspheres with a high surface area and mesoporous shell wall and studied the release of a model protein, bovine serum albumin (BSA), from the microspheres into phosphate-buffered saline (PBS). The present work is an extension of our previous work to study the release of BSA from similar HA microspheres into a biocompatible hydrogel, poly(ethylene glycol) (PEG). BSA-loaded HA microspheres were placed in a PEG solution which was rapidly gelled using ultraviolet radiation. The BSA release rate into the PEG hydrogel, measured using a spectrophotometric method, was slower than into PBS, and it was dependent on the initial BSA loading and on the microstructure of the microsphere shell wall. A total of 35–40% of the BSA initially loaded into the microspheres was released into PEG over ~ 14 days. The results indicate that these hollow HA microspheres have promising potential as an osteoconductive device for local drug or growth factor delivery in bone regeneration and in the treatment of bone diseases.     

Hollow Carbon Nanospheres with Tunable Hierarchical Pores for Drug,Gene, and Photothermal Synergistic Treatment

Design and synthesis of porous and hollow carbon spheres have attracted considerable interest in the past decade due to their superior physicochemical properties and widespread applications. However, it is still a big challenge to achieve controllable synthesis of hollow carbon nanospheres with center‐radial large mesopores in the shells and inner surface roughness. Herein, porous hollow carbon nanospheres (PHCNs) are successfully synthesized with tunable center‐radial mesopore channels in the shells and crater‐like inner surfaces by employing dendrimer‐like mesoporous silica nanospheres (DMSNs) as hard templates. Compared with conventional mesoporous nanospheres, DMSN templates not only result in the formation of center‐radial large mesopores in the shells, but also produce a crater‐like inner surface. PHCNs can be tuned from open center‐radial mesoporous shells to relatively closed microporous shells. After functionalization with polyethyleneimine (PEI) and poly(ethylene glycol) (PEG), PHCNs not only have negligible cytotoxicity, excellent photothermal property, and high coloading capacity of 482 µg of doxorubicin and 44 µg of siRNA per mg, but can also efficiently deliver these substances into cells, thus displaying enhanced cancer cell killing capacity by triple‐combination therapy.     

Aqueous Preparation and Evaluation of Albumin‐Chitosan Microspheres Containing Indomethacin

Controlled‐release egg albumin‐chitosan microspheres containing indomethacin as a model drug were successfully prepared by coacervation method. The proposed method can offer a simple method for microsphere preparation in an aqueous system with the elimination of the use of organic solvents that are usually needed in conventional techniques of microencapsulation. The interaction between negatively charged egg albumin molecules in phosphate buffer, pH 7.2, or sodium hydroxide solution and positively charged chitosan molecules dissolved in diluted acetic acid to form an insoluble precipitate was the principle for the formation of the microspheres. The effects of many process variables, such as amount of formaldehyde as a cross‐linking agent, stirring time, final pH of encapsulation medium, initial drug loading, and albumin concentration or albumin‐to‐chitosan weight ratio, on the properties of the prepared microspheres were investigated. Incorporation efficiencies of the microspheres to the drug were high in most cases and ranged between 63.3 ± 3.6% and 92.39 ± 3.2%, while particle sizes were 435.2 ± 12.6 up to 693.9 ± 34.6 µm for the different tested batches. On the other hand, the values of angles of repose and compressibility indices were in the range of 23.5 ± 0.4 to 32.0 ± 0.7 degrees and 11.1 ± 0.7% to 23.6 ± 0.7% respectively, which indicate overall good free flowing nature of the microspheres of all batches. The maximum required amount of the cross‐linking agent was determined to avoid excessive unnecessary chemicals. It was also noticed that excessive time of stirring and excessive initial drug loading are not recommended as it may lead to microspheres of low properties. The pH of the encapsulation media (pH 3.77 up to pH 4.91) significantly affected the properties of the microspheres. As the pH of the encapsulation media was increased, the incorporation efficiency, particle size, and flowability decreased, along with increase of drug release rate, which could be related to incomplete cross linking of the microspheres matrix. It was also observed that high concentration of albumin solution and accordingly the increase of albumin‐to‐chitosan weight ratio were accompanied with increases in incorporation efficiency and particle size with improved microsphere flowability and slow indomethacin release. Thus, the proposed microspheres showed the ability to control the release of indomethacin, and their properties were highly affected by many process variables that could be controlled to obtain an optimized system.     

