Partial dependence of human peripheral blood leukocyte binding to high molecular weight fucoidan on divalent cations

L-selectin (CD62L) is the principal leukocyte adhesion molecule for the high endothelial venules of peripheral lymph nodes. This adhesion has an absolute requirement for calcium ions. Nevertheless, some studies have shown carbohydrate adhesion receptor interactions on lymphocytes and neutrophils, including the L-selectin molecule, that are Ca-independent. In the present study fucoidan, a reportedly Ca2+ independent ligand of L-selectin, and Mabs to human CD62L were coupled to magnetic polystyrene beads (MPB), as a model of leukocyte-surface interactions, and the efficiency of human leukocyte separation was investigated. 30% of Ficoll-purified human mononuclear cells and 75% of dextran-purified human leukocytes (DPHL) were specifically bound by fucoidan-modified MPB in the presence of Ca2+; 55% of dextran-purified leukocytes were specifically bound in the absence of Ca2+. The specific binding was inhibited by an excess of free fucoidan. The data obtained show the presence of Ca-independent adhesion determinants, specific to fucoidan on human leukocytes. No significant specific binding of leukocytes to fucoidan-modified MPB was found after the incubation with fresh human Ca(2+)-depleted whole blood. More than 90% of DPHL were specifically bound to MPB modified with Mabs to human CD62L irrespective of Ca2+ presence. The same degree of separation was achieved after the incubation with fresh human Ca(2+)-depleted-whole blood with anti-CD62L modified beads.     

12-O-tetradecanoylphorbol-13-acetate disrupts actin filaments and focal contacts and enhances binding of fibronectin-coated latex beads to 3T3-L1 cells

The effect of a tumor-promoting phorbol ester on the binding of fibronectin-coated beads to 3T3-L1 cells was studied to clarify the relationship between the binding of fibronectin to the cells, cell adhesion, and the organization of actin filaments. Interference-reflection microscopy revealed focal contacts of 3T3-L1 cells with the substratum. Stress fibers observed after rhodamine-phalloidin staining were well-developed in the cells. Treatment of the cells for 20 min with 12-O-tetradecanoylphorbol-13-acetate (TPA), but not with phorbol, disrupted focal contacts and caused a reorganization of stress fibers to generate actin ribbons. Treatment of the cells with TPA enhanced the binding of beads coated with human plasma fibronectin to the cells, as observed after incubation for 6 h with the beads. The TPA-induced increase in the percentage of cells with bound beads was dependent on the duration of treatment with TPA and on the concentration of TPA. Treatment of the cells with TPA also enhanced proliferation of cells in a dose-dependent manner. The enhancement of binding of the beads by TPA was suppressed by addition of an adhesion-inhibitory peptide (Gly-Arg-Gly-Asp-Ser-Pro). Treatment with TPA did not enhance nonspecific binding of beads coated with heat-denatured bovine serum albumin. Furthermore, treatment of the cells with phorbol did not enhance the binding of beads coated with fibronectin. These results suggest that TPA specifically enhances the binding of fibronectin-coated beads to 3T3-L1 cells, and that TPA-induced binding of the beads may be related to disruption of focal contacts and reorganization of actin filaments.     

Abiotrophia defectiva adhere to saliva-coated hydroxyapatite beads via interactions between salivary proline-rich-proteins and bacterial glyceraldehyde-3-phosphate dehydrogenase

Abiotrophia defectiva is a species of nutritionally variant streptococci that is found in human saliva and dental plaques and that has been associated with infective endocarditis. In our previous study, it was found that A. defectiva could bind specifically to saliva-coated hydroxyapatite beads (SHA). This study identified a cell surface component of A. defectiva that promotes adherence to SHA beads. The binding of A. defectiva to SHA was reduced in the presence of antibodies against human proline-rich protein (PRP); these results suggested that PRP may be a critical component mediating interactions between A. defectiva and the salivary pellicle. Two-dimensional gel electrophoresis of whole A. defectiva cells followed by Far-Western blotting was conducted by probing with synthetic peptides analogous to the binding region of PRP known as PRP-C. The results indicate that an A. defectiva protein of 37 kDa interacts with PRP-C. The results of amino-terminal sequencing of the adhesive A. defectiva protein revealed significant similarity to glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Recombinant GAPDH bound to immobilized PRP-C in a dose-dependent manner and binding of A. defectiva to SHA or to PRP was reduced in the presence of anti-GAPDH antiserum. Western blotting or electron immunomicroscopic observations with anti-GAPDH antiserum revealed that this protein was expressed in both cytosolic and cell wall fractions. These results suggest that A. defectiva could specifically bind to PRP via interactions with cell surface GAPDH; the findings suggest a mechanism underlying A. defectiva-mediated adherence to saliva-coated tooth surfaces.     

