Plastic Beads Research Articles

The effect of selected basic dyes on the blood cells, in particular, the basophils, of the Pacific oyster, Crassostrea gigas

It was determined by means of light and electron microscopy that the granular blood cells of Pacific oysters could absorb basic dyes from dye-coated plastic beads injected into the mantles of oysters; in doing so, the granular blood cells became altered morphologically and behaviorally. In addition, it was observed that oysters could absorb the basic dye, toluidine blue O, in low concentration from sea water. It was shown that the absorption of toluidine blue O caused a shift in the blood cell levels of oysters, and it was suggested that this shift was occasioned by the absorption of toluidine blue O by granulocytes.

A simplified isotope release assay for cell-mediated cytotoxicity against anchorage dependent target cells

An assay for cell-mediated cytotoxicity has been developed in which anchorage-dependent target cells are cultured on small plastic beads in suspension. Confluent target cells on the beads are handled by methods appropriate to suspension-grown cells and labelled with chromium-51, iodine-125 and [ 3H]proline. Fetal human lung fibroblasts and HeLa cells were used as targets in model experiments measuring human natural killer cell activity. In 20 h experiments, chromium-51 was the most suitable isotope. In 40 h experiments, [ 3H]proline release assay was superior to chromium-51 and iodine-125 assays. The bead cytotoxicity assay offers a rapid and simple isotope release technique for anchorage dependent cells because no trypsinization and re-seeding of target cells is needed.

Contact inhibition and the movement of metal, glass and plastic beads within solid tissues.

Cell-size spheres of metal, glass and plastic can move from surface to subsurface positions within solid tissue masses in culture, demonstrating that the observed movement of cells in similar circumstances may not be due to ‘active cell locomotion’.

Optimal conditions for the separation of rat T lymphocytes on anti-immunoglobulin-immunoglobulin affinity columns

The separation of rat T lymphocytes was investigated on anti-IgIg columns. A simple and efficient method for the purification of rat Ig by precipitation of rat serum with sodium sulfate is presented. Protein binding characteristics of glass and plastic beads, as solid support of affinity columns, are described, as well as optimal parameters for coating beads with rat Ig (with BSA, ribonuclease and lysozyme, as comparison). Binding of Ig was primarily dependent on the concentration of the Ig solution. Maximal strong binding of Ig (6.2 × 10 3 molecules per μ 2 of bead surface) was reached at 400 μg per ml concentration of purified Ig solution during 20 min of incubation. Higher concentrations increased only the amount of loosely bound Ig on the surface of beads whereas the amount of firmly bound Ig remained unchanged. Fractionation of lymphoid cell suspensions on anti-IgIg affinity columns prepared at optimal conditions resulted in highly purified T-cell suspensions containing less than 1% of lymphocytes bearing surface Ig receptors.

A solid phase radioimmunoassay for detection of serum autoantibodies in systemic lupus erythematosus

Solid phase radioimmunoassay (RIA) methods for measuring autoantibodies in systemic lupus erythematosus (SLE) patients' serum were developed to improve the sensitivity and quantitative precision of these determinations. Two mechanical systems were studied: (1) acetone fixed cell monolayers in glass tubes and (2) antigen coated plastic beads. Both systems were sensitive and reproducible, giving serum dilution end-points between two and four orders of magnitude (100–10,000 times) greater than those obtained by fluorescence microscopy. The most sensitive, versatile system involves the coating of plastic beads with nuclear antigen(s), incubation overnight with sera and labelling with 125I conjugated antihuman globulin. Linear binding of this radioactive tag is obtained over a wide range of SLE serum dilutions and slopes of the serum dilution titration curves are almost identical for all SLE patients' sera we have tested. Therefore, a standard titration curve can be constructed from the results with a positive serum, and end-point dilutions of unknown sera estimated from results obtained with a single serum dilution. Alternatively, binding ratios of unknown sera can be usefully compared at fixed dilutions with standard positive and negative sera. For example, high binding ratios (> 3.0) were obtained with 19/20 SLE sera and 0/20 control sera. Antigens used in these systems include crude, whole-cell lysates and lysates from purified nuclei. These RIA methods appear to provide certain advantages over existing autoantibody assay methods because they are relatively simple, sensitive, reproducible and potentially capable of measuring a variety of autoantibody specificities.

