We present the first example of the analysis of long double-stranded (ds) DNA molecules by nanoparticle-filled capillary electrophoresis (NFCE). To avoid aggregation of the gold nanoparticles (GNPs) and to allow strong interactions with the DNA molecules, the gold nanoparticles were modified with poly(ethylene oxide) (PEO) via noncovalent bonding to form gold nanoparticle/polymer composites (GNPPs). The neutral GNPPs are heavy (approximately 2.0 x 10(8) g/mol for the 32-nm GNP) and thus slow the DNA molecules that they encounter during the electrophoretic process. Compared to linear polymer solutions, such as hydroxyethyl cellulose and PEO, the GNPPs provide greater efficiency and require significantly shorter times to separate long dsDNA. The separation of lambda-DNA (0.12-23.1 kbp) by NFCE at -250 V/cm was accomplished in 3 min. The ability to separate high molecular weight DNA markers (8.27-48.5 kbp) with plate numbers greater than 10(6) suggests that this novel method may hold great promise for the analysis of long-stranded DNA molecules such as chromosomes. Moreover, this method is simple and affordable when compared to those that use micro- and nanofabricated devices for separating long DNA molecules.

Polyurethane (PU) homopolymers and PU/polystyrene (PS) interpenetrating polymer networks (IPNs) were successfully synthesized changing the length of the pendant poly(ethylene oxide) (PEO) chains and the grafting density of PEO chains. All the PU/PS IPNs had the microphase-separated structures in which the PS-rich phase domains were dispersed in the matrix of the PU-rich phase. The domain size decreased a little, as the degree of grafting with PEO chains was increased. The water swelling ratio increased, and the interfacial energy decreased, as the length of the pendant PEO chains, and the grafting density of PEO chains of the PEO-grafted PU/PS IPNs were increased, since the mobile hydrophilic pendant PEO chains effectively induced and absorbed the water, when they were contacted with water. The hydrophilic and highly concentrated pendant PEO chains could easily prohibit the adhesion of the fibrinogens and the platelets on the surface, and the blood compatibility of IPNs was enhanced by increasing of grafting with PEO chains. The adsorption of the fibrinogens and the platelets was suppressed, as the length of pendant PEO chains, and the grafting density were increased.

Spray drying of complex liquids to form solid powders is important in many industrial applications. One of the challenges associated with spray drying is controlling the morphologies of the powders produced; this requires an understanding of how drying mechanics depend on the ingredients and conditions. We demonstrate that the morphology of powders produced by spray drying colloidal polystyrene (PS) suspensions can be significantly altered by changing the molecular weight of dissolved poly(ethylene oxide) (PEO). Samples containing high-molecular-weight PEO produce powders with more crumpled morphologies than those containing low-molecular-weight PEO. Observations of drying droplets suspended by a thin film of vapor suggest that this occurs because the samples with high-molecular-weight PEO buckle earlier in the drying process when the droplets are larger. Earlier buckling times are likely caused by the decreased stability, demonstrated by bulk rheology experiments, of PS particles in the presence of high-molecular-weight PEO at elevated temperatures. We present a consistent picture in which decreased particle stability hastens droplet buckling and leads to more crumpled powder morphologies; this underscores the importance of interparticle forces in determining the buckling of particle-laden droplets.

Because tissues are characterized by a well-defined three-dimensional arrangement of cells, tissue engineering scaffolds that facilitate the organization and differentiation of new tissue will have improved performance in comparison to scaffolds that only provide surfaces for cell attachment and growth. We hypothesize that instructions for cells can be incorporated into tissue engineering scaffolds by patterning the scaffold's architecture and surface chemistry. Our goals for the presented work were to collect data about cell response to three-dimensional, porous scaffolds with uniformly modified surfaces chemistries, and to demonstrate patterning of cell response by patterning surface chemistry. Our system was osteoblast response to poly(l-lactide-co-glycolide) scaffolds modified with poly(ethylene oxide) (PEO). Scaffolds were fabricated using the Three-Dimensional Printing (3DP) process which has control over scaffolds properties to a resolution of approximately 100 microm in all three dimensions. At higher PEO concentrations, adhesion, growth rates, and migration of rat osteoblasts were reduced; alkaline phosphate activity was increased, and cells were less spread and had microvilli. Patterned regions of low and high cell adhesion were demonstrated on scaffolds fabricated with 1 mm thick stripes of PEO and non-PEO regions.