Binding and phagocytosis of fibronectin-coated beads by BHK cells: receptor specificity and dynamics

Studies on the receptor specificity and dynamics involved in fibroblast phagocytosis of latex beads revealed the following: 1) Ligands other than fibronectin such as concanavalin A (ConA) and serum spreading factor, when coated on latex beads, were found to promote phagocytosis of the beads. This indicates that fibroblast phagocytosis, like spreading, is a ligand-receptor mediated phenomenon not specifically requiring fibronectin (pFN); 2) Anti-pFN antibodies were found to inhibit the ability of cells to ingest pFN-coated beads that previously were bound on the cell surfaces. Consequently, binding of beads to the cell surfaces per se is not a sufficient signal to promote ingestion of the beads; 3) Finally, divalent cations protected receptor function necessary for phagocytosis of pFN-coated beads from proteolysis by trypsin, as previously was found for receptors involved in cell attachment and spreading on pFN-coated culture dishes. Recovery experiments carried out with cells whose surface receptors had been destroyed indicated that there was an internal (or cryptic cell surface) pool of receptors that amounted to at least 50% of the receptors normally found on the cell surface. After complete destruction of the cell surface and cryptic pools of receptors, reappearance of receptors required for bead binding and phagocytosis required several hours and did not occur in the absence of new protein synthesis.     

Enhanced binding of fibronectin-coated latex beads to quiescent 3T3-L1 cells is correlated with escape from growth arrest

The possible involvement of fibronectin receptors in growth stimulation was investigated by an analysis of fibronectin-coated latex bead binding to 3T3-L1 cells under various conditions. 3T3-L1 cells, growth-arrested in a medium with a low concentration of calf serum, bound few fibronectin-coated beads. After addition of serum at concentrations of 1.0% or higher, there was a rapid and transient increase in the number of cells with bound beads and a subsequent increase in the incorporation of bromodeoxyuridine (BrdU) into cell nuclei. Incorporation of BrdU was observed in about 60% of the cells with bound beads. Fibroblast growth factor and platelet-derived growth factor at concentrations of 5 ng/ml or higher also enhanced binding of fibronectin-coated beads to cells. Stimulation of bead binding by epidermal growth factor and insulin was weak. Fibroblast growth factor, but not epidermal growth factor, increased the incorporation of BrdU into nuclei. These results indicate a relationship between stimulation of cell proliferation in quiescent cells and increased binding by cells of fibronectin-coated latex beads.     

Recruitment and dissociation of nonhomologous end joining proteins at a DNA double-strand break in Saccharomyces cerevisiae

Nonhomologous end joining (NHEJ) is an important DNA double-strand-break (DSB) repair pathway that requires three protein complexes in Saccharomyces cerevisiae: the Ku heterodimer (Yku70-Yku80), MRX (Mre11-Rad50-Xrs2), and DNA ligase IV (Dnl4-Lif1), as well as the ligase-associated protein Nej1. Here we use chromatin immunoprecipitation from yeast to dissect the recruitment and release of these protein complexes at HO-endonuclease-induced DSBs undergoing productive NHEJ. Results revealed that Ku and MRX assembled at a DSB independently and rapidly after DSB formation. Ligase IV appeared at the DSB later than Ku and MRX and in a strongly Ku-dependent manner. Ligase binding was extensive but slightly delayed in rad50 yeast. Ligase IV binding occurred independently of Nej1, but instead promoted loading of Nej1. Interestingly, dissociation of Ku and ligase from unrepaired DSBs depended on the presence of an intact MRX complex and ATP binding by Rad50, suggesting a possible role of MRX in terminating a NHEJ repair phase. This activity correlated with extended DSB resection, but limited degradation of DSB ends occurred even in MRX mutants with persistently bound Ku. These findings reveal the in vivo assembly of the NHEJ repair complex and shed light on the mechanisms controlling DSB repair pathway utilization.     

