Biomedical subjects

M V Sefton

Publications and source records attributed to M V Sefton.

At least 37 records · Page 2Linked to original sources

PubMed1998

Surface grafting of poly(ethylene glycol) onto poly(acrylamide-co-vinyl amine) cross-linked films under mild conditions.

Poly(ethylene glycol) (PEG) was grafted onto poly(acrylamide-co-vinyl amine) (poly(AM-co-VA)) film using tresylated PEG (TPEG) at 37 degrees C in aqueous buffers (pH 7.4) with a view to surface-modifying microencapsulated mammalian cells. Poly(AM-co-VA) film was synthesized by Hofmann degradation of a cross-linked poly(acrylamide) film. Conversion to vinyl amine on the surface of the film was approximately 50%, but bulk conversion was not observed; surface specificity was thought to be the result of cleavage of aminated polymer chains at the surface due to chain scission. Reaction between primary amine and TPEG gave a graft yield of 2 mol% (based on XPS) with respect to available surface amine groups, equivalent to 54 mol% ethylene oxide based on monomer units. Physical adsorption of non-activated polymer was done under identical conditions as a control and the difference in oxygen content was significant compared to TPEG. The type of buffer agent and buffer concentration did not influence graft yields. This graft reaction, which was completed in as little as 2 h was considered to be mild enough to be used for a surface modification of microcapsules containing cells without affecting their viability. Such a surface modification technique may prove to be a useful means of enhancing the biocompatibility of microcapsules (or any tissue engineering construct) even after cell encapsulation or seeding.

Acrylic Resins↗

PubMed1998

Tissue engineering.

Wound care has become one of the first fields to see the benefit of a new technology: tissue engineering. Tissue engineering involves the development of new materials or devices capable of specific interactions with biological tissues. In wound care, these materials may be based entirely on naturally occurring tissues and cells, or may be materials that combine synthetics, usually polymers, with biological layers. Both wound dressings and skin substitutes are available. The complexity of the materials depends on the end uses. Generally, synthetics made from polymeric materials such as Tegaderm and Opsite are used as wound dressings over relatively simple and shallow wounds or as coverings over more complex dressings. Their function is one of protection from water loss, drying, and mechanical injury. More complex dressings vary from dermal replacements made of reconstituted collagen and chondroitan sulfate backed by a polymer layer such as Integra(R) to the complex Apligraftrade mark that contains collagen and seeded cells. This last is designed as a complete skin replacement or skin substitute. Ultimately, engineered skin will contain all of the components necessary to modulate healing and provide the desired response: a wound closed with limited scar tissue that retains all of the characteristics of natural skin.

Biocompatible Materials↗

PubMed1997

Selected aspects of the microencapsulation of mammalian cells in HEMA-MMA.

Microencapsulation of live mammalian cells is one means of creating hybrid artificial organs, like an artificial pancreas or an artificial liver. In addition to creating and developing the methodologies for enclosing cells within the appropriate semipermeable and biocompatible membranes, novel techniques are needed to assess the various features of the resulting capsules. The small size of a capsule or its heterogeneity can lead to additional complexities that go beyond the problem of examining cell behavior in the presence of biomaterials. These problems are illustrated here by comparison of protein release by microencapsulated HepG2 cells within large and small HEMA-MMA (hydroxyethyl methacrylate-methyl methacrylate) capsules, by assessment of the effect of processing conditions on HEMA-MMA microcapsule permeability to horseradish peroxidase at the individual capsule level, and by a confocal microscopy technique for assessing intracapsule cell viability.

Animals↗

PubMed1997

Flow cytometric analysis of material-induced platelet activation in a canine model: elevated microparticle levels and reduced platelet life span.

Assessment of material-induced platelet activation is important given that it is thought to be a major mechanism of biomaterials thrombogenicity. We monitored, by flow cytometry, platelet microparticle (MP) levels in the circulation during the connection of polyvinyl alcohol (PVA) hydrogel and polyethylene (PE) test segments (3.18 mm ID, 20 and 50 cm L) to our chronically shunted beagle dogs. We report that circulating microparticle levels were dependent on test segment material, length, and time. The connection of 50-cm lengths of PVA hydrogel test segments led to MP levels two to three times greater than background at 48 h, while the connection of polyethylene test segments did not lead to elevated microparticle levels. MP levels were near background 24 h after removal of the PVA test segment. To determine platelet life span during the connection of test segments, platelets were labeled in vivo with biotin and their disappearance monitored flow cytometrically. While platelet life span for shunted dogs (no test segment) was 4.7 +/- 0.2 days, the connection of PVA hydrogel test segments led to a platelet life span of < 2 days.