Azido-functional amphiphilic macromolecules based on a biodegradable aliphatic polyester (poly-ε-caprolactone, PCL) and a bioeliminable hydrophilic poly(ethylene oxide) (PEO) block have been used in order to build micellar drug delivery systems. Such azido groups being able to react by alkyne-azide 1,3 Huisgens cycloaddition (a click reaction) have been used further in order to cross-link the micelles via redox-sensitive disulfide bridges. This reversible cross-linking allows to prevent micelle dissociation at high dilution upon injection and to trigger their dissociation in more reductive environment, such as the cytosol. Copolymers having three different architectures, i.e. able to cross-link either the core or the shell of core-shell-corona system have been used to investigate their micellization, cross-linking and cross-linking reversibility. The stealthiness of these micelles cross-linked in the hydrophobic segment has also been studied in vitro.

To date the self-assembly of ordered metal nanoparticle (NP)/block copolymer hybrid materials has been limited to NPs with core diameters (D(core)) of less than 10 nm, which represents only a very small fraction of NPs with attractive size-dependent physical properties. Here this limitation has been circumvented using amphiphilic brush block copolymers as templates for the self-assembly of ordered, periodic hybrid materials containing large NPs beyond 10 nm. Gold NPs (D(core) = 15.8 ± 1.3 nm) bearing poly(4-vinylphenol) ligands were selectively incorporated within the hydrophilic domains of a phase-separated (polynorbornene-g-polystyrene)-b-(polynorbornene-g-poly(ethylene oxide)) copolymer via hydrogen bonding between the phenol groups on gold and the PEO side chains of the brush block copolymer. Well-ordered NP arrays with an inverse cylindrical morphology were readily generated through an NP-driven order-order transition of the brush block copolymer.

OBJECTIVE

To understand the mechanism of nano-crystalline drug formation in Pluronic (i.e., poly(ethylene oxide-block-propylene oxide) triblock copolymers) based drug-polymer solid dispersions.

METHODS

Four polymers, Pluronic F127, F108, F68 and PEG 8000, which have different poly(ethylene oxide) (PEO) and poly(propylene oxide) (PPO) ratio and chain length, were co-spray dried with BMS-347070, a COX-2 inhibitor, to form 50/50 (w/w) drug-polymer solid dispersions. The solid dispersions were analyzed by powder X-ray diffraction (PXRD), modulated differential scanning calorimetry (mDSC), and hot-stage microscopy. Average size of drug crystallites in different polymers was calculated by the Scherrer equation based on peak-broadening effect in PXRD. Two other drug compounds, BMS-A and BMS-B, were also spray dried with Pluronic F127, and the solid dispersions were analyzed by PXRD and mDSC.

RESULTS

The average size of BMS-347070 crystallites in PEG 8000, F127, F108 and F68 polymers was 69, 80, 98 and 136 nm, respectively, and the degree of BMS-347070 crystallinity is the lowest in PEG 8000. Hot-stage microscopy showed that 50/50 drug-polymer dispersions crystallized in a two-step process: a portion of the polymer crystallizes first (Step 1), followed by crystallization of drug and remaining polymer (Step 2). The T (g) value of the BMS-347070/Pluronic dispersions after Step 1 (i.e., T(g1)) was measured and/or calculated to be 15-26 degrees C, and that of BMS-347070/PEG 8000 was 60 degrees C. Solid dispersions of BMS-A and BMS-B in Pluronic F127 have T(g1) of 72 and 3 degrees C, respectively; and PXRD showed BMS-A remained amorphous after approximately 3 weeks under ambient condition, while BMS-B crystallized in F127 with an average crystallite size of 143 nm.

