High Molecular Weight Polyacrylic Acid Research Articles

Effect of Different Molecular Weights of Polyacrylic Acid on Rat Lung Following Intratracheal Instillation.

Background: We conducted intratracheal instillations of different molecular weights of polyacrylic acid (PAA) into rats in order to examine what kinds of physicochemical characteristics of acrylic acid-based polymer affect responses in the lung. Methods: F344 rats were intratracheally exposed to a high molecular weight (HMW) of 598 thousand g/mol or a low molecular weight (LMW) of 30.9 thousand g/mol PAA at low and high doses. Rats were sacrificed at 3 days, 1 week, 1 month, 3 months and 6 months post exposure. Results: HMW PAA caused persistent increases in neutrophil influx, cytokine-induced neutrophil chemoattractants (CINC) in the bronchoalveolar lavage fluid (BALF), and heme oxygenase-1 (HO-1) in the lung tissue from 3 days to 3 months and 6 months following instillation. On the other hand, LMW PAA caused only transient increases in neutrophil influx, CINC in BALF, and HO-1 in the lung tissue from 3 days to up to 1 week or 1 month following instillation. Histopathological findings of the lungs demonstrated that the extensive inflammation and fibrotic changes caused by the HMW PAA was greater than that in exposure to the LMW PAA during the observation period. Conclusion: HMW PAA induced persistence of lung disorder, suggesting that molecular weight is a physicochemical characteristic of PAA-induced lung disorder.

Upregulation of mitochondrial dynamics is responsible for osteogenic differentiation of mesenchymal stem cells cultured on self-mineralized collagen membranes

Collagen membranes crosslinked with high molecular weight polyacrylic acid (HPAA) are capable of self-mineralization via in situ intrafibrillar mineralization. These HPAA-crosslinked collagen membranes (HCM) have been shown to promote osteogenic differentiation of mesenchymal stem cells (MSCs) and enhance bone regeneration in vivo. Nevertheless, the biological triggers involved in those processes and the associated mechanisms are not known. Here, we identified the contribution of mitochondrial dynamics in HCM-mediated osteogenic differentiation of MSCs. Mitochondriogenesis markers were significantly upregulated when MSCs were cultured on HCM, committing the MSCs to osteogenic differentiation. The mitochondria fused to form an interconnected mitochondrial network in response to the high energy requirements. Mitochondrial fission in MSCs was also triggered by HCM; fission slightly declined at 14 days to restore the equilibrium in mitochondrial dynamics. Mitophagy, another event that regulates mitochondrial dynamics, occurred actively to remove dysfunctioned mitochondria and isolate damaged mitochondria from the rest of network. The mitophagy level of MSCs was significantly elevated in the presence of HCM. Taken together, the present findings indicate that upregulation of mitochondrial dynamics via mitochondriogenesis, fusion, fission and mitophagy is responsible for HCM-mediated osteogenic differentiation of MSCs. Statement of significanceHigh molecular weight polyacrylic acid (HPAA)-crosslinked collagen membrane (HCM) was found to promote in-situ bone regeneration because of it can stimulate osteogenic differentiation of mesenchymal stem cells (MSCs). Nevertheless, the biological triggers involved in those processes and associated mechanisms are not known. This study identifies that activation of mitochondrial dynamics is centrally involved in HCM-mediated osteogenic differentiation of MSCs. The HCM accelerates mitochondriogenesis and regulates homeostasis of the mitochondrial network in response to the increased energy demand for osteogenic differentiation. Concomitantly, mitophagy actively occurs to remove dysfunctioned mitochondria from the rest of the mitochondrial network. Identification of the involvement of mitophagy in HCM-mediated osteogenic differentiation of MSCs opens new vistas in the application of biomimetic mineralization in bone tissue regeneration.