Ristocetin-dependent reconstitution of binding of von Willebrand factor to purified human platelet membrane glycoprotein Ib-IX complex

Whether the human platelet membrane glycoprotein (GP) Ib-IX complex is the receptor for ristocetin-dependent binding of von Willebrand factor (vWF) has been examined by reconstitution with the purified components using a solid-phase bead assay. Purified GP Ib-IX complex was bound and orientated on the beads via a monoclonal antibody, FMC 25, directed against the membrane-associated region of the complex. Specific binding of 125I-labeled vWF to the GP Ib-IX complex coated beads was strictly ristocetin dependent with maximal binding occurring at ristocetin concentrations greater than or equal to 1 mg/mL. Ristocetin-dependent specific binding of 125I-labeled vWF was saturable. The observed binding was specific to the interaction between vWF and the GP Ib-IX complex since there was no ristocetin-dependent specific binding of vWF if the physicochemically related platelet membrane glycoprotein, GP IIb, was substituted for the GP Ib-IX complex in a corresponding bead assay. Further, neither bovine serum albumin nor other adhesive glycoproteins, such as fibrinogen or fibronectin, specifically bound to the GP Ib-IX complex in the presence of ristocetin. Ristocetin-dependent binding of vWF to platelets and to GP Ib-IX complex coated beads was inhibited by monoclonal antibodies against a 45,000 molecular weight N-terminal region of GP Ib but not by monoclonal antibodies directed against other regions of the GP Ib-IX complex. Similar correspondence between platelets and purified GP Ib-IX complex with respect to the ristocetin-dependent binding of vWF was obtained with anti-vWF monoclonal antibodies.(ABSTRACT TRUNCATED AT 250 WORDS)     

Impact of protein binding on the analytical detectability and anticancer activity of thymoquinone

Thymoquinone (TQ), an active component of Nigella sativa L., is known to have anti-cancer and anti-inflammatory effects; however, no studies on its analytical detection in serum and its protein binding have been published. Using high performance liquid chromatography analysis, we show that the average recovery of TQ from serum is 2.5% at 10 μg/ml of TQ and 72% at 100 μg/ml. The low recovery of TQ from serum is due to its extensive binding to plasma proteins, as more than 99% of TQ was bound within 30 min of incubation. The binding of TQ to the major plasma proteins, bovine serum albumin (BSA) and alpha −1 acid glycoprotein (AGP), was studied and found to be 94.5 ± 1.7% for BSA and 99.1 ± 0.1% for AGP. Mass spectrometric analysis revealed that TQ was bound covalently to BSA, specifically on Cyst-34. Using WST-1 proliferation assay, we showed that BSA plays a protective role against TQ-induced cell death; pre-incubation with BSA prevented TQ from exerting its anti-proliferative effects against DLD-1 and HCT-116 human colon cancer cells. On the other hand, binding of TQ to AGP did not alter its anti-proliferative activity against both cell lines. When TQ was pre-incubated with AGP prior to the addition of BSA, the activity of TQ against DLD-1 was maintained, suggesting that AGP prevented the binding of TQ to BSA. This is the first time the covalent binding and inhibitory effect of BSA on TQ is documented. These data offer new grounds for TQ future pharmacokinetic analysis in vivo.     

Interaction of purified actin-binding protein with the platelet membrane glycoprotein Ib-IX complex

The interaction of platelet membrane glycoprotein (GP) Ib-IX complex with the cytoplasmic membrane skeleton is potentially of major importance in regulating platelet function. Indirect evidence suggested that this interaction is mediated by actin-binding protein, but it is not known whether GP Ib-IX and actin-binding protein associate directly. To examine more closely the nature of this association, purified GP Ib-IX complex was specifically bound and oriented on the surface of impermeable polymer beads via a monoclonal antibody, AK 2, directed against the extracytoplasmic domain of GP Ib alpha (glycocalicin). Binding was specific since 1) it was abolished by excess unlabeled actin-binding protein; 2) there was no detectable specific binding of radiolabeled actin-binding protein to beads coated with glycocalicin, the major extracytoplasmic proteolytic fragment of GP Ib alpha; and 3) unlike actin-binding protein, there was no specific binding of bovine serum albumin or human platelet vinculin to the GP Ib-IX complex-coated beads. Binding of actin-binding protein to the GP Ib-IX complex-coated beads, but not to the glycocalicin-coated beads, was saturable and reversible (apparent Kd = 1 x 10(-7) M). These experiments provide direct evidence that actin-binding protein can bind to the cytoplasmic domain of a membrane glycoprotein. Because actin-binding protein is found submembranously in cells other than the platelet, it is possible that this protein may link actin filaments to the plasma membrane in those cells.     