CONCLUSIONS

The size of drug crystallites in the drug-polymer solid dispersions is independent of polymer topology, but is caused kinetically by a combined effect of nucleation rate and crystal growth rate. When drug-Pluronic solid dispersions crystallize at room temperature, that is close to the T(g1) of the systems, a fast nucleation rate and a relatively slow crystal growth rate of the drug synergistically produced small crystallite size. While the much higher T(g1) value of drug-PEG 8000 led to a slower nucleation rate and an even slower crystal growth rate at room temperature, therefore, small crystallite size and low drug crystallinity were observed. Results from BMS-A/Pluronic and BMS-B/Pluronic systems confirmed this kinetic theory.

Water-in-water emulsions were formed by mixing incompatible aqueous solutions of dextran and poly(ethylene oxide) (PEO) in the presence of latex or protein particles. It was found that particles with a radius as small as 0.1 μm become trapped at the interface between the PEO- and dextran-rich phases with interfacial tensions down to 10(-6) N/m. The particles were visualized at the interface of the emulsion droplets using confocal laser scanning microscopy (CLSM) allowing determination of the contact angle. Various degrees of coverage with particles could be observed. On densely covered droplets, the particles had a hexagonal crystalline order. At intermediate coverage, transient clustering of the particles was observed. The diffusion coefficient of the particles at the interface was determined using multiparticle tracking. Fusion of droplets was observed in all cases leading eventually to macroscopic phase separation.

We describe the synthesis of pH-responsive miktoarm star block terpolymers mu-[polystyrene][poly(ethylene oxide)][poly(2-(dimethylamino)ethyl acrylate)] (mu-SODA) using a combination of two successive living anionic polymerizations and one reversible addition-fragmentation chain-transfer polymerization. Poly[2-(dimethylamino)ethyl acrylate] (PDMAEA) is a weak polybase that is hydrophilic at low pH and hydrophobic at high pH because of the protonation of the dimethylamino functional group with decreasing pH. In addition, our results suggest that PDMAEA is immiscible with polystyrene (PS), a feature that is desirable for the formation of multicompartment micelles. Using a combination of dynamic light scattering and cryogenic transmission electron microscopy, we demonstrate that mu-SODA micelles formed in water evolve from mixed corona (PEO + PDMAEA corona; PS core) and predominantly spherical micelles to multicompartment (PEO corona; PS + PDMAEA core) micelles with increasing pH.

A range of poly(ethylene oxide)-polybutylene terephthalate (PEO-PBT) copolymers (70-30% PEO), both as coating on titanium alloy as well as bulk cylinders, was press-fit implanted in the diaphyseal femur of 16 goats. At early survival times (4 wk), a high degree of cortical bone contact was observed for bulk implants using light microscopy and this was confirmed by backscatter electron microscopy. This was attributed to the swelling behaviour of PEO-PBT copolymers. At this stage, bone contact was also revealed for coated implants, but to a lesser extent. At a later stage (12 wk), bone bonding was demonstrated both morphologically and by X-ray microanalysis, at the interface of 70:30 PEO-PBT bulk as well as 70:30 PEO-PBT-coated implants. Bone bonding increased with time (26 and 52 wk) for this PEO-PBT proportion and was also observed for 60:40 and 55:45 implants, although less frequently. For 40:60 and 30:70 PEO-PBT proportions, bone bonding was not shown. Based on these qualitative data, it was not possible to differentiate between coated and bulk implants with respect to bone bonding. This study demonstrated that the application of PEO-PBT elastomers as coatings does not alter the bone-bonding properties. It was therefore concluded that PEO-PBT coatings are beneficial over the bone-bonding but brittle ceramic coatings, due to their flexibility. In addition, the bone-bonding capacities of these PEO-PBT coatings surpass the non-bonding behaviour of currently available flexible coatings.