Matrix stiffening by self-mineralizable guided bone regeneration

Collagen membranes produced in vitro with different degrees of intrafibrillar mineralization are potentially useful for guided bone regeneration (GBR). However, highly-mineralized collagen membranes are brittle and difficult for clinical manipulation. The present study aimed at developing an intrafibrillar self-mineralization strategy for GBR membrane by covalently conjugating high-molecular weight polyacrylic acid (HPAA) on Bio-Gide® membranes (BG). The properties of the self-mineralizable membranes (HBG) and their potential to induce bone regeneration were investigated. The HBG underwent the progressive intrafibrillar mineralization as well as the increase in stiffness after immersed in supersaturated calcium phosphate solution, osteogenic medium, or after being implanted into a murine calvarial bone defect. The HBG promoted in-situ bone regeneration via stimulating osteogenic differentiation of mesenchymal stromal cells (MSCs). Hippo signaling was inhibited when MSCs were cultured on the self-mineralized HBG, and in HBG-promoted MSC osteogenesis during in-situ bone regeneration. This resulted in translocation of the transcription co-activators Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ) into the nucleus to induce transcription of genes promoting osteogenic differentiation of MSCs. Taken together, these findings indicated that HBG possessed the ability to self-mineralize in situ via intrafibrillar mineralization. The increase in stiffness of the extracellular matrix expedited in-situ bone regeneration by inactivating the Hippo-YAP/TAZ signaling cascade. Statement of significanceGuided bone regeneration (GBR) membranes made of naturally derived collagen have been widely used in the bone defect restoration. However, application of collagen GBR membranes run into the bottleneck with the challenges like insufficient stress strength, relatively poor dimensional stability and unsatisfactory osteoinductivity. This study develops a modified GBR membrane that can undergo progressive self-mineralization and matrix stiffening in situ. Increase in extracellular matrix stiffness provides the mechanical cues required for MSCs differentiation and expedites in-situ bone regeneration by inactivating the Hippo-YAP/TAZ signaling cascade.

Enabling Aqueous Processing for Low/No-Cobalt Lithium-Ion Battery Cathode Active Materials

Industrially, lithium-ion battery cathodes are manufactured through a wet slurry mixing and coating process. There is great interest in replacing the presently-used N-methyl-2-pyrrolidone (NMP) solvent with water due to the cost and toxicity associated with NMP. In this study, the modifications made to the active materials as a result of water exposure were studied and two materials, LiMn2O4 (LMO) and LiNi0.8Co0.15Al0.05O2 (NCA), were assembled in high-performing aqueous-processed cathodes for the first time.Five cathode active materials were the subject of this study: LiCoO2 (LCO), LiFePO4 (LFP), LMO, NCA, and LiNi0.5Mn0.3Co0.2O2 (NMC532). All materials exposed to DI water exhibited rapid increases in pH over the first 10 min, with stable pH values ranging from 9.0 (LFP and LMO) to 12.5 (NCA). Given that the protective aluminum oxide layer on the current collector breaks down at a pH of 8.5, corrosion is predicted for all materials, with NCA being the most extreme case. Pristine samples of the same five materials were submerged in water for 24 h, then the moisture was dried in a vacuum oven. The crystal structures of these materials (henceforth referred to as the wetted samples) were evaluated using X-ray diffraction and compared with pristine samples. No changes were observed in the crystal structure of any material as confirmed by Rietveld refinement. X-ray photoelectron spectroscopy was used to evaluate the changes of the surface composition of each wetted and pristine material.Based on inductively coupled plasma-mass spectrometry results, the amount of Li and transition metal leaching in water was found to be independent of time in excess of 1 h, and the amount of Li leaching in water for NCA was found to be five times that of NMC532 and more than 30 times that of the other materials. As a result, using a traditional aqueous slurry formulation (active material, carbon black, emulsion binder, carboxymethylcellulose thickener) resulted in a severely cracked and unusable NCA cathode. By including high-molecular weight polyacrylic acid (PAA), the adhesion of the cathode was restored. Half coin cells of this PAA-processed and an NMP-processed NCA cathode were constructed using punches with 1.7 mAh·cm-2 areal capacity and a charge and discharge rate of C/3 between 3.0-4.3 V. The initial capacity of the PAA-processed NCA cathode was slightly lower (176.3 mAh·g-1 to 185.2 mAh·g-1), though it did exhibit better capacity retention through 100 cycles (84.2% to 78.1%).