Animals↗

PubMed1997

Effect of C4-, C8- and C18-alkylation of poly(vinyl alcohol) hydrogels on the adsorption of albumin and fibrinogen from buffer and plasma: limited correlation with platelet interactions.

Polyvinyl alcohol (PVA) hydrogel was partially alkylated with short (C4), intermediate (C8) and long (C18) alkyl chains to test the hypothesis that an alkylated surface might promote enhanced interaction with albumin and thus exhibit low platelet thrombogenicity. PVA hydrogel was reacted with alkyl halides (C4, C8 or C18) and coated onto polyethylene. The effect of surface alkylation (extent of alkylation and alkyl chain length) on the adsorption of human serum albumin and fibrinogen to these surfaces was investigated in both buffer and plasma. Platelet interactions were investigated in vitro using flow cytometry methods. The maximum surface concentrations of albumin and fibrinogen adsorbed from buffer onto PVA and alkylated PVA were characteristic of monolayers. At low concentrations differences in adsorption among the surfaces appeared to be related to hydrophobicity as determined by dynamic advancing water contact angle, and to degree of alkylation as determined by angle dependent XPS analysis. Alkyl chain length dependence was not observed. Adsorption from plasma was considerably lower than from buffer, except for albumin on C8-PVA where monolayer adsorption was observed. Fibrinogen adsorption from plasma was similar on PVA, C8-PVA and C18-PVA, but was higher on C4-PVA. For albumin adsorption from plasma, the initial slope of the adsorption-concentration curve was highest for C18-PVA, suggesting higher albumin affinity despite the low degree of substitution of the C18-PVA material. These data suggest possible selectivity of the C18 alkylated PVA for albumin. Platelet studies showed that C4-PVA was the least platelet reactive (microparticle generation and P-selectin expression) of the alkyl derivatized materials.

Adsorption↗

PubMed1997

Coagulation on biomaterials in flowing blood: some theoretical considerations.

Are truly inert biomaterials feasible? Recent mathematical models of coagulation which are reviewed here suggest that such materials are impossible. This conclusion, which is certainly consistent with our collective experimental evidence, arises from the calculation that conversion of Factor XI to XIa never drops to zero even at the highest flow rates and with virtually no Factor XIIa bound to a surface. Residual amounts of XIa are still formed which can in principle kick-off the coagulation cascade. Furthermore, if the flow rates and corresponding mass transfer coefficients are low and in spite of these near-vanishing levels of the initiating coagulants, the surprising result is that substantial amounts of thrombin are produced. On the contrary, under slightly higher flow conditions, there can be more substantial levels of initiating coagulants, yet paradoxically thrombin production is near zero. This article presents a theoretical understanding of the events which take place during the interaction of biomaterials with flowing blood. We follow these events from the time of first contact to the final production of thrombin. The effect of flow and surface activity on the contact phase reactions is examined in detail and the two are found to be intertwined. The common pathway is also examined and here the main feature is the existence of three flow dependent regions which produce either high or very low levels of thrombin, as well as multiple thrombin steady states. In a final analysis we link the two segments of the cascade and consider the events which result. In addition, we note that multiple steady states arise only in the presence of two (thrombin) feedback loops. Single loops or the bare cascade will produce only single steady states. With some imagination one can attribute to the feedback loops the role of providing the cascade with a mechanism to produce high thrombin levels in case of acute need (e.g. bleeding) or to allow levels to subside to 'stand-by' when there is no need for clotting. We present this as a partial answer to the question: Why is the coagulation cascade so complex and what is the importance of the feedback loops?

Biocompatible Materials↗

PubMed1997

X-ray photoelectron spectroscopy (XPS) surface analysis of HEMA-MMA microcapsules.