Development of mammalian serum albumin affinity purification media by peptide phage display

Several phage isolates that bind specifically to human serum albumin (HSA) were isolated from disulfide-constrained cyclic peptide phage-display libraries. The majority of corresponding synthetic peptides bind with micromolar affinity to HSA in low salt at pH 6.2, as determined by fluorescence anisotropy. One of the highest affinity peptides, DX-236, also bound well to several mammalian serum albumins (SA). Immobilized DX-236 quantitatively captures HSA from human serum; mild conditions (100 mM Tris, pH 9.1) allow release of HSA. The DX-236 affinity column bound HSA from human serum with a greater specificity than does Cibacron Blue agarose beads. In addition to its likely utility in HSA and other mammalian SA purifications, this peptide media may be useful in the proteomics and medical research markets for selective removal of mammalian albumin from serum prior to mass spectrometric and other analyses.     

Functional binding analysis of human fibrinogen as an iron- and heme-binding protein

Human fibrinogen is a metal ion-binding protein, but its mechanism of binding with iron and heme has not been elucidated in detail. In this study, human fibrinogen was immobilized on CNBr-activated Sepharose 4B beads. The fibrinogen beads bound hemin (iron–protoporphyrin IX: PPIX) as well as iron ion released from ferrous ammonium sulfate (FAS) more efficiently than Sepharose 4B beads alone. Hemin bound to fibrinogen still exhibited pseudo-peroxidase activity. The affinity of fibrinogen binding to hemin, Sn–PPIX, Zn–PPIX and metal-free PPIX followed the order Sn–PPIX < metal-free PPIX < hemin < Zn–PPIX; PPIX bound more non-specifically to control beads. FAS significantly enhanced the binding of hemin to fibrinogen beads. These results suggest that human fibrinogen directly recognizes iron ion, the PPIX ring and metal ions complexed with the PPIX ring, and that the binding of hemin is augmented by iron ions.     

Affinity binding of cells to cryogel adsorbents with immobilized specific ligands: effect of ligand coupling and matrix architecture

The capture of human acute myeloid leukemia KG-1 cells expressing the CD34 surface antigen and the fractionation of human blood lymphocytes were evaluated on polyvinyl alcohol (PVA)-cryogel beads and dimethyl acrylamide (DMAAm) monolithic cryogel with immobilized protein A. The affinity ligand (protein A) was chemically coupled to the reactive PVA-cryogel beads and epoxy-derivatized monolithic cryogels through different immobilization techniques and the binding efficiency of the cell surface receptors specific antibody-labeled cells to the gels/beads was determined. The binding of cells to monolithic cryogel was higher (90-95%) compared with cryogel beads (76%). B-lymphocytes, which bound to the protein A-cryogel beads, were separated from T-lymphocytes with yields for the two cell types 74 and 85%, respectively. About 91% of the bound B-cells could be recovered without significantly impairing their viability. Our results show differences in the percentage of cell-binding to the immunosorbents caused by ligand density, flow shear forces and bond strength between the cells and the affinity surface once distinct chemical coupling of protein A, size of beads, sequence of antibody binding to protein A adsorbents, morphology and geometry of surface matrices were compared.     

Determination of the affinity of vitamin D metabolites to serum vitamin D binding protein using assay employing lipid-coated polystyrene beads.

