Showing papers on "Human serum albumin published in 1991"
TL;DR: A plasmid (palb3) was constructed containing the structural gene for human serum albumin driven by mouse albumin enhancer-rat albumin promoter elements and capable of targeting specifically to hepatocytes via asialoglycoprotein receptors present on these cells.
420 citations
Journal Article•
TL;DR: It is concluded that human serum globulins play a negligible role in acid-base equilibria, and that in normal human serum at pH 7.40 with [TP] = 7 and [Alb] = 4.3 gm/dl, the charges attributed to proteins are substantially less than the value given by many contemporary texts.
376 citations
TL;DR: Protein G was solubilized from 31 human group C and G streptococcal strains with the muralytic enzyme mutanolysin and a peptide corresponding to a single IgG-binding unit was obtained by the cloning and expression of a 303-base pair polymerase chain reaction-generated DNA fragment.
175 citations
TL;DR: The study indicates that chromatography on the silica-immobilized HSA can detect interactions between binding sites on the protein, which can be of great importance in the determination of drug-drug interactions.
125 citations
01 Jan 1991
TL;DR: Conformational stability evaluated through thermal shock and addition of moderate amounts of guanidine indicate that the oxidized protein is more heat-resistant, less flexible, and more rigid than the native one.
Abstract: Mild oxidative stress, as elicited by ascorbate, oxygen, and trace metals, affects the binding properties of human serum albumin via purely conformational changes. In fact, no gross alteration can be observed in the electrophoretic and chromatographic patterns of albumin, whereas localized modifications are indicated by the changes in absorption and fluorescence spectra and in polarization degree. The oxidized protein presents a small increase of bityrosine production and a time-dependent increase in the content of carbonyl groups, whereas proteolytic susceptibility is unchanged. A higher affinity for cis-parinaric acid and a slight loss of solubility in high salt indicate a greater surface hydrophobicity. Pinpoint denaturation of the albumin molecule is also suggested by a decreased “esterase” activity in the presence of p-nitrophenyl acetate. Conformational stability evaluated through thermal shock and addition of moderate amounts of guanidine indicate that the oxidized protein is more heatresistant, less flexible, and more rigid than the native one. Although limited, structural damages afforded by the oxidative stress cause alterations of albumin binding properties as documented by experiments with probes and physiological ligands. The loss of biological activity of human serum albumin induced by ascorbate system appears of medical relevance, because it can affect drug metabolism and particularly drug tolerance in the elderly.
119 citations
TL;DR: In this article, the binding properties of human serum albumin via purely conformational changes are investigated. But no gross alteration can be observed in the electrophoretic and chromatographic patterns of albumin, whereas localized modifications are indicated by the changes in absorption and fluorescence spectra and in polarization degree.
110 citations
TL;DR: In vitro NMR results suggest that the threshold FA/albumin mole ratio value for alterations in FA distributions in the human circulation may be 1, rather than 3, as previously held.
Abstract: A nonperturbing 13C nuclear magnetic resonance (NMR) method was used to monitor the equilibrium distribution of carboxyl 13C-enriched fatty acids (FA) between distinct binding sites on human serum albumin, native human lipoproteins, and/or phospholipid model membranes, under conditions that mimic the normal and diabetic human circulation. Two variables pertinent to the diabetic circulation were examined: FA/albumin mole ratio (as elevated in insulin deficiency and/or nephrosis) and pH (as decreased in acidosis). 13C NMR spectra for samples containing carboxyl 13C-enriched palmitate, human serum albumin, and phospholipid vesicles or native lipoproteins (all samples at pH 7.4, 37 degrees C) exhibited up to six carboxyl NMR resonances corresponding to FA bound to distinct binding sites on albumin and nonalbumin components. When the sample FA/albumin mole ratio was 1, three FA carboxyl resonances were observed (182.2, 181.8, and 181.6 ppm; designated peaks beta, gamma, and beta', respectively). These resonances corresponded to FA bound to three distinct high-affinity binding sites on human serum albumin. When the sample mole ratio value exceeded 1, additional carboxyl resonances corresponding to FA bound to phospholipid vesicles (179.0 ppm, peak phi), lipoproteins (180.7 ppm, peak sigma), and lower affinity sites on albumin (183.8 ppm, peak alpha; 181.9 ppm, peak gamma'), were observed. The intensity of peaks phi and sigma increased with increasing mole ratio or decreasing pH. Using Lorentzian lineshape analysis, the relative mole quantities of FA bound to albumin and nonalbumin binding sites were determined. Plots of the fraction of FA associated with nonalbumin components as a function of FA/albumin mole ratio were linear and extrapolated to the abscissa at a mole ratio value of 1. This pattern of FA distribution was observed regardless of the type of nonalbumin acceptor used (phospholipid vesicles, human high- or low-density lipoproteins) or the type of FA used (palmitate, oleate, or stearate), and provided evidence for negative cooperativity for human serum albumin upon binding of 1 mol of FA per mole albumin. These in vitro NMR results suggest that the threshold FA/albumin mole ratio value for alterations in FA distributions in the human circulation may be 1, rather than 3, as previously held. The pathophysiological implications of these findings are discussed.
