Development of VH81X transgene-bearing B cells in fetus and adult: sites for expansion and deletion in conventional and CD5/B1 cells

Vitamin E (+/-alpha-tocopherol) was recently investigated as an antioxidant for implanted poly(etherurethane urea) (PEUU) elastomers. In that work, vitamin E prevented chemical degradation of biaxially strained PEUU up to 5 weeks implantation, and prevented pitting and cracking of the PEUU surface for the duration of the 10-week cage implant study. The promising results of the in vivo studies motivated a detailed comparison of vitamin E with Santowhite, the standard antioxidant used in PEUU elastomers. To evaluate vitamin E and Santowhite as antioxidants in PEUU, an accelerated in vitro treatment system was used that mimics the in vivo degradation of PEUUs. Vitamin E was even more effective than Santowhite in preventing pitting and cracking to the biaxially strained PEUU elastomers. The inhibition of ether oxidation was greater with vitamin E than with Santowhite when compared by equivalent concentrations and molar concentrations, respectively. It is hypothesized that the increased effectiveness of vitamin E in this system, compared to Santowhite, is due to differences in antioxidant mechanism(s). Vitamin E is more efficient in preventing PEUU oxidation than Santowhite because its phenoxy radical is more stable and it can terminate more than one chain per vitamin E molecule.     

Vascular cell attachment and procoagulant activity on metal alloys

The attachment and growth of vascular smooth muscle cells on biomaterials used as components of devices implanted in the vascular space may influence the biocompatibility of such materials. The nature of the materials may affect the attachment and/or the activation of these cells' procoagulant responses. Therefore, the main objective of this study was to measure the strength of adhesion of these vascular cells to potential biomaterials (titanium, zirconium alloys, and stainless steel) by exposing them to a range of shear stresses (50-300 dyn cm(-2)) in a parallel plate flow chamber. The procoagulant responses of the cells were evaluated by measuring the tissue factor (TF) activity promoted by the different materials under flow conditions. The materials supported distinctly different levels of initial cell adhesion in static culture. However, the fraction of adherent cells did not decline significantly with incrementally increasing shear stress within the range tested. TF expression, as measured by factor Xa (FXa) production. was material-dependent. For example, cells cultured on Ti1313 exhibited more FXa production (13.2 nM 10(-5) cells) than Ti1313(DH) (8.5 nM 10(-5) cells) or stainless steel (2 nM 10(-5) cells). Thus, our studies indicate that the level of adhesion, strength of attachment and the expression of procoagulant activity of adherent vascular cells depend strongly on the nature of the underlying biomaterial.     

In vitro side-view imaging technique and analysis of human T-leukemic cell adhesion to ICAM-1 in shear flow

The objective of the present study is to apply a novel side-view imaging technique to investigate T-leukemic Jurkat cell adhesion to a surface-immobilized ICAM-1 in shear flow, a ligand for leukocyte LFA-1. Images have revealed that Jurkat cell adhesion on ICAM-1 under flow conditions in vitro is quasistatic. The cell-substrate contact length steadily increased with time during the initial cell attachment to the ICAM-1-coated surface and subsequently decreased with time as the trailing edge of the cell membrane peeled away from the substrate under the influence of fluid shear forces. Changes in flow shear stresses, cell deformability, or substrate ligand strength resulted in a significant change in the characteristic adhesion binding time and contact length. A 3-D flow field with shear stresses acting on an adherent cell was calculated by using finite element methods based on cell shapes obtained from the in vitro images. The maximum shear stress acting on an actual cell body was found to be 3-5 times greater than the upstream inlet wall shear stress and was influenced by the extent of cell deformation within the flow channel. Therefore, the application of such a side-view imaging technique has provided a practical assay to study the mechanics of cell-surface adhesion in 3-D. The elongation of cells in shear flow tempers hydrodynamic shear forces on the cell, which affects the transients in cell-surface adhesion.     