Dissociation of the vitreo-retinal junction following experimental embolization of retinal circulation

Spheric plastic beads (7-25 mu in diameter) were injected into the central retinal artery of the owl monkey (Aotus trivirgatus) using a micro-surgical technique. The occlusion of retinal arteriolar branches resulted in posterior vitreous detachment and microcystoid degeneration in inner retinal layers. The dissociation of the vitreoretinal junction showed three different entities: detachment of the vitreous from the basal lamina (inner limiting lamina of the retina), separation of the basal lamina from the retina, and detachment of shreds of glial cells and other degenerated retinal tissue (intraretinal schisis).

Specific affinity fractionation of lymphocytes using glass or plastic bead columns.

This paper describes column fractionation techniques for separating different lymphocyte subpopulations. For this purpose columns can be coupled with antigen, specific antibodies to membrane determinants on lymphocytes, or immune complexes or aggregated immunoglobulins, which interact with Fc receptors.

Cell surface-associated and released proteolytic activities of bovine aorta endothelial cells

We have demonstrated cell surface-associated and released proteolytic activity on bovine aorta endothelial cells, representing normal cells with regulated invasive properties. To demonstrate these proteolytic activities on viable cells grown in monolayer cultures, a new method was developed. The method consists of rolling modified plastic beads carrying covalently-linked [ 125I]-labeled casein in contact with the cell surfaces or adjacent to the cell monolayers without actual contact. The rate of radioactive peptide release is proportional to the proteolytic activity. Released proteases are detected when no contact occurs between the substrate and the cells. When the bead-substrate complex is rolled over the surface of endothelial cells, a significant increase in released labeled peptides is observed which represents a cell surface-associated proteolytic activity. These activities may be relevant to the endothelial cell's invasive capacity and appear to be similar to the neutral proteolytic activity of transformed cells.

Insolubilization of protein antigens on polyacrylic plastic beads using poly-l-lysine

A new model system for quantitation of immunofluorescense on single cells is descibed using poly-L-lysine (PLL) coated polyacrylic plastic beads of approximately cell sizes as carriers for protein antigen. By increasing PLL concentration on the beads increased amounts of 125I labeled antigen were attached to the particles. Excess binding sites of PLL could be completely blocked by unrelated proteins. After staining with FITC-conjugated antibodies and quantitative fluorescence measurements of individual beads using a microspectrofluorimeter, strong correlations were found between antibody and antigen concentration on the beads. Neither repeated washings with PBS nor storage of the beads for two months caused detectable shedding of antigen—antibody complexes. There was a strong linear correlation between fluorescence intensity and the volume of beads, but the correlation between surface area and fluorescence was nonlinear. The described procedure was shown to be a simple method for quantitative and stable coating of particles with proteins. It can be applied as a useful model system for quantitative immunofluorescence studies on intracellular antigens.

Effect of plastic bead on gastric carcinogenesis in rats treated with N-methyl-N'-nitro-N-nitrosoguanidine.

Experiments were made on induction of cancer of the glandular stomach of rats by a combination of oral administration of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and insertion of a plastic bead into the gastric lumen. The incidence of cancer with this combination of treatments was greater than by administration of MNNG alone. Fluoroscopic examination showed that barium sulfate remained in the stomach longer when a bead had been inserted into the gastric lumen. This indicates that after insertion of a bead, MNNG must have remained in the stomach longer, and so the period of exposure of the gastric mucosa to the carcinogen must have increased.

Retinal detachment and pathology following experimental embolization of choroidal and retinal circulation.

By means of a micro-surgical technique, the central retinal artery of the owl monkey (Aotus trivirgatus) was embolized with spheric plastic beads (7-25 mu in diameter). Beads (4-15 mu) were also injected into choroidal capillaries through a supero-temporal vortex vein. Choroidal ischemia induced proliferation or degerneration of the pigment epithelium, frequently accompanied by photoreceptor degeneration and circumscribed retinal detachment. Occlusion of retinal vessels resulted in posterior vitreous detachment and micro-cystoid degeneration in inner retinal layers, followed by retinal schisis. In combined retinal and choroidal ischemia long-lasting retinal detachments regularly developed.

Antibody-dependent cell-mediated cytotoxic activity in syngeneic moouse ascites tumors.