High resolution carbon, C 1s, X-ray photoelectron spectroscopy (XPS) of the surface of hydroxyethyl methacrylate-methyl methacrylate (HEMA-MMA; 75 mol% HEMA) capsules maintained in PBS for 1 week showed that the surface was not pure HEMA-MMA. In these spectra, more carbon was bonded in the C-O form than in the C-C form indicating the presence of the Pluronic surfactant, L101, adsorbed from the precipitation bath to the surface during microcapsule preparation. Capsules maintained in medium containing fetal bovine serum for 1 week showed a nitrogen signal consistent with the presence of adsorbed serum proteins. There was a decrease in the amount of nitrogen on the surface after phosphate buffered saline (PBS) washing, however this did not decrease to zero. These preadsorbed proteins, present on the surface of capsules incubated in serum-containing medium before their implantation, may affect the tissue response to these capsules. Calcium was not detected on freshly-made capsules or capsules maintained in PBS for 1 week but was detected on capsules maintained in medium containing serum. Calcium deposits, if formed in vitro, could act as nucleation sites for calcification of the polymer in vivo.

Adsorption↗

PubMed1996

Material-induced up-regulation of leukocyte CD11b during whole blood contact: material differences and a role for complement.

Material-induced thrombogenicity is in part a consequence of leukocyte activation. To evaluate and compare material-induced platelet damage, we have expanded our in vitro flow cytometric immunoassay to include assessment of leukocyte activation. We have used a very simple system whereby fresh, heparinized whole blood contacts materials for 1 h at 37 degrees C under low shear. Unlike other tests that focus on adherent leukocytes, this assay evaluates the leukocytes in the whole blood drained from the tube (1.57 mm internal diameter, 25 cm length) after material contact. We demonstrate that whole blood contact with a polyvinyl alcohol (PVA) hydrogel surface leads to a twofold up-regulation in CD11b surface expression of all monocytes and neutrophils. The activation is metal-ion dependent and highly material dependent in that blood contact with polyethylene and Silastic surface leads to minimal activation. The shedding of L-selection as a marker of leukocyte activation was found to be unsuitable in our assay given it ease of shedding in resting heparinized whole blood. Further, plasma levels of complement components Bb and sC5b-9 (ELISA assays) were significantly elevated only after blood contact with PVA hydrogel surfaces (9.4 micrograms/mL sC5b-9 and 9.6 micrograms/mL Bb). Use of recombinant soluble human CR1 (sCR1) to inhibit the action of the C3 and C5 convertases completely inhibited sC5b-9 levels in whole blood after contact with PVA hydrogel surfaces and inhibited CD11b up-regulation by over 70%, suggesting that material-induced leukocyte activation is partially mediated by C5a production.

Biocompatible Materials↗

PubMed1996

Preparation of purified atactic polypropylene and polyvinyl methyl ether surfaces for thrombogenicity studies.

Commercial samples of atactic polypropylene (aPP) and polyvinyl methyl ether (PVME) were purified and spin-cast onto glass coverslips with a view to using these as model surfaces in thrombogenicity studies. These materials differ from polyvinyl alcohol (PVA) in a single functional group and are similarly amorphous: with the same backbone they have a hydroxyl, a methoxy, or a methyl group. The objective was to understand the role of the hydroxyl group in the platelet reactivity of PVA. Surface characterization showed that they were chemically pure (as determined by X-ray photoelectron spectroscopy) but not smooth (as determined by scanning electron microscopy or interferometry), presumably due to the difficulties of spin-casting optically clear films from hot solutions (aPP or polyethylene [PE]) or because of imperfect adhesion to the saline-treated substrate (PVME). PVME was also gamma-irradiated to insolubilize it. Fewer platelets adhered to PVA than to PVME or to aPP and PE, but roughness effects and limited data preclude definitive conclusions regarding the effect of functional groups. Less protein was found on PVA than on the hydrophobic surfaces, but the significance of this observation is unclear. Further studies with more sensitive protocols are called for to examine the extent of platelet activation and its relationship to surface chemistry.

Adsorption↗

PubMed1996

Dopamine secretion by PC12 cells microencapsulated in a hydroxyethyl methacrylate--methyl methacrylate copolymer.