Role of P-selectin and leukocyte activation in polymorphonuclear cell adhesion to surface adherent activated platelets under physiologic shear conditions (an injury vessel wall model)

Carbohydrate moieties on leukocytes adhere to activated platelets via P-selectin under static binding condition studies. We characterize polymorphonuclear cell (PMN) surface interactions with surface adherent platelets and the PMNs response, under physiologic flow conditions corresponding to a shear of 100 s-1, in an in vitro flow chamber. Fluorescent labeled PMNs with red blood cells were drawn through a transparent flow channel and visually quantitated over 30 minutes, interacting with a confluent monolayer of activated, shear-spread platelets expressing P-selectin. PMN adhesion was saturable (2,250 +/- 350/mm2), and time and cation (Ca2+, Mg2+) dependent, and PMNs did not bind to the experimental surface in the absence of a platelet monolayer. P-selectin antibodies completely abolished PMN adhesion in a concentration-dependent manner with half inhibition at 70 micrograms/mL. Antibodies to a putative P-selectin receptor CD15 (80H5 and MMA) maximally inhibited PMN adhesion by 73% and 10%, respectively. Adherent PMNs appeared morphologically activated and flow cytometric analysis of adherent PMNs confirmed activation because CD11b and CD18 surface expression was upregulated (100% and 27%, respectively), whereas L-selectin was downregulated (55%) compared with control nonadherent PMNs. In the presence of the metabolic inhibitor sodium azide (0.02% and 0.1%) there was a 23% +/- 9% and 51% +/- 3% decrease, respectively, in PMN adhesion at 100 s-1. Thus, P-selectin is required for PMN adhesion to a pathophysiologic surface of activated adherent platelets at physiologic shear rates. Furthermore, a secondary step involving PMN activation after platelet binding appears necessary for complete (irreversible) adhesion to occur. This unique flow cell provides a model to explore, under controlled conditions, biologic mechanisms and ligands involved in leukocyte-platelet binding that play important roles in PMN localization at sites of thrombosis and vascular injury.     

Through and beyond the wall: late steps in leukocyte transendothelial migration

A combination of approaches has provided firm experimental support for a step-wise model of leukocyte adhesion to vascular endothelium under hemodynamic shear stress. However, leukocyte adhesion to vascular endothelium acquires functional significance only if coupled to transmigration and progression into extravascular tissues. As discussed here by Elisabetta Bianchi and colleagues, these latter processes involve the sequential activation of adhesion-dependent functional programs, carried out by structurally diverse multimolecular complexes.     

Protein and platelet interactions with thermally denatured fibrinogen and cross-linked fibrin coated surfaces

In this work the hypothesis that a mature, cross-linked fibrin clot, pre-formed on a biomaterial, may be relatively nonthrombogenic was investigated. A cross-linked fibrin layer was formed on polyethylene which had been precoated with thermally denatured fibrinogen. Plasma protein adsorption and platelet interactions with the cross-linked fibrin and denatured fibrinogen surfaces were investigated. The adsorption of albumin, fibrinogen, and fibronectin from plasma was measured. For all three proteins, the cross-linked fibrin surface exhibited much higher levels of adsorption than either the thermally denatured fibrinogen or the polyethylene surface. Vroman peaks were observed for fibrinogen and fibronectin on polyethylene but not on the cross-linked fibrin and thermally denatured fibrinogen materials. In dilute plasma the thermally denatured fibrinogen surface showed considerable resistance to protein adsorption. However, at plasma concentrations greater than about 5% normal, this protein resistance was apparently lost. Platelet interactions (adhesion and release of granule constituents from adherent platelets) using suspensions of washed platelets in the presence of red cells were investigated at shear rates of 50, 300, and 525 s(-1) using a cone and plate apparatus. The levels of platelet adhesion on the different surfaces were in the order: adsorbed fibrinogen > cross-linked fibrin > thermally denatured fibrinogen = polyethylene. Platelets on the cross-linked fibrin surface also showed high levels of release indicating significant platelet activation. Scanning electron microscopic observations were in agreement with the platelet adhesion and release data, showing only a few (but well-spread) adherent platelets on the cross-linked fibrin surface.     