Eight different syngeneic murine ascites tumor preparations were tested for their ability to lyse antibody-coated chicken erythrocytes in an assay for antibody-dependent cell-mediated cytotoxicity (ADCC). All of the tumor preparations demonstrated significant ADCC effector activity, even at effector-cell:target-cell ratios of less than 1:1. In contrast, none of eight in vitro murine tumor-cell lines showed any consistent ability to function as ADCC effector cells. Several cell fractionation experiments were performed to ascertain the nature of the effector cells in the ascites tumors. Adherent cells were removed from tumor preparations by passage of cells over columns of Degalan plastic beads and 19S Fc-receptor-bearing cells were removed by sedimentation of EA rosette-forming cells. Although each of these procedures depleted only a minority of the total cells, all the ADCC activity was simultaneously removed in each case. Depletion of phagocytic cells by treatment of tumor preparations with carbonyl iron and magnetism only slightly diminished the ADCC activity. These results indicate that the ADCC activity associated with ascites tumors is due to effector cells most probably of host origin, but possibly representing an atypical sub-population of tumor cells. The characteristics of the effector cell closely parallel those defined for the major non-phagocytic ADCC effector cell in normal mouse spleen and peritoneal exudate.

Effect of carrageenan on the development of hypersensitivity (Schistosoma mansoni egg) and foreign body (divinyl-benzene copolymer beads and bentonite) granulomas.

The effect of carrageenan on the development of hypersensitivity and foreign body granulomas was examined. Repeated intraperitoneal injections of the polysaccharide caused a strong suppression of the primary, hypersensitivity granulomas forming around the eggs of Schistosoma mansoni worms in the lungs of mice, but had no effect on the secondary, enhanced egg granulomas in the previously sensitized animals. Carrageenan treatment virtually abolished the granulomatous reactions forming around plastic beads or bentonite particles. It is concluded that carrageenan exerted its suppressive effect not through macrophage toxicity but rather by affecting plasma factors which regulate vascular permeability and cellular traffic during the process of inflammation.

The participation of thymus-derived and of bone marrow-derived lymphocytes of sensitized mice, in the proliferative response to specific antigen, in vitro.

AbstractSpleen cells of mice primed with sheep red cells (SRC) responded with increased thymidine uptake when stimulated with SRC in vitro. Depletion of bone marrow‐derived (B) cells by filtering the spleen cells through columns of plastic beads coated with anti‐mouse immunoglobulin serum resulted in a drastic loss of the proliferative response. A similar loss occurred on depleting the thymus‐derived (T) cells by treatment with anti‐Θ serum and complement. Addition of peritoneal exudate cells failed to restore the reactivity of the purified T cells to SRC. The proliferative response could be restored to a level exceeding the sum of the individual responses, by mixing the purified T with the purified B cells. A synergistic interaction between T and B cells is suggested.

In Vivo Suppression by Cholera Toxin of Cell-Mediated and Foreign Body Inflammatory Responses

Abstract Cholera toxin (CT) injected i.v. into mice resulted in a rise in cyclic AMP levels in splenic white cells. This was associated with and followed by a marked and relatively prolonged decrease in both the spleen cells and circulating lymphocytes. The effect of CT on a series of cell-mediated immunologic reactions related to the pathogenesis of schistosomiasis was then tested. CT profoundly suppressed dermal footpad swelling to soluble schistosome egg antigens, granuloma formation around schistosome eggs injected into the pulmonary microvasculature of unsensitized and sensitized mice, production of macrophage migration inhibition factor by intact granulomas (obtained from the livers of infected mice) maintained in vitro in tissue culture, and it ameliorated portal hypertension and esophageal varices in hepatosplenic schistosomiasis. In addition CT significantly prolonged allograft survival in mice even though it was first administered 6 days after grafting. Finally, CT completely eliminated foreign body granuloma formation around plastic beads (used as controls for the egg granulomas) which is a nonimmunologic reaction due to activation of chemical mediators of inflammation.

The enrichment of T-cell functions in a lymphocyte population by anti-immunoglobulin columns.