We have developed an assay to measure the affinity of serum vitamin D binding protein for 25-hydroxyvitamin D3, 1,25-dihydroxyvitamin D3, and vitamin D3, using uniform diameter (6.4 microns) polystyrene beads coated with phosphatidylcholine and vitamin D metabolites as the vitamin D donor. The lipid metabolite coated beads have a solid core, and thus all of the vitamin D metabolites are on the bead surface from which transfer to protein occurs. After incubating these beads in neutral buffer for 3 h, essentially no 3H-labeled vitamin D metabolites desorb from this surface. Phosphatidylcholine/vitamin D metabolite-coated beads (1 microM vitamin D metabolite) were incubated with varying concentrations of serum vitamin D binding protein under conditions in which the bead surfaces were saturated with protein, but most of the protein was free in solution. After incubation, beads were rapidly centrifuged without disturbing the equilibrium of binding and vitamin D metabolite bound to sDBP in solution was assayed in the supernatant. All three vitamin D metabolites became bound to serum vitamin D binding protein, and after 10 min of incubation the transfer of the metabolites to serum vitamin D binding protein was time independent. The transfer followed a Langmuir isotherm, and the Kd for each metabolite binding to serum vitamin D binding protein was derived by nonlinear least-squares fit analysis. From this analysis the following values for the Kd were obtained: 5.59 x 10(-6) M, 25-hydroxyvitamin D; 9.45 x 10(-6) M, 1,25-dihydroxyvitamin D; and 9.17 x 10(-5) M, vitamin D. In conclusion, we have developed a method which avoids problems encountered in previous assays and allows the precise and convenient determination of binding affinities of vitamin D metabolites and serum vitamin D binding protein.     

Surface imprinted beads for the recognition of human serum albumin

The synthesis of poly-aminophenylboronic acid (ABPA) imprinted beads for the recognition of the protein human serum albumin (HSA) is reported. In order to create homogeneous recognition sites, covalent immobilisation of the template HSA was exploited. The resulting imprinted beads were selective for HSA. The indirect imprinting factor (IF) calculated from supernatant was 1.6 and the direct IF, evaluated from the protein recovered from the beads, was 1.9. The binding capacity was 1.4 mg/g, which is comparable to commercially available affinity materials. The specificity of the HSA recognition was evaluated with competitive experiments, indicating a molar ratio 4.5/1 of competitor was necessary to displace half of the bound HSA. The recognition and binding of the imprinted beads was also tested with a complex sample, human serum and targeted removal of HSA without a loss of the other protein components was demonstrated. The easy preparation protocol of derivatised beads and a good protein recognition properties make the approach an attractive solution to analytical and bio-analytical problems in the field of biotechnology.     

Sorting of chromosomes by magnetic separation

Summary Chromosomes were isolated from Chinese hamster x human hybrid cell lines containing four and nine human chromosomes. Human genomic DNA was biotinylated by nick translation and used to label the human chromosomes by in situ hybridization in suspension. Streptavidin was covalently coupled to the surface of magnetic beads and these were incubated with the hybridized chromosomes. The human chromosomes were bound to the magnetic beads through the strong biotin-streptavidin complex and then rapidly separated from nonlabeled Chinese hamster chromosomes by a simple permanent magnet. The hybridization was visualized by additional binding of avidin-FITC (fluorescein) to the unoccupied biotinylated human DNA bound to the human chromosomes. After magnetic separation, up to 98% of the individual chromosomes attached to magnetic beads were classified as human chromosomes by fluorescence microscopy.     

Binding of Streptococcal Lipoteichoic Acid to Fatty Acid-Binding Sites on Human Plasma Fibronectin

The ability of Streptococcus pyogenes lipoteichoic acid and palmitic acid to bind to purified human plasma fibronectin was investigated. Initial studies indicated that intact fibronectin formed soluble complexes with lipoteichoic acid, resulting in a change in the mobility of fibronectin in an electrical field. Fibronectin covalently linked to agarose beads bound radiolabeled lipoteichoic acid in the acylated form but not in the deacylated form. An 18-M excess of fibronectin inhibited binding of lipoteichoic acid to the immobilized protein by 92%. Fibronectin-bound [(3)H]lipoteichoic acid could be specifically eluted with unlabeled lipoteichoic acid, as well as by fatty acid-free serum albumin. Serum albumin, which is known to contain fatty acid-binding sites capable of binding to the lipid moieties of lipoteichoic acid, inhibited the binding of lipoteichoic acid to fibronectin in a competitive fashion. The fibronectin-bound lipoteichoic acid could be eluted by 50% ethanol and various detergents but not by 1.0 M NaCl, various amino acids, or sugars. Similarly, radiolabeled palmitic acid adsorbed to fibronectin could be eluted with 50% ethanol but not with 1.0 M NaCl. Fibronectin adsorbed to a column of palmityl-Sepharose was eluted with 50% ethanol in 0.5% sodium dodecyl sulfate but not with 1.0 M NaCl or 1% sodium dodecyl sulfate alone. The binding of lipoteichoic acid to fibronectin followed first-order kinetics and was saturable. A Scatchard plot analysis of the binding data indicated a heterogeneity of lipoteichoic acid-binding sites similar to that previously found for serum albumin. Nevertheless, fibronectin contains at least one population of high-affinity binding sites for lipoteichoic acid. The binding affinity (nKa approximately 250 muM(-1)) is 2 orders of magnitude greater than the binding affinity of serum albumin. These data suggest that human plasma fibronectin contains specific binding sites for fatty acids and that lipoteichoic acid binds to these sites by way of its glycolipid moiety.     