Albumin-binding surfaces: in vitro activity

Immobilized monoclonal antibodies (Mabs) have been used to attract specific molecules to a solid surface from complex mixtures such as blood, plasma or serum, thereby directing the response to the modified substrate, a key goal in rational biomaterial design. The nature of the Mab dictated the nature of the response: anti-albumin antibodies were used to prevent cell and platelet adhesion in vitro, whilst anti-fibronectin Mabs promoted attachment. Patterned surfaces could be formed, bearing Mabs that generated adhesive and non-adhesive regions. Fibrinogen adsorption from plasma showed a Vroman peak on unmodified control polymer, which was reduced by 64% in the presence of surface-bound anti-albumin Mab. Immobilization of a control Mab reduced fibrinogen adsorption only slightly, implying an albumin-mediated effect. In static tests, platelet adhesion from human platelet rich plasma was significantly reduced by the immobilization of anti-HSA Mab when compared to the untreated FEP surface (p < 0.0001). This effect was also seen with citrated blood flowing through Mab-treated polyurethane tubing at a shear rate of 132 s(-1) (p=0.034). Since platelets and proteins (as blood, plasma or serum) were introduced to the surface simultaneously, the generation of a defined protein film must have been sufficiently rapid as to shape the platelet or cell response.     

The role of air plethysmography in the diagnosis of chronic venous insufficiency

The role of protein and cellular components of thrombi in mediating bacterial adhesion on artificial surfaces was investigated in this study. The attachment of Staphylococcus aureus on polyurethane surfaces was observed directly using an automated video microscopy system. Surfaces were preconditioned with components of platelet-fibrin thrombi, including fibrinogen, thrombin, plasma, and isolated platelets. Experiments were performed in a radial flow chamber, and attachment rate constants were compared on the preconditioned surfaces in an effort to understand the complex relationship that exists between bacterial infection and thrombosis on synthetic biomaterials. Preadsorption of fibrinogen to surfaces significantly increased S. aureus adhesion compared to those preadsorbed with albumin alone while the presence of fibrin dramatically increased bacterial attachment compared to plasma preadsorbed surfaces. While the presence of adherent platelets also increased bacterial attachment, fibrin appeared to play a larger role in mediating bacterial adhesion on polyurethane surfaces. Striking results were obtained on the zwitterionic phosphonated polyurethane for a number of pretreatment conditions with regard to decreased bacterial adhesion and fibrinogen deposition.     

Survey of rheumatic heart disease in school children of Kinshasa town

Leukocyte adhesion under flow is preferentially mediated by the selectins. In this study we used intravital microscopy to investigate whether E-selectin may promote firm leukocyte adhesion in vivo. E-Selectin is expressed by endothelial cells activated with tumor necrosis factor-alpha (TNF-alpha) and causes slow leukocyte rolling. Microinjection of formyl-methionyl-leucyl-phenylalanine (fMLP) or macrophage inflammatory protein-2 (MIP-2) next to a venule of the TNF-alpha-treated mouse cremaster muscle significantly increased the number of adherent leukocytes. In gene-targeted mice homozygous for a null mutation in the E-selectin gene or in wild-type mice treated with an E-selectin monoclonal antibody (mAb), this response was significantly attenuated (by >80%). No such defect was seen in intercellular adhesion molecule-1 (ICAM-1)-deficient mice. E-Selectin-null mice showed more rapid leukocyte rolling than wild-type or ICAM-1-deficient mice, resulting in significantly shortened leukocyte transit times through venules. Topical application of fMLP onto the whole cremaster muscle generated the same number of adherent leukocytes in wild-type and E-selectin-deficient mice. We conclude that slow leukocyte rolling through E-selectin results in long transit times, which are essential for efficient leukocyte adhesion in response to a local chemotactic stimulus.     