Influence of molecular weight on polyacrylic acid flocculation of sub-micron titanium dioxide

This paper reports the effect of the molecular weight of polyacrylic acid (PAA) on the flocculation of sub-micron titanium dioxide particles. The average molecular weight (MW) of the three studied PAAs were 140,000, 512,000 and 1,483,000 g/mol, respectively. The flocculation process was studied by several techniques including focused beam reflectance measurement (FBRM), photometric dispersion analyses (PDA), settling tests in graduated cylinders. The adsorption density of the PAAs on the titanium dioxide surface was measured by an elemental analyzer, and the floc structure was studied by scanning electron microscope and fractal dimension analysis using an optical microscope. The results showed that the flocs formed with the high MW PAA had larger sizes than the flocs generated with medium MW PAA. However, the medium MW PAA resulted in better flocculation performance, with faster settling rate and clearer supernatant. This observation was in contrast to the general perception in the literature that high MW polymer flocculants have better flocculation performance. The porosity of the flocs generated by the high MW PAA was larger than flocs by the medium MW PAA, leading to lower density flocs. This was found to be the main reason for the slow settling rate of the high MW PAA despite the larger floc size. Finally, despite the different settling rates, the PAAs with different MWs all showed their optimum flocculation performance at about 3.3 % of a monolayer coverage on the TiO2 surfaces.

Lithium and transition metal dissolution due to aqueous processing in lithium-ion battery cathode active materials

Enabling aqueous processing for lithium-ion battery cathodes is essential as solvents like N-methyl-2-pyrrolidone (NMP) are expensive, hazardous, and being phased out of usage around the world. Using water as a solvent can reduce electrode manufacturing cost and environmental impact, but it presents unique challenges for cathodes such as Li and transition metal dissolution from the active material and current collector corrosion. In this study, the suitability of aqueous processing for five cathode active materials is evaluated: LiCoO2 (LCO), LiFePO4 (LFP), LiMn2O4 (LMO), LiNi0.80Co0.15Al0.05O2 (NCA), and LiNi0.5Mn0.3Co0.2O2 (NMC532). After three days of water exposure, NCA and NMC532 exhibit significantly greater pH values (11.5–12.5) than the Ni-free materials (9.0–10.5), though all pH values suggest corrosion of the Al substrate would occur. Surface compositions change to various extent while little change is observed in the crystal structures. The transition metal dissolution in water and electrolyte is relatively low for all materials, though the Li dissolution in water is high for NCA (~0.1 mg mL−1). Electrochemical testing in half coin cells reveals that high-molecular weight polyacrylic acid addition is able to modify the pH and provide adequate binding to the current collector to permit aqueous processing of NCA.

Recyclable nanoscale zerovalent iron (nZVI)-immobilized electrospun nanofiber composites with improved mechanical strength for groundwater remediation

Nanoscale zero-valent iron (nZVI), as a promising material, has been widely used in groundwater remediation. Membrane-supported nZVI can both avoid nZVI agglomeration for better reactivity and recycle nZVI/contaminants to lower the risk of secondary pollution. However, membrane mechanical strength is a critical property for long-term operation and the regeneration of nZVI membranes. This study tried to use a high molecular weight dual-crosslinking method to improve the mechanical strength of polymeric electrospun nanofiber membranes. Specifically, high molecular weight polyacrylic acid (PAA, Mw = 450,000) was dual-crosslinked by adding polyvinyl alcohol (PVA) as covalent cross-linker (named as M450k) and Fe(II) or Fe(III) as the ionic cross-linker (named as M450k-II and M450k-III). The results indicated that the M450k had better thermal resistance against membrane shrinkage, thus having larger surface areas and more –COOH groups to immobilize more nZVI particles. Besides, M450k-III had the highest tensile strength at 8.5 MPa, 5 times the figure for the mono-crosslinked low molecular weight membrane (M2k). In terms of nZVI immobilization and filtration performance, the Fe(II)-crosslinked membrane had better nZVI immobilization with the highest removal capacity at 463 mg/g while Fe(III)-crosslinked membrane had overwhelming mechanical strength with decent and stable removal capacity under multiple nZVI regenerations over 15 filtration cycles. Generally, the high molecular weight Fe(III)-crosslinked PAA-PVA electrospun nZVI showed better potential for long-term filtration process.