Polymer microspheres with active carboxyl groups: preparation, structure and hydrophilicity

A quantitative research of microspheres, poly(ethylene glycol dimethacrylate-co-acrylic acid) (P(EGDMA-co-AA)) and poly(divinvlbenzene-80-co-acrylic acid) (P(DVB-co-AA)), with active carboxyl groups on surface prepared by distillation-precipitation polymerization was presented in this paper. The loading capacity of active carboxyl group on microspheres which was investigated by titration technique would be increased and the contact angle was decreased following the increase of the feed ratio of acrylic acid (AA) monomer. This phenomenon indicated that the hydrophilicity of particles was mainly determined by the feed ration of hydrophilic AA monomer. However, when the AA fraction was at a fixed level, a slight difference of the loading capacity of carboxyl groups on microsphere surfaces with different crosslinkers existed. The microspheres with EGDMA as crosslinker had a higher loading capacity of carboxyl groups and lower contact angle than those of P(DVB-co-AA) microspheres, which indicated that P(EGDMA-co-AA) microspheres were more hydrophilic than P(DVB-co-AA) microspheres.     

A Microfluidic Strategy for Controllable Generation of Water‐in‐Water Droplets as Biocompatible Microcarriers

Droplet microfluidics has been widely applied in functional microparticles fabricating, tissue engineering, and drug screening due to its high throughput and great controllability. However, most of the current droplet microfluidics are dependent on water‐in‐oil (W/O) systems, which involve organic reagents, thus limiting their broader biological applications. In this work, a new microfluidic strategy is described for controllable and high‐throughput generation of monodispersed water‐in‐water (W/W) droplets. Solutions of polyethylene glycol and dextran are used as continuous and dispersed phases, respectively, without any organic reagents or surfactants. The size of W/W droplets can be precisely adjusted by changing the flow rate of dispersed and continuous phases and the valve switch cycle. In addition, uniform cell‐laden microgels are fabricated by introducing the alginate component and rat pancreatic islet (β‐TC6) cell suspension to the dispersed phase. The encapsulated islet cells retain high viability and the function of insulin secretion after cultivation for 7 days. The high‐throughput droplet microfluidic system with high biocompatibility is stable, controllable, and flexible, which can boost various chemical and biological applications, such as bio‐oriented microparticles synthesizing, microcarriers fabricating, tissue engineering, etc.     

Solvent‐Assisted Micromolding of Biohybrid Hydrogels to Maintain Human Hematopoietic Stem and Progenitor Cells Ex Vivo

Array‐format cell‐culture carriers providing tunable matrix cues are instrumental in current cell biology and bioengineering. A new solvent‐assisted demolding approach for the fabrication of microcavity arrays with very small feature sizes down to single‐cell level (3 µm) of very soft biohybrid glycosaminoglycan–poly(ethylene glycol) hydrogels (down to a shear modulus of 1 kPa) is reported. It is further shown that independent additional options of localized conjugation of adhesion ligand peptides, presentation of growth factors through complexation to gel‐based glycosaminoglycans, and secondary gel deposition for 3D cell embedding enable a versatile customization of the hydrogel microcavity arrays for cell culture studies. As a proof of concept, cell‐instructive hydrogel compartment arrays are used to analyze the response of human hematopoietic stem and progenitor cells to defined biomolecular and spatial cues.     

Covalently coating dextran on macroporous polyglycidyl methacrylate microsphere enabled rapid protein chromatographic separation

Protein denaturation and nonspecific adsorption on polymer media as a chromatographic support have been a problem which needs to be overcome. Macroporous poly(glycidyl methacrylate–divinylbezene) (PGMA–DVB) microspheres prepared in this study were firstly covalently coated with dextran through a three-step method. The dextran was firstly adsorbed onto the microspheres and then covalently bound to the PGMA–DVB microsphere through ether bonds which were formed by hydroxyl group reacting with epoxy group at the presence of 4-(Dimethylamino) pyridine. Finally, the coating dextran layer was crosslinked by ethylene glycol diglycidyl ether to form the continuous network coating. The coated microspheres were characterized by Fourier transform infrared spectra, scanning electron microscope, mercury porosimetry measurements, laser scanning confocal microscope, and protein adsorption experiments. Results showed that PGMA–DVB microspheres coated with dextran successfully maintained the macroporous structure and high permeability. The backpressure was only 1.69 MPa at a high flow rate of 2891 cm/h. Consequently, the hydrophilicity and biocompatibility of modified microspheres were greatly improved, and the contact angle decreased from 184° to 13°, and nonspecific adsorption of proteins was decreased to little or none. The clad dextran coating with large amounts of hydroxyl group was easily derived to be various functional groups. The derived media have great potential applications in rapid protein chromatography.     