Endothelial cells exposed to erythrocytes under shear stress: an in vitro study

After injury and vascular replacement, endothelial cell recovery is limited and could lead to thrombosis. Seeding small diameter vascular prosthesis with endothelial cells has been proposed to fulfil cell lining and improve surface hemocompatibility. However, detachment of seeded cells occurs following implantation. Previous in vitro studies have looked at the fluid shear stress as a major cause of cell detachment. To our knowledge, the role of erythrocyte collisions has not been investigated. The present in vitro study aims at investigating whether endothelial cell adhesion depends on (i) the presence of erythrocytes in flow and (ii) the latent culture period (1, 24 and 48 h) between seeding and exposure to flow. Endothelial cells were exposed to culture media containing different erythrocyte concentrations using a steady laminar flow of 1350 ml min(-1) in a parallel plate flow chamber. Endothelial cell morphology in dynamic conditions was quantified and compared to that in static conditions. The projected area of cells were mostly found smaller under dynamic than static conditions, particularly at a wall shear stress of 23 dyn cm(-2). Cells from the 1 h latent culture period were oriented parallel to the flow axis and were more elongated than under static conditions. Conversely, endothelial cell shape was slightly modified when either the latent period or the wall shear stress was increased. Disparate orientation was observed on confluent endothelial cells (24-48 h latent period) exposed to shear stress with or without erythrocytes. Increasing fluid viscous forces due to erythrocytes play a critical role on the behaviour of freshly seeded endothelial cells upon exposure to blood flow.     

A comparison of the efficacy and tolerability of dorzolamide and acetazolamide as adjunctive therapy to timolol. Oral to Topical CAI Study Group

BACKGROUND: Recent observations provide evidence that complement is involved in the pathophysiology of ischemia/reperfusion injury. In this study, we assessed the impact of complement inhibition on hepatic microcirculation and graft function using a rat model of liver transplantation. METHODS: Arterialized orthotopic liver transplantation was performed in Lewis rats after cold preservation (University of Wisconsin solution, 4 degrees C, 24 h). Eight animals received the physiological complement regulator soluble complement receptor type 1 (sCR1) intravenously 1 min before reperfusion. Controls received Ringer's solution (n=8). Microvascular perfusion, leukocyte adhesion, and Kupffer cell phagocytic activity were studied 30-100 min after reperfusion by in vivo microscopy. RESULTS: Microvascular perfusion in hepatic sinusoids was improved in the sCR1 group (87+/-0.7% vs. 50+/-1%; P < 0.001). The number of adherent leukocytes was reduced in sinusoids (68.3+/-4.7 vs. 334.1+/-15.8 [adherent leukocytes per mm < or = liver surface]; P < 0.001) and in postsinusoidal venules after sCR1 treatment (306.6+/-21.8 vs. 931.6+/-55.9 [adherent leukocytes per mm < or = endothelial surface]; P < 0.001). Kupffer cell phagocytic activity was decreased in the sCR1 group compared to controls. Postischemic bile production reflecting hepatocellular function was increased by almost 200% (P = 0.004) after complement inhibition. Plasmatic liver enzyme activity was decreased significantly upon sCR1 treatment, indicating reduced parenchymal cell injury. CONCLUSIONS: Our results provide further evidence that the complement system plays a decisive role in hepatic ischemia/reperfusion injury. We conclude that complement inhibition by sCR1 represents an effective treatment to prevent reperfusion injury in liver transplantation.     