White/blue-emitting, water-dispersible CdSe quantum dots prepared by counter ion-induced polymer collapse

The synthesis and characterization of water-dispersible, luminescent CdSe/ZnS semiconductor quantum dots that exhibit nominal “white” fluorescence emission and have potential applications in solid-state lighting is described. The nanomaterials, prepared through counter ion-induced collapse and UV cross-linking of high-molecular weight polyacrylic acid in the presence of appropriate aqueous inorganic ions, were of ∼2–3nm diameter and could be prepared in gram quantities. The quantum dots exhibited strong luminescence emission in two bands, the first in the blue-region (band edge) of the optical spectrum and the second, a broad emission in the red-region (attributed to deep trap states) of the optical spectrum. Because of the relative strength of emission of the band edge and deep trap state luminescence, it was possible to achieve visible white luminescence from the quantum dots in aqueous solution and in dried, solid films. The optical spectroscopic properties of the nanomaterials, including ensemble and single-molecule spectroscopy, was performed, with results compared to other white-emitting quantum dot systems described previously in the literature.

Precipitation of biomimetic fluorhydroxyapatite/polyacrylic acid nanostructures

Ordered structures of fluorhydroxyapatite (FHA) nanoparticles that resemble the nanostructure of natural human enamel have been prepared. Wet precipitation in the presence of polyacrylic acid (PAA) was used, and the particle morphology was altered by varying several reaction conditions. High molecular weight PAA increased particle length from around 54nm to several hundred nanometres, while maintaining particle width at 15nm. PAA concentration and the order of mixing the reactants also influenced crystal morphology. Optimum conditions produced dense, aligned bundles of highly elongated nanorods, which are very similar to the hierarchical nanostructure of human tooth enamel.

Low Cost, Scalable and Selective Electrochemical TSV Fill Technology for 3D IC Interconnects

Physical and economical limitations for 2D scaling (so called “Moore’s Law”) prevents further increase of integration density to improve the performance of integrated circuits (IC). These challenges have stimulated the development of 3D through-silicon via (TSV) technology (so called “More than Moore”) in order to increase the speed and the bandwidth of the devices as well as to decrease the form factor and power consumption of integrated microsystems. However, the implementation of 3D TSV technology is limited by the high cost of 3D TSV fill (due to the use of expensive vapor deposition and chemical-mechanical processes) and its poor scalability (due to low conformality of physical and chemical vapor deposited films) to high TSV aspect ratios and smaller via sizes (to increase I/O). The objective of this investigation is to address the cost and scalability issues in order to enable cost-effective and reliable fill of high aspect ratio 3D TSVs. In this paper, we will review the low temperature and scalable process technology for 3D TSV fill employing the selective electrochemical through-via metal (Ni alloy/Cu or Co alloy/Cu) filling technology which is more economical because it will eliminate expensive chemical-mechanical polishing (CMP) process and replace expensive plasma-enhanced chemical vapor deposition (PE CVD) and physical vapor deposition (PVD) processes with low cost, lower temperature (<100 oC), more scalable (AR > 30:1) selective electrochemical process (LoCoSS process). LoCoSS process technology includes the following major electrochemical process steps: anodic oxidation to form isolation layer, selective electroless barrier deposition and selective electrochemical fill that can be implemented using one LoCoSS electrochemical processing tool instead of four POR processing tools utilizing different technologies – PE CVD isolation, PVD barrier/seed, Electrofill and CMP.The activation of thin continuous electroless barrier films (Ni or Co alloys) with high adhesion on the isolation surface (SiO2) is challenging. To overcome this technical risk, a novel nanoactivation process has been developed with the use of self-assembled monolayers [Si-(CH2)x-NH2-OH] and catalytic functionalized nanoparticles (Pd/Cu). To increase the adhesion of the deposited barrier films, the covalent bonding between substrate and the SAM binding layer has been engineered through the proper choice of the functional groups. Selective activation has been achieved through the mechanical removal of catalysts from the front surface (except inside the TSV’s) and the use of photosensitive TiO2/catalyst process. Besides, robust and defect-free fill of high aspect ratio TSV’s requires the development of electroless Cu superfill process with the use of suppressing additives in the plating solution such as high molecular weight polyacrylic acid (PAA) and polyethylene glycols (PEG). Selective electroless Cu plating into high aspect ratio TSV was successfully demonstrated with complete coverage of electroless barrier layer and low overplating on the field. Copper electroplating from NANO3D suppression-based plating solutions was also used to enable high speed and uniform plating of flat Cu bumps that can be used to interconnect chips in 3D microelectronics systems. LoCoSS technology could accelerate the mass-scale adoption of low cost and scalable TSV technology in semiconductor manufacturing. Implementation of our proprietary LoCoSS process will lower the cost of 3D ICs with the potential to reduce the cost by over 2X and increase scalability by over 3X for volume production. Besides, enabling low cost 3D ICs with our technology will allow heterogeneous system generation for next generation smart phones and other communication devices as well as increase of the bandwidth, memory capacity and decrease of the signal transmission delay within computing chips.