In this article, we have prepared hot-melt-extruded solid dispersions of bicalutamide (BL) using poly(ethylene oxide) (PEO) as a matrix platform. Prior to preparation, miscibility of PEO and BL was assessed using differential scanning calorimetry (DSC). The onset of BL melting was significantly depressed in the presence of PEO, and using Flory-Huggins (FH) theory, we identified a negative value of -3.4, confirming miscibility. Additionally, using FH lattice theory, we estimated the Gibbs free energy of mixing which was shown to be negative, passing through a minimum at a polymer fraction of 0.55. Using these data, solid dispersions at drug-to-polymer ratios of 1:10, 2:10 and 3:10 were prepared via hot-melt extrusion. Using a combination of DSC, powder X-ray diffractometry and scanning electron microscopy, amorphous dispersions of BL were confirmed at the lower two drug loadings. At the 3:10 BL to PEO ratio, crystalline BL was detected. The percent crystallinity of PEO was reduced by approximately 10% in all formulations following extrusion. The increased amorphous content within PEO following extrusion accommodated amorphous BL at drug to polymer loadings up to 2:10; however, the increased amorphous domains with PEO following extrusion were not sufficient to fully accommodate BL at drug-to-polymer ratios of 3:10.

A study is presented of the preparation of gold nanoparticles incorporated into biodegradable micelles. Poly(ethylene oxide)-b-poly(epsilon-caprolactone) (PEO-b-PCL) copolymer was synthesized by ring-opening polymerization, and the hydroxyl end group of the PCL block was modified with thioctic acid using dicyclohexyl carbodiimide as the coupling reagent. The PEO-b-PCL-thioctate ester (TE) thus obtained was used in a later step to form monolayer protected gold nanoparticles via the thioctate spacer. Gold nanoparticles stabilized with the PEO-b-PCL block (named Au/Block (x/y), where x/y is the mole feed ratio between HAuCl4 and PEO-b-PCL-TE) were prepared and analyzed. Au/Block (1/1), Au/Block (2/1), and Au/Block (3/1) nanoparticles were found to form stable dispersions in the organic solvents commonly used to dissolve the unlabeled block copolymer. The average diameter of the nanoparticles was determined by transmission electron microscopy (TEM) and found to be 6+/-2 nm. Au/Block (4/1) nanoparticle dispersions in organic solvents, on the other hand, were not stable and produced large gold clusters (50-100 nm). Cluster formation was attributed to the low grafting density of the block copolymer, which facilitates agglomeration. For Au/Block (12/1), along the same trend, only an insoluble product was isolated. Micelles in water were prepared by the slow addition of the dilute Au/Block solution in dimethylformamide into a large excess of water with vigorous stirring. Au/Block (1/1) and Au/Block (2/1) formed nanosized structures of 5-7 nm. TEM images of stained Au/Block (1/1) micelles, made in water, clearly showed the formation of core-shell structures. Au/Block (3/1) micelles, on the other hand, were not stable and large agglomerates a few microns in size were observed. The study focuses on the synthesis, characterization, and aggregation behavior of gold-loaded PEO-b-PCL block copolymer micelles, a potential system for drug delivery in conjunction with tissue and subcellular localization studies.

A new three-dimensional (3D) scaffold containing a functional drug delivery system (DDS) consisting of electrospun micro/nanofibers is proposed. In the DDS scaffold, a core-shell laminated, structured, electrospun mat of hydrophobic polycaprolactone (PCL) and hydrophilic poly(ethylene oxide) (PEO)/rhodamine-B fibers was embedded in the normal 3D PCL scaffold, which was fabricated by a melt-plotting system. Rhodamine release from the scaffold was controlled physically by the thickness change of the PCL layer, and initial burst in drug release was eliminated by an appropriate thickness of the PCL layer. This simple technique may be useful in fabricating DDS-functional scaffolds for the clinical areas not only of bone and skin regeneration, but also of other tissue regeneration areas, regardless of the degradation rate of the structural scaffold.