聚氨酯泡沫塑料对Au(Ⅲ)吸附动力学性能的研究

通过静态吸附实验研究了不同条件下聚氨酯泡沫塑料对Au(Ⅲ)的吸附动力学,并运用3种动力学方程拟合,分析了吸附动力学机制。研究结果表明:聚氨酯泡沫塑料吸附Au(Ⅲ)过程中,反应30 min基本达到吸附平衡,平衡吸附量随着温度的升高而降低;吸附过程符合准二级动力学方程,主要以物理吸附为主,吸附速率由颗粒内扩散和膜扩散共同控制。该研究对聚氨酯泡沫塑料法富集金的应用和改进具有指导意义。     

Novel anti-inflammatory compounds induce shedding of L-selectin and block primary capture of neutrophils under flow conditions

Leumedins are small molecules that inhibit neutrophil movement into inflamed tissues. These compounds have been shown to inhibit the adherence of neutrophils in static adhesion assays mediated by beta2-integrins. We now report that leumedins, like activating agents, induce the loss of L-selectin from the neutrophil surface. The loss of L-selectin is unrelated to the inhibition of static adhesion, since neutrophils that have been pretreated with leumedins to cause shedding of L-selectin, followed by removal of drug, adhere normally in a static adhesion assay, and this adhesion is inhibited upon readdition of leumedin. In an assay of adhesion to endothelial cells under conditions of physiologic wall shear stress, leumedins prevent both primary capture of neutrophils mediated by L-selectin and firm adherence mediated by beta2-integrins.     

In vivo and in vitro responses to poly(ethylene terephthalate-co-diethylene glycol terephthalate) and polyethylene oxide blends

Although biocompatible polymeric compounds are generally nontoxic, nonimmunogenic, and chemically inert, implants made from these materials may trigger acute and chronic inflammatory responses. These inflammatory reactions may induce degeneration of implanted biopolymer. Interactions between implanted biomaterial and inflammatory cells are mediated by many cellular events involving cellular adhesion and activation. We studied the inflammatory responses in vivo and in vitro to samples of biopolymers composed of poly(ethylene terephthalate-co-diethylene glycol terephthalate) plus 0, 5, 25% of polyethylene oxide. We observed that these biopolymers did not induce inflammatory responses when implanted in the peritoneal cavity of mice for 28 days. However we observed deposition of hyaluronic acid at the surface of implanted biomaterial, suggesting that tolerance to biomaterial occurred after surgical implantation. No significant adhesion of inflammatory cells such as mononuclear phagocytes and peripheral leukocytes were observed in vitro, when poly(ethylene terephthalate-co-diethylene glycol terephthalate) blends were used as substratum to cellular adhesion. These results suggest that blends composed of poly(ethylene terephthalate-co-diethylene glycol terephthalate) induce low inflammatory cell adhesion, since no rejection of biopolymer was observed when implanted in experimental animal models.     

Mitogen-activated protein kinase (ERK1/2) activation by shear stress and adhesion in endothelial cells. Essential role for a herbimycin-sensitive kinase

Fluid shear stress modulates vascular function and structure by stimulating mechanosensitive endothelial cell signal events. Cell adhesion, mediated by integrin-matrix interactions, also regulates intracellular signaling by mechanosensitive events. To gain insight into the role of integrin-matrix interactions, we compared tyrosine phosphorylation and extracellular signal-regulated kinase (ERK1/2) activation in adhesion- and shear stress-stimulated human umbilical vein endothelial cells (HUVEC). Adhesion of HUVEC to fibronectin, but not to poly-L-lysine, rapidly activated ERK1/2. Fluid shear stress (12 dyn/cm2) enhanced ERK1/2 activation stimulated by adhesion, suggesting the presence of a separate pathway. Two differences in signal transduction were identified: focal adhesion kinase phosphorylation was increased rapidly by adhesion but not by shear stress; and ERK1/2 activation in response to adhesion was inhibited to a significantly greater extent when actin filaments were disrupted by cytochalasin D. Two similarities in activation of ERK1/2 were observed: protein kinase C (PKC) activity was necessary as shown by complete inhibition when PKC was downregulated; and an herbimycin-sensitive (genistein- and tyrphostin-insensitive) tyrosine kinase was required. c-Src was identified as a candidate tyrosine kinase as it was activated by both shear stress and adhesion. These findings suggest that adhesion and shear stress activate ERK1/2 via a shared pathway that involves an herbimycin-sensitive tyrosine kinase and PKC. In addition, shear stress activates ERK1/2 through another pathway that is partially independent of cytoskeletal integrity.     