A rat pheochromocytoma cell line (PC12) was encapsulated in a water-insoluble hydroxyethyl methacrylate-methyl methacrylate copolymer by interfacial precipitation from a polyethylene glycol 200 solution into phosphate-buffered saline. The resulting capsules (660 +/- 44 microns in diameter; 84 +/- 27 microns wall thickness) contained viable PC12 cells in a spheroidal arrangement, much like tumour spheroids, the latter grown on surfaces unsuitable for cell attachment. In these spheroids, the viable cells formed a band approximately 100 microns thick, surrounding an inner core of necrotic cells. A similar arrangement was seen 14, 28 and 42 days after encapsulation, with capsules maintained in an in vitro tissue culture environment; the annular ring was roughly constant in size, although the packing density appeared to increase over the 6 week observation period. During the first 4 weeks, when measurements were made the encapsulated cells converted a tetrazolium dye (MTT) into an insoluble formazan product, in a time-after-encapsulation-dependent manner. This indicated that PC12 cells retained viability despite encapsulation and an ability to increase (at least in part) their metabolic capacity, presumably by a combination of proliferation and altered cellular activity. The encapsulated PC12 cells also secreted dopamine when incubated in a high potassium release medium but not in a low potassium, conventional tissue culture medium (RPMI 1640). Consistent with the MTT results, the amount of dopamine released was also dependent on the time after encapsulation, as well as the cell density at the time of encapsulation.

Animals↗

PubMed1996

Preparation and characterization of alkylated poly(vinyl alcohol) hydrogels using alkyl halides.

A poly(vinyl alcohol) hydrogel coated onto polyethylene was partially alkylated by reaction with an alkylhalide (C4, C8, or C18) in the presence of a deprotonating agent (sodium ethoxide or potassium tert-butoxiDATE Surface coverage determined by X-ray photoelectron spectroscopy (XPS) was respectively approximately 34, 25, and approximately 8% for the C4, C8, and C18 modified surfaces. Statistically significant differences were observed in the fraction of C8 and C18 grafted alkyl groups as a function of depth (i.e. take-off angle) indicating the presence of a verticle composition gradient. All three surfaces showed maximal surface coverage of alkyl groups after 1 h reaction. At this reaction time, no further coverage was observed beyond a base/PVA ratio twenty times greater than the stoichiometric ratio. The advancing contact angle data exhibited an increase in hydrophobicity that correlated with the degree of coverage obtained by XPS: 90 +/- 1, 83 +/- 0.5, and 71 +/- 1 deg for C4, C8 and C18 alkylated PVA, and 55 +/- 2 deg for PVA respectively. Large contact angle hysteresis was observed on all three surfaces consistent with surface heterogeneity.

Alkylation↗

PubMed1995

Platelet activation in whole blood by artificial surfaces: identification of platelet-derived microparticles and activated platelet binding to leukocytes as material-induced activation events.

Because the lack of thromboresistant vascular biomaterials is in part due to platelet activation, we have attempted, by using fluorescence-activated flow cytometry, to fully characterize the platelet population after in vitro material contact with whole blood. We have used a very simple, near-physiologic system whereby whole blood, anticoagulated with D-phenylalanyl-L-prolyl-arginyl chloromethyl ketone (thrombin inhibitor), contacts materials for 1 hour at 37 degrees C, under low shear. Unlike other tests of platelet compatibility that focus on adherent platelets, this assay evaluates the platelets in the whole blood drained from the tube (1.57 mm internal diameter, 25 cm length) after material contact. We demonstrate for the first time significant materials-induced microparticle formation. One-hour contact with Silastic, polyethylene, and polyvinyl alcohol hydrogel surfaces lead to 30 +/- 1, 33 +/- 4, and 43 +/- 4 x 10(9) microparticles/L, respectively, whereas resting blood samples contained only 10 +/- 1 x 10(9) microparticles/L. In addition, significant increases in activated platelet(s) binding to neutrophils/monocytes after material contact were noted for all surfaces tested. For polyvinyl alcohol hydrogel surfaces a greater than 500% increase in the fluorescent intensity over that of resting whole blood was attained. The addition of monoclonal antibodies to GPIIb/IIIa (A2A9), the tetrapeptide adhesion ligand RGDS (arginine-glycine-aspartate-serine), or the calcium ion chelator ethyleneglycol-bis-(B-aminoethyl-ether)- N,N,N',N'-tetraacetic acid to the whole blood before material contact fully inhibited all platelet reactivity noted for all surfaces--platelet microparticles, platelet P-selectin expression, loss of platelets from bulk, and the formation of platelet/leukocyte aggregates--thereby indicating that material-induced platelet activation is a calcium-dependent process involving GPIIb/IIIa receptors.

Antibodies↗

PubMed1994

Immobilization of a lysine-terminated heparin to polyvinyl alcohol.