This article reports on the synthesis of a well-defined hydrophilic ABA triblock copolymer composed of a poly(ethylene oxide) (PEO) middle block and thermo- and pH-sensitive outer blocks and the study of sol-gel transitions of its aqueous solutions at various pH values. The doubly responsive linear triblock copolymer, poly(methoxydi(ethylene glycol) methacrylate-co-methacrylic acid)-b-PEO-b-poly(methoxydi(ethylene glycol) methacrylate-co-methacrylic acid) (P(DEGMMA-co-MAA)-b-PEO-b-P(DEGMMA-co-MAA)), was prepared by atom transfer radical polymerization of a mixture of DEGMMA and tert-butyl methacrylate with a molar ratio of 100: 5 from a difunctional PEO macroinitiator and subsequent removal of tert-butyl groups using trifluoroacetic acid. Dynamic light scattering studies showed that the critical micellization temperature (CMT) of this ABA triblock copolymer in a 0.2 wt % aqueous solution was dependent on the solution pH and can be varied in a large temperature range (>20 degrees C). To study the sol-gel transitions, a 12.0 wt % aqueous solution of the triblock copolymer with a pH of 4.89 was made; its pH value can be readily changed and well controlled by the injection of either a 1.0 M HCl or a 1.0 M KOH solution. From rheological measurements, the sol-gel transition temperature (T(sol-gel)) versus pH curve was found to closely trace the CMT versus pH curve, though there was a shift. By cycling the solution pH between 3.2 and 5.4, we showed that the T(sol-gel) at a specific pH was reproducible. Moreover, multiple sol-gel-sol transitions were realized by judiciously controlling the temperature and pH simultaneously, demonstrating the possibility of achieving on-demand sol-gel transitions by using two external stimuli. In addition, the effect of polymer concentration on T(sol-gel) at pH = 4.0 was investigated. The sol-gel transition temperature increased with the decrease of polymer concentration and the critical gelation concentration was found to be between 4 and 6 wt %.

Charging ahead: separate values for the simultaneous electronic and ionic conductivity of a conjugated polymer containing poly(3-hexylthiophene) and poly(ethylene oxide) (P3HT-PEO) were determined by using ac impedance and dc techniques. P3HT-PEO was used as binder, and transporter of electronic charge and Li(+) ions in a LiFePO(4) cathode, which was incorporated into solid-state lithium batteries.

Protein-resistant poly(ethylene glycol methyl ether acrylate-co-polyethylene glycol diacrylate) monoliths were prepared in 150 microm i.d. capillaries using novel binary porogenic solvents consisting of ethyl ether and poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) (PEO-PPO-PEO) or PPO-PEO-PPO copolymer with molecular weights (MWs) from 2700 to 5800. The effects of the MWs and concentrations of these surfactants in the porogenic solvent mixture on the pore properties of the resultant monoliths were investigated. Several of the monoliths showed improvements in protein and peptide separations over an extended MW range compared to monoliths synthesized using non-surfactant porogens (i.e., low MW organic liquids). The pore size distributions were examined using inverse size-exclusion chromatography (ISEC) of a select series of proteins and peptides covering a wide MW range. It was found that the best monolith had relatively large fractions of micropores (<2 nm, 11.9%) and mesopores in the range from 2.8 to 15.7 nm (8.5%), which are important for size-exclusion separation of peptides and proteins, respectively. The new monoliths possessed high mechanical rigidity that enabled them to withstand pressures up to approximately 4000 psi.

Injectable hydrogels with pH and temperature triggered drug release capability were synthesized based on biocompatible glycol chitosan and benzaldehyde-capped poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol) (PEO-PPO-PEO). Aqueous solutions of the above polymers formed hydrogel under physiological conditions, allowing a desirable injectability, through the formation covalent benzoic-imine bond with pH and temperature changes. Rheological characterization demonstrated that the gelation rate and the moduli of the hydrogels were able to be tuned with chemical composition as well as pH and temperature of the polymer solution. Both hydrophobic and hydrophilic drugs could be incorporated inside the hydrogel through the in situ gel forming process and undergo a controlled release by altering pH or temperature. In vivo tests proved the formation and biocompatibility of the hydrogel in rat model.