Effect of hypertonic saline on leukocyte activity after spinal cord injury

STUDY DESIGN: The effect of intravenous administration of hypertonic saline on leukocyte adhesion after compression injury of the spinal cord was evaluated. OBJECTIVES: To investigate changes in leukocyte adhesion after spinal cord injury and to evaluate the effect of hypertonic saline on this process. SUMMARY OF BACKGROUND DATA: Leukocytes have been thought to exacerbate tissue injury after ischemia-reperfusion. Downregulating and reducing the number of circulating leukocytes has attenuated tissue damage in various models of cerebral ischemia. Recently, investigators have reported that leukocytes exacerbate injury in the spinal cord after trauma. Other recent findings have indicated that hypertonic saline may play a role in decreasing leukocyte adhesion and activation. METHODS: Sprague-Dawley rats were anesthetized, and a C3-C5 laminectomy was performed. Injury was caused by 35 g of compression applied to the cord for 10 minutes. Animals were divided into three groups: sham treated, untreated, and treated. The treated animals received 7.5% hypertonic saline (5 mL/kg, intravenously) 5 minutes after the injury. Sticking leukocytes and shear rate were measured using fluorescence microscopy. RESULTS: Administration of 7.5% hypertonic saline after injury significantly decreased the number of sticking leukocytes in the venules and arterioles. Shear rate was unchanged between the groups. CONCLUSIONS: The results show that an increase in leukocyte adhesion after a compressive injury is attenuated by the administration of 7.5% hypertonic saline. The decrease in adhesion cannot be attributed to changes in the shearing forces, because no significant change was observed in the shear rate. Hypertonic saline may interfere with leukocytes directly by interfering with their ability to swell and thus may prevent activation.     

Endothelial cells in culture: an experimental model for the study of vascular dysfunctions

Culture of endothelial cells started two decades ago and is now a useful tool in understanding endothelial physiology and the study of the interaction of endothelial cells with blood cells and various mediators. In vitro proliferation can be measured by [3H]thymidine incorporation in defined conditions and gives reproducible results. Endothelial cells can be activated by several stimuli, including cytokines such as tumor necrosis factor-alpha and interleukin-1. Part of endothelial cell activation is defined by expression or overexpression of leukocyte adhesion molecules. Intracellular adhesion molecule (ICAM), E-selection and vascular adhesion molecule (VCAM) are receptor molecules for leukocyte adhesion. Leukocyte adhesion to endothelium can be measured in static but also in rheologically defined flow conditions. Normal red blood cells (RBCs) do not adhere to endothelium, while RBC from patients with sickle cell anemia, diabetes mellitus, and malaria have an increased adhesion to endothelium which is mediated by specific VCAM, receptor for advanced glycated end-products (RAGE), and ICAM, respectively. Binding of blood cells or activation by cytokine is followed by a series of reactions in endothelial cells associated with the modulation of prostacyclin, nitric oxide, tissue factor, and cytokine production. Modification of endothelial cell functions in culture is correlated to in vivo alteration of vascular wall properties, further supporting these cells in culture as a relevant experimental model.     

Microjet impingement followed by scanning electron microscopy as a qualitative technique to compare cellular adhesion to various biomaterials

Adhesion of cells to biomaterial surfaces is one of the major factors which mediates their biocompatibility. Quantitative or qualitative cell adhesion measurements would be useful for screening new implant materials. Microjet impingement has been evaluated by scanning electron microscopy, to determine to what extent it measures cell adhesion. The shear forces of the impingement, on the materials tested here, are seen to be greater than the cohesive strength of the cells in the impinged area, causing their rupture. The cell bodies are removed during impingement, leaving the sites of adhesion and other cellular material behind. Thus the method is shown not to provide quantification of cell adhesion forces for the metals and culture plastic tested. It is suggested that with highly adherent biomaterials, the distribution and patterns of these adhesion sites could be used for qualitative comparisons for screening of implant surfaces.     