Graphene/silicon nanocomposite anode with enhanced electrochemical stability for lithium-ion battery applications

A graphene/silicon nanocomposite has been synthesized, characterized and tested as anode active material for lithium-ion batteries. A morphologically stable composite has been obtained by dispersing silicon nanoparticles in graphene oxide, previously functionalized with low-molecular weight polyacrylic acid, in eco-friendly, low-cost solvent such as ethylene glycol. The use of functionalized graphene oxide as substrate for the dispersion avoids the aggregation of silicon particles during the synthesis and decreases the detrimental effect of graphene layers re-stacking. Microwave irradiation of the suspension, inducing reduction of graphene oxide, and the following thermal annealing of the solid powder obtained by filtration, yield a graphene/silicon composite material with optimized morphology and properties.Composite anodes, prepared with high-molecular weight polyacrylic acid as green binder, exhibited high and stable reversible capacity values, of the order of 1000 mAh g−1, when cycled using vinylene carbonate as electrolyte additive. After 100 cycles at a current of 500 mA g−1, the anode showed a discharge capacity retention of about 80%. The mechanism of reversible lithium uptake is described in terms of Li–Si alloying/dealloying reaction. Comparison of the impedance responses of cells tested in electrolytes with or without vinylene carbonate confirms the beneficial effects of the additive in stabilizing the composite anode.

Electrical double layer and rheological properties of yttria-stabilized zirconia suspensions in solutions of high molecular weight polyacrylic acid polymers

This work deals with the effect of the adsorption of two high molecular weight polyacrylic acid polymers (Carbopol) on the interfacial properties, and the rheology of aqueous zirconia suspensions. Since the Carbopol-covered particles can be thought of as “soft” colloids, Ohshima’s theory was used to gain information on the surface potential and the charge density of the polymer layer (Ohshima H (1995) Electrophoretic mobility of soft particles. Colloids Surf A Physicochem Eng Aspects 103:249–255). The effect of the pH of the solution on the double layer characteristics is related to the different conformations of the adsorbed molecules provoked by the dissociation of the acrylic groups present in polymer molecules. The electrokinetic properties of the suspensions are studied for different pH and Carbopol concentrations in solution in order to investigate the possible stabilization of the suspensions by electrostatic repulsion between the particles. The rheological behavior of the suspensions was investigated in steady-state and dynamic conditions, and the corresponding yield stress and storage modulus were obtained in absence and presence of polymer in solution. The competition between bridging flocculation provoked by polymer adsorption and electrosteric stabilization determines the rheological properties of the suspensions. In the pH range investigated, bridging flocculation predominates at the neutral pH because of the graft of the uncoiled polymer to more than one particle, while at the extreme pH values (pH 3, pH 9) steric or electrosteric stabilization seems to be the predominant mechanism that explains the rheological results. These facts were confirmed by estimating the zirconia particle (or aggregate) diameter in the liquid medium by means of light scattering measurements.