Solution-phase surface modification in intact poly(dimethylsiloxane) microfluidic channels

An improved approach composed of an oxidation reaction in acidic H2O2 solution and a sequential silanization reaction using neat silane reagents for surface modification of poly(dimethylsiloxane) (PDMS) substrates was developed. This solution-phase approach is simple and convenient for some routine analytical applications in chemistry and biology laboratories and is designed for intact PDMS-based microfluidic devices, with no device postassembly required. Using this improved approach, two different functional groups, poly(ethylene glycol) (PEG) and amine (NH2), were introduced onto PDMS surfaces for passivation of nonspecific protein absorption and attachment of biomolecules, respectively. X-ray electron spectroscopy and temporal contact angle experiments were employed to monitor functional group transformation and dynamic characteristics of the PEG-grafted PDMS substrates; fluorescent protein solutions were introduced into the PEG-grafted PDMS microchannels to test their protein repelling characteristics. These analytical data indicate that the PEG-grafted PDMS surfaces exhibit improved short-term surface dynamics and robust long-term stability. The amino-grafted PDMS microchannels are also relatively stable and can be further activated for modifications with peptide, DNA, and protein on the surfaces of microfluidic channels. The resulting biomolecule-grafted PDMS microchannels can be utilized for cell immobilization and incubation, semiquantitative DNA hybridization, and immunoassay.     

RNA‐Capturing Microsphere Particles (R‐CAMPs) for Optimization of Functional Aptamers

RNA aptamers are useful building blocks for constructing functional nucleic acid‐based nanoarchitectures. The abilities of aptamers to recognize specific ligands have also been utilized for various biotechnological applications. Solution conditions, which can differ depending on the application, impact the affinity of the aptamers, and thus it is important to optimize the aptamers for the solution conditions to be employed. To simplify the aptamer optimization process, an efficient method that enables re‐selection of an aptamer from a partially randomized library is developed. The process relies on RNA‐capturing microsphere particles (R‐CAMPs): each particle displays different clones of identical DNA and RNA sequences. Using a fluorescence‐activated cell sorter, the R‐CAMPs that are linked to functional aptamers are sorted. It is demonstrated that after a single round of reselection, several functional aptamers, including the wild‐type, are selected from a library of 16 384 sequences. The selection using R‐CAMPs is further performed under the solution containing high concentration of ethylene glycol, suggesting applicability in various conditions to optimize an aptamer for a particular application. As any type of RNA clone can be displayed on the microspheres, the technology demonstrated here will be useful for the selection of RNAs based on diverse functions.     

Antibacterial activity of ciprofloxacin-loaded zein microsphere films

Our aim was to produce an antibiotic-emitting coating composed of zein microspheres for the prevention of bacterial infection on implanted devices. Ciprofloxacin-loaded zein microspheres were prepared using a phase separation procedure, with particle sizes between 0.5 and 2 µm. Drug encapsulation and drug loading varied with the amount of both zein and ciprofloxacin, and the highest encapsulation efficiency was 8.27% (2 mg/ml ciprofloxacin and 20 mg/ml zein; n = 3). A ciprofloxacin-loaded zein microsphere film (CF-MS film) was generated via solvent evaporation. Continuous drug release from a trypsin-degraded microsphere film was observed for up to 28 days. The liberation of ciprofloxacin from the trypsin-degraded film and the biodegradation of the microsphere film were highly correlated. Proliferation assay of the growth of human umbilical vein endothelial cells (HUVECs) by the MTT method showed that the microsphere film had no toxicity when compared with cells grown on Corning culture plates alone and plates with a zein film alone. Quantification of bacteria adhesion showed that adhesion on the microsphere film is significantly suppressed. In addition, according to the results of bacterial growth tests, ciprofloxacin-loaded microsphere films maintained antibacterial activity for more than 6 days. In contrast, a control medium containing a zein film allowed constant bacterial growth. These results indicate that CF-MS films might be useful as antibacterial films on implanted devices.