Lysine terminated heparin, prepared by the nitrous acid partial depolymerization and reductive amination of heparin, failed to increase the active heparin content of a heparin-polyvinyl alcohol (heparin-PVA) hydrogel relative to the unmodified commercial heparin. The depolymerization of heparin resulted in a loss of biological activity which outweighed the increase in the terminal amine groups (produced by reductive amination), that were used for glutaraldehyde immobilization to the PVA. The loss in anti-thrombin activity (thrombin time or chromogenic substrate) paralleled the increase in anhydromannose end groups due to depolymerization making it necessary to optimize the loss of activity against the increase in terminal amine groups after amination. For example, depolymerization at a high sodium nitrite concentration (81 g/l) at pH4 and 25 degrees C for 20 min, resulted in a loss of 22-40% of the biological activity but achieved an anhydromannose content of 600 nmoles/mg (approximately 7 cleavage sites/molecule). After the anhydromannose groups were reductively aminated by lysine, the anhydromannose content was reduced to 190 nmol/mg indicating a terminal lysine content of 410 nmol/mg. This resulted in an increase in heparin content of the final hydrogel by 53% on mass terms. However, given the reduction in biological activity, it was not surprising that the modified heparin-PVA hydrogel coated on a polyethylene tube was no better than the hydrogel with unmodified heparin in inactivating thrombin in a flow circuit. These results point out the need for care in interpreting heparin immobilization results and for new strategies to increase the active heparin content of this hydrogel.

Gels↗

PubMed1993

Platelet-derived microparticle formation involves glycoprotein IIb-IIIa. Inhibition by RGDS and a Glanzmann's thrombasthenia defect.

While the physiologic role of platelet microparticles may include a stable, physical dispersion of concentrated surface procoagulant activity the mechanism(s) of platelet vesiculation remains unknown. We demonstrate using flow cytometric methods a central role for the beta 3 integrin glycoprotein (GP) IIb-IIIa complex and its ligand tetrapeptide Arg-Gly-Asp-Ser (RGDS) binding site in platelet vesiculation. Time- and calcium-dependent vesiculation of platelets in response to ADP, collagen, thrombin, phorbol myristate acetate, and the thrombin peptide SFLLRN were dramatically inhibited, in a concentration-dependent manner, by monoclonal antibodies to GPIIb-IIIa (A2A9, 7E3, PAC1) and RGDS. Complete inhibition with A2A9 and RGDS occurred at 7.5 micrograms/ml and 75 microM, respectively, while control antibodies and a mock peptide had no effect. Platelet vesiculation requires intact GPIIb-IIIa and is fully supported by the intracellular pool of GPIIb-IIIa alone since de-complexing of this heterodimer by calcium chelation completely abolished microparticle formation in response to collagen (no alpha-granule release) but not to thrombin or SFLLRN. A central role for GPIIb-IIIa is supported by the near total inability of Glanzmann's thrombasthenic (type I) platelets to vesiculate in response to thrombin, ADP, collagen, and phorbol 12-myristate 13-acetate. This extends the biologic roles of GPIIb-IIIa to include platelet vesiculation and suggests that one or all of its binding ligands play a role.

Amino Acid Sequence↗

PubMed1993

Thrombin and albumin adsorption to PVA and heparin-PVA hydrogels. 2: Competition and displacement.

Thrombin adsorption to polyvinyl alcohol (PVA) was different from its adsorption to polyethylene (PE)--not so much in amount, but in its affinity. Thrombin was more easily displaced from polyethylene and its adsorption was more readily prevented by prior or simultaneous exposure to albumin. From PVA (or heparin-PVA), only approximately 30% of the adsorbed protein could be removed by a series of eluents, including even harsh ones such as 2.5M NaOH and 6M guanidine; > 85% could be removed from PE. Thrombin adsorption to PVA was not affected by the presence of BSA in solution or at the surface, but was virtually prevented on PE by preexposure to or adsorption with BSA. Heparin-PVA was not much different than PVA in most of these experiments, but did exhibit a "Vroman effect". In the absence of fibrinogen or antithrombin III, there was a maximum in thrombin adsorption from plasma at a plasma concentration of 1%. The behavior on this surface was dependent on both exposure time and protein concentration. These studies highlight the complexity of the interaction between plasma proteins and polymer surfaces (particularly hydrogel surfaces) and the difficulty of obtaining a clear picture of what happens when a single protein interacts with a polymer in the presence of other proteins.