Synthesis, Characterization, and First Application of High Molecular Weight Polyacrylic Acid Derivatives Possessing Perfluorinated Side Chains and Chemically Linked Pyrene Labels

The authors describe a new and unique nanosensing system: a derivative of poly(acrylic acid) (PAA, Mw = 1.250.000) possessing a comb-like structure. This structure is achieved by attaching perfluorinated (−O−C8F17) functions and chemically linked pyrene fluorescent units to the PAA backbone. The average composition of the new materials was investigated by UV/vis absorption spectroscopy, DOC, and elementary (CHN) analysis. Gel permeation chromatography (GPC) led to the conclusion that the new polymeric materials are almost monodisperse. The change of pH from the basic to the acidic range causes a collapse of the macromolecules in an aqueous environment, and consequently the formation of preformed excimers is suppressed. These novel materials are designed for use in sensor heads for the photochemical detection of the pH and CO2 content of water solutions. Using a model experiment, we show how these characteristics can be used for the measurement principles.

Enhanced cytotoxicity of alkyl viologens and N,N'-diamino analogs toward cultured murine leukemia L1210 cells under vortex-stirring with a high molecular weight polyacrylic acid.

Alkyl viologens showed cytotoxicity when incubated with cultured murine leukemia L1210 cells for 48 h, whereas they were not cytotoxic when briefly incubated for 10 min. Under the permeabilizing conditions achieved by vortex-stirring with a high molecular weight polyacrylic acid (A-119), appreciable cytotoxicity was shown even after a 10-min exposure to any of the alkyl and amino viologens examined. Acetylamino viologen showed no cytotoxicity regardless of the presence or absence of A-119. This permeabilizing procedure was demonstrated to be applicable to internalize water-soluble positively charged viologen molecules into the cell.

A new method for permeabilization of the plasma membrane of cultured mammalian cells. VI. Enhanced cytotoxicity of bleomycin toward leukemia P388 cells inoculated into the tail vein of mice together with high molecular weight polyacrylic acid--similarity of blood stream to vortex-stirring.

Bleomycin (BLM) cytotoxicity was greatly enhanced by vortex-stirring BLM and mammalian cells for a few seconds together with 1 to 10 microg/ml high molecular weight polyacrylic acid (A-119). When suspensions of murine leukemia P388 cells were injected together with BLM and A-119 into the tail vein of CDF1 mice, cell viability was reduced to 1/1000, the reduction being similar to that obtained by vortex-stirring the cells with BLM and A-119 in vitro. This was corroborated by an increase in the survival time of these mice. The reduction in cell viability was noted only when the cells, BLM, and A-119 were simultaneously injected. There was absolutely no effect when there was a time-lag between cell inoculation and injection of BLM/A-119. These findings suggest that the conditions created by the blood stream may simulate those of vortex-stirring and that, in both cases, rapid uni-directional movement of cells with high molecular weight polyacrylic acid may affect the plasma membrane facilitating internalization of non-permeant materials into cells.

Internalization of non-permeant materials into cultured mammalian cells by vortex-stirring in the presence of a high molecular weight polyacrylic acid: direct evidence of internalization and its dependence on the cell lines.