Purification of a recombinant, thermostable alpha-amylase (MJA1) from the hyperthermophile, Methanococcus jannaschii, was investigated in the ethylene oxide-propylene oxide random copolymer (PEO-PPO)/(NH(4))(2)SO(4), and poly(ethylene glycol) (PEG)/(NH(4))(2)SO(4) aqueous two-phase systems. MJA1 partitioned in the top polymer-rich phase, while the remainder of proteins partitioned in the bottom salt-rich phase. It was found that enzyme recovery of up to 90% with a purification factor of 3.31 was achieved using a single aqueous two-phase extraction step. In addition, the partition behavior of pure amyloglucosidase in polymer/salt aqueous two-phase systems was also evaluated. All of the studied enzymes partitioned unevenly in these polymer/salt systems. This work is the first reported application of thermoseparating polymer aqueous two-phase systems for the purification of extremophile enzymes.

Thermoresponsive hydrogels with efficient water-release channels were prepared by incorporating star-shaped macromolecular pore precursors, with degradable disulfide crosslinked cores and hydrophilic poly(ethylene oxide) (PEO) arms, into the gel network. The gel framework exhibiting lower critical solution temperature (LCST) behavior was synthesized by atom transfer radical polymerization (ATRP) of 2-(2-methoxyethoxy)ethyl methacrylate and ethylene glycol dimethacrylate. The incorporation of degradable star macromolecules (dSM) was facilitated by growing the gel from ATRP initiator sites contained within their cores. Following the formation of the gel, the dSM cores were degraded, yielding uniform pores lined with hydrophilic PEO chains. The effect of hydrophilic pores on thermoresponsive hydrogel performances was studied by comparing hydrogels containing hydrophilic pores with analogous hydrogels with neutral pores or with pore-free controls. Dye absorption/release experiments pointed to the suitability of newly synthesized hydrogels as controlled-release media, for example, for drug delivery. Cell culture experiments confirmed their nontoxicity and biocompatibility (cell viability >98%).

The rheological properties of aqueous solutions of poly(ethylene oxide) (PEO) of different molecular weights (1x10(5), 4x10(5), 1x10(6) and 4x10(6) g mol(-1)) and concentrations were investigated using shear viscosity and dynamic rheological measurements. It was found that the aqueous solutions of PEO do not exhibit a yield stress and that, above a critical shear rate, all PEO solutions exhibit shear-thinning behavior, well described by the Cross model, except for the solutions made by the lowest molecular weight (1x10(5) g mol(-1)) which were almost Newtonian. The parameters of the Cross model, namely the zero-shear rate viscosity and reciprocal of the time constant, allowed the determination of the critical concentrations c(*) and c(**) (respectively, the transition to semi-dilute network solution and concentrated solution). At concentrations higher than c(**) and below a critical shear rate, solutions made of PEO of high molecular weight exhibited a clearly shear-thickening behavior at very low shear rates. In addition, the dynamic tests showed that PEO solutions exhibit concentration-dependent viscoelastic properties, with a dominant viscous behavior at PEO concentrations lower than c(**) and a dominant elastic behavior at PEO concentrations greater than c(**).

Aiming at developing new reverse thermo-responsive polymers, poly(ethylene oxide)-poly(propylene oxide) multiblock copolymers were synthesized by covalently binding the two components using carbonyl chloride and diacyl chlorides as the coupling molecules. The appropriate selection of the various components allowed the generation of systems displaying much enhanced rheological properties. For example, 15 wt% aqueous solutions of an alternating poly(ether-carbonate) comprising PEOPEO/PPO ratio on the aggregate size was observed. By incorporating short aliphatic oligoesters into the backbone, prior to the chain extension stage, reverse thermal gelation-displaying biodegradable poly(ether-ester-carbonate)s, were generated.