Adsorption↗

PubMed1993

Microencapsulated human hepatoma (HepG2) cells: in vitro growth and protein release.

The feasibility of a microencapsulation process ultimately for cell transplantation was investigated by encapsulating human hepatoma (HepG2) cells in hydroxyethyl methacrylate-methyl methacrylate (HEMA-MMA) membranes through an interfacial precipitation process. Changes in viability and metabolic activity as well as protein secretion by the encapsulated cells were studied in vitro. When encapsulated at either low or high density (1 or 5 x 10(6) cells/mL, respectively), HepG2 cells retained their active metabolic state and/or proliferated during the initial 1-week period, after which a significant drop in cell viability was obtained. Encapsulation of a biological attachment substrate, Matrigel, along with the cells, however, resulted in rapid proliferation in both low and high density capsules with prolonged maintenance of an active metabolic state. The secretion of four model proteins (alpha 1-acid glycoprotein, alpha 1-antitrypsin, haptaglobin and fibrinogen) was demonstrated during the 2-week study period for the Matrigel encapsulated cells. Furthermore, the encapsulated cells remained responsive to interleukin 6 (IL6), a physiological stimulator of plasma protein secretion, as determined by the elevated secretion of haptaglobin in response to IL6 treatment. We conclude that HEMA-MMA capsules, in the presence of an attachment substrate, provide a suitable environment for the growth and expression of differentiated functions of encapsulated hepatoma cells.

Biocompatible Materials↗

PubMed1993

Immobilization of poly(ethylene glycol) onto a poly(vinyl alcohol) hydrogel: 2. Evaluation of thrombogenicity.

Immobilized polyethylene glycol (PEG) reduced the amount of bovine serum albumin (BSA) adsorbed on polyvinyl alcohol (PVA) hydrogel, but did not reduce the platelet reactivity of the hydrogel surface. PEG, molecular weight (MW) 2000 or 5000, with or without a monomethoxy end group, was covalently bound to glutaraldehyde-crosslinked PVA either through a cyclic acetal or an urethane functional group with a surface coverage of 70% (as measured by x-ray photoelectron spectroscopy [XPS]). Immobilization of monomethoxy-PEG via a cyclic acetal reduced BSA adsorption to PVA from 11 +/- 2 nmol/m2 to 3.9 +/- 0.3 nmol/m2 and 3.3 +/- 0.3 nmol/m2 for MW 2000 and 5000, respectively. Similarly, urethane bound PEG reduced adsorption to 3.5 +/- 1.6 nmol/m2 for MW 2000 and 5.4 +/- 1.0 nmol/m2 for MW 5000. Whole blood clotting times of PVA (using a Chandler loop) were not affected by covalently linked PEG, although the initial rate of thrombin generation at the surface, measured using a fluorogenic substrate, was marginally reduced; a rate constant of 4.2 +/- 0.1 cm/sec and 3.5 +/- 0.1 cm/sec were obtained for MW 2000 and 5000, respectively, compared to 5.6 +/- 1.0 cm/sec for PVA. Ex vivo evaluation using a canine arteriovenous shunt revealed that the hydrogel, with or without bound PEG, reduced circulating platelet levels by 35-70% after 4 days. The initial fractional rate of platelet destruction determined from measurement of platelet cyclooxygenase activity, indicated that cyclic acetal or urethane bound PEG of either molecular weight had no effect on platelet consumption produced by PVA.(ABSTRACT TRUNCATED AT 250 WORDS)

Adsorption↗

PubMed1993

Towards the preparation of a MMA-PEO block copolymer for the microencapsulation of mammalian cells.

Polymethyl methacrylate-polyethylene glycol-polymethyl methacrylate triblock copolymers (PMMA-PEO-PMMA) were synthesized by reductive amination coupling of preformed aldehyde-terminated PEO and amine-terminated PMMA. These were intended for use as high water content and therefore high permeability, biocompatible encapsulating materials for mammalian cells. Evidence for the formation of the block copolymer was obtained indirectly from precipitation experiments and IR analysis of the water-soluble extract of the polymer. Unfortunately the pure copolymer could not be separated from the homopolymers, because of the difficulty in finding appropriate non-solvents and the apparently limited yield of the product. Further work is necessary to confirm the underlying hypothesis of this work, i.e. that such a block copolymer would have a high permeability to the small proteins critical to microencapsulated cell survival or function.

Animals↗