Gum Arabic coated magnetic nanoparticles with affinity ligands specific for antibodies

A novel magnetic support based on gum Arabic (GA) coated iron oxide magnetic nanoparticles (MNP) has been endowed with affinity properties towards immunoglobulin G (IgG) molecules. The success of the in situ triazine ligand synthesis was confirmed by fluorescence assays. Two synthetic ligands previously developed for binding to IgG, named as ligand 22/8 (artificial Protein A) and ligand 8/7 (artificial Protein L) were immobilized on to MNPs coated with GA (MNP_GA). The dimension of the particles core was not affected by the surface functionalization with GA and triazine ligands. The hydrodynamic diameters of the magnetic supports indicate that the coupling of GA leads to the formation of larger agglomerates of particles with about 1 µm, but the introduction of the triazine ligands leads to a decrease on MNPs size. The non‐functionalized MNP_GA bound 28 mg IgG/g, two times less than bare MNP (60 mg IgG/g). MNP_GA modified with ligand 22/8 bound 133 mg IgG/g support, twice higher than the value obtained for ligand 8/7 magnetic adsorbents (65 mg/g). Supports modified with ligand 22/8 were selected to study the adsorption and the elution of IgG. The adsorption of human IgG on this support followed a Langmuir behavior with a Q_máx of 344 mg IgG/g support and K_a of 1.5 × 10⁵ M. The studies on different elution conditions indicated that although the 0.05 M citrate buffer (pH 3) presented good recovery yields (elution 64% of bound protein), there was occurrence of iron leaching at this acidic pH. Therefore, a potential alternative would be to elute bound protein with a 0.05 M glycine‐NaOH (pH 11) buffer. Copyright © 2010 John Wiley & Sons, Ltd.     

Human alveolar macrophage fibronectin: synthesis, secretion, and ultrastructural localization during gelatin-coated latex particle binding

Human pulmonary alveolar macrophages synthesized and secreted several characteristic high molecular weight proteins for at least 7 d in vitro. Immunoprecipitates of medium and cell lysates from metabolically labeled cultures with specific anti-human plasma fibronectin IgG contained one major labeled polypeptide of molecular weight 440,000 (unreduced) or 220,000 (reduced). An identical polypeptide in conditioned medium from radiolabeled macrophages bound specifically to gelatin-Sepharose, demonstrating that alveolar macrophages synthesized and secreted a molecule immunologically and functionally similar to fibronectin. Fibronectin was the major newly synthesized and secreted polypeptide of freshly harvested alveolar macrophages. Pulse-chase experiments revealed that newly synthesized fibronectin was rapidly secreted into medium, approximately 50 percent appearing by 1 h and 80 percent by 8 h. Immunoperoxidase staining using antifibronectin F(ab’)(2)-peroxidase conjugates revealed the majority of immunoreactive fibronectin to be intracellular, localized to endoplasmic reticulum and Golgi apparatus. No extracellular matrix fibronectin was visualized, and cell surface staining was rarely seen, usually appearing only at sites where cells were closely apposed and not at sites of macrophage-substrate attachment. Similar immunostaining of fibroblast cultures revealed cell surface-associated fibrillar fibronectin. Ultrastructural localization of fibronectin during binding and phagocytosis of gelatin-coated and plain latex particles revealed fibronectin only on gelatin-latex beads and at their cell binding sites. Neigher plain latex beads nor their cell membrane binding sites stained for fibronectin. These results demonstrate that fibronectin is a major product of human alveolar macrophages, is rapidly secreted, and is localized at cell membrane binding sites for gelatin-coated particles. In view of the known binding properties of fibronectin, it may serve as an endogenous opsonic factor promoting the binding of staphylococcus, denatured collagen, fibrin, or other macromolecules to macrophages in the lower respiratory tract.