Well-defined alpha-methoxy-omega-amino and a alpha-hydroxy-omega-amino poly(ethylene oxide)s (PEOs) were obtained after chemical modifications of alpha-hydroxy-omega-allyl PEO which was synthesized by anionic polymerization of ethylene oxide (EO) with allyl alcoholate as initiator; molecular weights of the prepolymer were controlled by the monomer/initiator ratio. Addition of methyl iodide on the hydroxy function of this prepolymer led to an alpha-methoxy-omega-allyl PEO; completion of the reaction and purity of the resulting polymer were demonstrated by 1H, 13C NMR and GPC studies. Addition reactions of 2-amino-ethanethiol hydrochloride on alpha-hydroxy-omega-allyl PEO and alpha-methoxy-omega-allyl PEO in the presence of azobisisobutyronitrile (AIBN) led to the expected homopolymers without any side reactions as shown by 1H and 13C NMR spectra.

We present a detailed study of the kinetics of crystallization for thin films of poly(ethylene oxide) (PEO). Measurements of the growth rate have been carried out using optical-microscopy techniques on films of monodisperse PEO. Films with thicknesses from 13 nm to approximately 2 microm were crystallized isothermally at temperatures approximately 20 degrees C below the melting point. A remarkable non-monotonic slowing-down of the crystal growth is observed for films with thickness less than approximately 400 nm. The changes in the growth rate from bulk-like values is significant and corresponds to a factor of 40 decrease for the thinnest films studied. The morphologies of isothermally crystallized samples are studied using atomic-force microscopy. We find that a morphology, similar to diffusion-controlled growth (dendritic growth and densely branched growth), is observed for films with h<150 nm. In addition, changes in the morphology occur for thicknesses consistent with changes in the growth rate as a function of film thickness.

Poly(ethylene oxide) grafted with 1.8 kDa branched polyethylenimine (PEO-g-PEI) copolymers with varying compositions, that is, PEO(13k)-g-10PEI, PEO(24k)-g-10PEI, and PEO(13k)-g-22PEI, were prepared and investigated for in vitro nonviral gene transfer. Gel electrophoresis assays showed that PEO(13k)-g-10PEI, PEO(24k)-g-10PEI, and PEO(13k)-g-22PEI could completely inhibit DNA migration at an N/P ratio of 4/1, 4/1, and 3/1, respectively. Dynamic light scattering (DLS) and zeta potential measurements revealed that all three graft copolymers were able to effectively condense DNA into small-sized (80-245 nm) particles with moderate positive surface charges (+7.2 ∼ +24.1 mV) at N/P ratios ranging from 5/1 to 40/1. The polyplex sizes and zeta-potentials intimately depended on PEO molecular weights and PEI graft densities. Notably, unlike 25 kDa PEI control, PEO-g-PEI polyplexes were stable against aggregation under physiological salt as well as 20% serum conditions due to the shielding effect of PEO. MTT assays in 293T cells demonstrated that PEO-g-PEI polyplexes had decreased cytotoxicity with increasing PEO molecular weights and decreasing PEI graft densities, wherein low cytotoxicities (cell viability >80%) were observed for polyplexes of PEO(13k)-g-22PEI, PEO(13k)-g-10PEI, and PEO(24k)-g-10PEI up to an N/P ratio of 20/1, 30/1, and 40/1, respectively. Interestingly, in vitro transfection results showed that PEO(13k)-g-10PEI polyplexes have the best transfection activity. For example, PEO(13k)-g-10PEI polyplexes formed at an N/P ratio of 20/1, which were essentially nontoxic (100% cell viability), displayed over 3- and 4-fold higher transfection efficiencies in 293T cells than 25 kDa PEI standard under serum-free and 10% serum conditions, respectively. Confocal laser scanning microscopy (CLSM) studies using Cy5-labeled DNA confirmed that these PEO-g-PEI copolymers could efficiently deliver DNA into the perinuclei region as well as into nuclei of 293T cells at an N/P ratio of 20/1 following 4 h transfection under 10% serum conditions. PEO-g-PEI polyplexes with superior colloidal stability, low cytotoxicity, and efficient transfection under serum conditions are highly promising for safe and efficient in vitro as well as in vivo gene transfection applications.