Laser scanning confocal fluorescence microscopic analysis provided a direct evidence of the internalization of non-permeant Lucifer Yellow and a dextran of MW 4400 into leukemia L1210 cells when the cells were vortex-stirred in the presence of a high molecular weight polyacrylic acid, A-119 (MW ca. 9 x 10(6)). The morphology of A-119-treated cells was probed by scanning electron microscopic analysis using 2% glutaraldehyde for fixation. The cells immediately after vortex-stirring with A-119 showed many blebs on the cell surface, indicative of local weakening of the plasma membrane. The blebby surface returned to normal within 5 min at 37 degrees C, but not completely at 0 degree C even after 20 min. Several cultured cell lines, murine splenocytes, and Ehrlich ascites carcinoma cells were particularly susceptible to permeabilization by this method, although the efficiency varied from one type of cell to another.

Delicate and Crucial pH Dependence in Permeabilization of Cultured Mammalian Cells by Vortex-Stirring with a High Molecular Weight Polyacrylic Acid.

We previously reported that cultured mammalian cells are effectively permeabilized by vortex-stirring the cells with a high molecular weight polyacrylic acid. The present study revealed that the efficiency of permeabilization of a non-permeant dye, Lucifer Yellow (LY), was sensitively affected by the pH of the medium. When the cells are vortex-stirred in RPMI 1640 culture medium, the pH of the medium should be adjusted with HEPES and NaOH, instead of NaHCO3, to a pH higher than 7.6 for successful permeabilization; the higher the pH, the better the result obtained. Internalization of LY was near the background level at a pH below 7.4. When phosphate-buffered saline was substituted for RPMI 1640 medium, the optimal pH range was slightly shifted to a more acidic region. This requirement for the pH of the medium is indispensable as a supplement to the standard permeabilization procedure tentatively proposed in the preceding report.

A new method for permeabilization of the plasma membrane of cultured mammalian cells. VII. Permeabilization of growing and resting cells by vortex-stirring with high molecular weight polyacrylic acid.

The permeability of murine leukemia L1210 cells toward non-permeant fluorescent Lucifer Yellow (LY) was evaluated by flow cytometry following vortex-stirring of cells with LY in the presence of high molecular weight polyacrylic acid (A-119). Permeabilization by this technique was greatly enhanced in growth phase cells, whereas it was appreciably reduced in resting cells which had been maintained in unchanged culture medium for 2 to 4d. This reduction in permeabilization of resting cells gradually disappeared when resting cells were re-suspended in fresh culture medium, and the degree of permeabilization was restored to that of the growing cells. It is recommended that the mammalian cells used for internalization of non-permeant materials by vortex-stirring with A-119 should be in the early log phase.

A New Method for Permeabilization of the Plasma Membrane of Cultured Mammalian Cells. V. A Standard Procedure Recommended for Permeabilization by Vortex-Stirring with High Molecular Weight Polyacrylic Acid.

As we previously reported, permeabilization of the plasma membrane was effectively achieved by vortex-stirring the mammalian cells in the presence of high molecular weight polyacrylic acid. Based on our present findings, as well as those of our previous studies, optimal conditions for permeabilization were considered in terms of the structure and dosage of a polymer, the constituent of the medium, the conditions of cells, vortex-stirring and post-vortex-stirring, etc. A standard procedure tentatively recommended is described.

A new method for permeabilization of the plasma membrane of cultured mammalian cells. III. Internalization of fluorescent dextrans into cultured mammalian cells by vortex-stirring in the presence of high molecular weight polyacrylic acid.

Using a new technique of vortex-stirring of cells with high molecular weight polyacrylic acid (A-119), non-permeant fluorescein isothiocyanate-labeled dextrans (FDs) of various molecular weights were tested to determine the degree of their internalization into cultured murine leukemia L1210 cells. Flow cytometric analyses of the treated cells revealed that FD4 with a molecular weight (MW) of 4400 was incorporated in an amount 13.2 times greater than that of the control and that FDs with MW of 19000 and 38260 were incorporated at levels several times greater than control levels, by vortex-stirring cells with A-119 for 10 s followed by maintenance at 0 degree C for 10 min. Results of the present study indicate that this permeabilization procedure might be useful to introduce poorly permeable materials into cultured mammalian cells, which would be beneficial in various fields of biological research.