Role of erythrocytes in leukocyte-endothelial interactions: mathematical model and experimental validation

The binding of circulating cells to the vascular wall is a central process in inflammation, metastasis, and therapeutic cell delivery. Previous in vitro studies have identified the adhesion molecules on various circulating cells and the endothelium that govern the process under static conditions. Other studies have attempted to simulate in vivo conditions by subjecting adherent cells to shear stress as they interact with the endothelial cells in vitro. These experiments are generally performed with the cells suspended in Newtonian solutions. However, in vivo conditions are more complex because of the non-Newtonian flow of blood, which is a suspension consisting of 20-40% erythrocytes by volume. The forces imparted by the erythrocytes in the flow can contribute to the process of cell adhesion. A number of experimental and theoretical studies have suggested that the rheology of blood can influence the binding of circulating leukocytes by increasing the normal and axial forces on leukocytes or the frequency of their collision with the vessel wall, but there have been no systematic investigations of these phenomena to date. The present study quantifies the contribution of red blood cells (RBCs) in cell capture and adhesion to endothelial monolayers using a combination of mathematical modeling and in vitro studies. Mathematical modeling of the flow experiments suggested a physical mechanism involving RBC-induced leukocyte dispersion and/or increased normal adhesive contact. Flow chamber studies performed with and without RBCs in the suspending medium showed increases in wall collision and binding frequencies, and a decrease in rolling velocity in the presence of erythrocytes. Increased fluid viscosity alone did not influence the binding frequency, and the differences could not be attributed to large near-wall excesses of the lymphocytes. The results indicate that RBCs aid in the transport and initial engagement of lymphocytes to the vascular wall, modifying the existing paradigm for immune cell surveillance of the vascular endothelium by adding the erythrocyte as an essential contributor to this process.     

Immunoblot analysis of proteins associated with HEMA-MMA microcapsules: human serum proteins in vitro and rat proteins following implantation

Human serum proteins and their fragments, associated with hydroxyethyl methacrylate-methyl methacrylate (HEMA-MMA) copolymer microcapsules, were characterized using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot analysis. Capsules were incubated with serum for 1 week in vitro and then dissolved in ethanol to also precipitate the adsorbed protein. The precipitate was dissolved in 2% (w/v) SDS (the 'capsule eluate') to be assayed by electrophoresis. The majority of proteins probed for in the immunoblots were detected in the capsule eluates. These included fibronectin, plasminogen, IgG, vitronectin, Factor B, Factor H, Factor I, C3, but not beta-lipoprotein, fibrinogen, HMWK, or IgM. Complement activation fragments were detected in both the immunoblots of the capsule eluates and the medium containing serum without capsules. Thus, the adsorption of these fragments, formed independent of capsule presence, may be partially or completely responsible for the complement fragments associated with capsules. The prevention of complement activation by the addition of 5.8 mM EDTA, at the beginning of the week-long incubation, resulted in fewer low-molecular-weight C3 fragments associated with capsules. Rat proteins were also detected in immunoblots of the eluate of 'free-floating' capsules from the rat peritoneal cavity following implantation for 1 day using anti-human antibodies. Detected proteins included HMWK, fibrinogen, antithrombin III, transferrin, alpha1-antitrypsin, fibronectin, albumin, alpha2-macroglobulin, vitronectin, beta2-microglobulin, Factor B and Factor I. Rat fibrinogen, IgG, and complement C3 fragments were also detected in these immunoblots, but with monoclonal antibodies against the rat proteins.