Mouse lymphocytes have been fractionated using purified anti‐light chain antibodies adsorbed onto plastic beads. B cells were specifically bound on these columns, as evidenced by the absence of anti‐light chain immunofluorescent staining of the effluent cells and a two‐fold enrichment of T cells as judged by cytotoxicity and fluorescent staining using specific anti‐T‐cell sera. The effluent population was still functional and showed a 10–30% enrichment with respect to the killing of allogeneic mastocytoma cells. Although only 71% of spontaneous and 58% of immune rosettes formed by unfractionated lymphocytes with sheep red cells could be inhibited by an AKR anti‐CBA θ serum, virtually all the rosettes formed by the anti‐light chain effluent cells were inhibited. θ‐bearing rosette‐forming cells from immunized animals were retained to a greater extent Dn the anti‐LC as compared with control columns.

Food Size Selection Among Copepods

The marine copepod Acartia tonsa was fed small plastic beads (7—70 microns) in a variety of size—frequency distributions and a concentration of 3,000—4,000 beads/ml. Both selective and non—selective feeding were observed. Selective feeding was intense over a narrow size range (selected particle sizes over 20 times more abundant in the gut than in the water); the actual sizes selected depended on the size frequency distribution offered, associating always with the largest abundant particles. A hypothesis for the mechanism of food size selection is that the animals "scan" the size distribution by capturing particles larger than the previous one ingested more efficiently than those that are smaller; the copepods then "locate" an abundance peak and halt the process just to the right of it.

The specific separation of enzyme antigen-binding cells by substrate affinity chromatography.

The elegant work of Wigzell and his collaborators (1) has clearly established that lymphocyte populations can be deprived of a subfraction of cells reactive with a specific antigen when cells are passed through a column of glass or plastic beads coated with the test antigen. However, little success was experienced in using such columns to enrich for lymphocytes specific to a given antigen. Recently, Wofsy and coworkers (2), using Polyacrylamide beads to which azophenyl-β-lactoside (=LAC) was coupled, have succeeded in not only depleting lymphocyte populations of, but have also enriched for, LAC-re-active cells which are involved in anti-LAC antibody production.KeywordsBinding CellThymus CellMouse Bone Marrow CellPlastic BeadElegant WorkThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Immunoglobulins on the Surface of Lymphocytes

Abstract The presence of immunoglobulin (Ig) on the surface of lymphocytes that were negative for Ig by immunofluorescence was investigated by combining a quantitative inhibition assay with immunofluorescence. Thymus cells; thymus, spleen and lymph node cells that had been passed through anti-Ig-coated plastic bead columns; and ϑ-positive mouse lymphomas were evaluated by using these two systems. These cell preparations had 0 to 1% of cells with detectable surface immunoglobulin by immunofluorescence. If the assumption is made that the fluorescent-positive cells were relatively homogeneous with respect to the amount of surface Ig, then it could be calculated that there was more Ig present in the various cell populations studied than was accountable by the small percentage of fluorescent-positive cells. Based on these calculations, the amount of Ig present in the different populations was found to be: fluorescent-positive cells: 5 to 14 ng N/106 cells; fluorescent-negative thymus cells: 0.01 ng N/106 cells; fluorescent-negative spleen and lymph node cells 0.07 to 0.1 ng N/106 cells. Three of four ϑ-positive lymphomas also had detectable Ig in amounts 3 to 10 times that found in normal thymus. Routinely there was more IgM found in both fluorescent-positive and -negative normal cells than IgG2.

Removal of immunoglobulin-bearing lymphocytes by anti-immunoglobulin-coated columns

As measured by several immunochemical techniques, lymphocytes can be divided into two categories: those with easily detectable surface immunoglobulin and those with undetectable immunoglobulin. This paper describes a technique by which mouse spleen cell populations can be depleted of these immunoglobulin-bearing cells. By utilizing anti-immunoglobulin-coated plastic bead columns, we have been able to deplete cell populations of immunofluorescent-positive cells. This depletion was about 90% effective after a single passage through an anti-immunoglobulin-coated column. while serial passage of the cells through several columns resulted in a 99.8% depletion. In addition, two sets of experiments demonstrated that these columns deplete cell populations of certain immunocompetent cells. First, preexisting direct and indirect plaque-forming cells are specifically retained by these columns; and second, a population of cells which is necessary for reconstitution of lethally irradiated mice is removed from normal mouse spleens by these columns.