94 citations
TL;DR: Preliminary studies show that in addition to being a useful chiral analytical tool for this therapeutically important series of compounds, the HSA chiral stationary phase may provide useful information on the affinity and binding mechanism of small molecules to HSA.
Abstract: The native enantioselectivity in binding of human serum albumin (HSA) towards 2-aryl propionic acid non-steroidal anti-inflammatory drugs (2-APA-NSAIDs, the “profens”) was found to be preserved when the protein was immobilized within a commercially available diol high-performance liquid chromatographic column. High capacity factors were obtained, reflecting the previously observed extensive binding of the 2-APA-NSAIDs to free HSA. The capacity factors were modified by the addition of octanoic acid to the mobile phase. Chiral resolution of the enantiomers of all nine 2-APA-NSAIDs studied was achieved. Preliminary studies show that in addition to being a useful chiral analytical tool for this therapeutically important series of compounds, the HSA chiral stationary phase may provide useful information on the affinity and binding mechanism of small molecules to HSA.
92 citations
TL;DR: Results suggest that, although transferrin at pH 7.4 is the major Al(3+)-binding component of plasma, an appreciable amount of Al3+ present in patients on haemodialysis may be bound to albumin.
Abstract: The binding of Al3+ by human serum transferrin has been investigated by u.v.-visible difference spectroscopy. In the presence of 25 mM-HCO3- at pH 7.4, the apparent association constants were found to be 1.69 x 10(12) M-1 and 5.36 x 10(11) M-1. These association constants are pH-dependent, reducing with both increasing and decreasing pH. The apparent pKa values were found to be 6.7 and 8.2. Competitive assays of binding of Al3+ to transferrin in the presence of citrate and human serum albumin at molar ratios corresponding to those found in normal plasma showed that a considerable amount of Al3+ was not bound to transferrin. Taking a concentration of 5 microM as a typical value observed for the plasma of patients on haemodialysis [Harris & Sheldon (1990) Inorg. Chem. 29, 119-124] the competitive binding assay indicate that approximately 60% of it is bound to transferrin, approximately 34% to albumin and the remainder to citrate. These results therefore suggest that, although transferrin at pH 7.4 is the major Al(3+)-binding component of plasma, an appreciable amount of Al3+ present in patients on haemodialysis may be bound to albumin.
83 citations
TL;DR: The results are consistent with the proposal that the physiologically important high-affinity Zn (II) and Cd(II) binding sites of albumins are located not at the Cu(II)-specific NH2-terminal site, but at internal sites, involving mostly nitrogen and oxygen ligands and no sulphur ligand.
Abstract: The binding of Cd(II) and Zn(II) to human serum albumin (HSA) and dog serum albumin (DSA) has been studied by equilibrium dialysis and 113Cd(II)-NMR techniques at physiological pH. Scatchard analys...
77 citations
TL;DR: Competition experiments showed that when the concentration ratio between total fatty acids and retinol is similar to that found in milk, palmitic acid competes with the binding ofretinol to β-lactoglobulin.
Abstract: The interaction of bovine and human whey proteins with retinol and palmitic acid has been studied. Using gel filtration it was found that bovine β-lactoglobulin and α-lactalbumin and serum albumin from both species bind retinol in vitro while the ability to bind palmitic acid is restricted to bovine β-lactoglobulin and bovine and human serum albumin. Using equilibrium dialysis, β-lactoglobulin was found to display two binding sites for retinol per dimeric molecule with an association constant of 1.5 × 104m-1. Competition experiments showed that when the concentration ratio between total fatty acids and retinol is similar to that found in milk, palmitic acid competes with the binding of retinol to β-lactoglobulin.
TL;DR: In this paper, an electrosynthetic method for direct incorporation of antibodies into conducing polymeric coatings has been developed and the voltammetric behavior of an anti-human serum albumin (anti-HSA) containing electrode has been considered.
TL;DR: Analysis of the data suggests that the reductions in binding represent competition for a common high-affinity binding site within the albumin molecule and a region seems to exist that is capable of binding one molecule of these diverse ligands with a high affinity.
Abstract: Binding of L-tryptophan, diazepam and octanoate to defatted human serum albumin was studied at pH 7.0 by equilibrium dialysis at low ligand/protein molar ratios. L-Tryptophan binding takes place at only one site of the protein with an association constant of 4.4 x 10(4) M-1. Under the present experimental conditions, binding of diazepam and octanoate could be accounted for by high-affinity binding alone with primary association constants of 3.8 x 10(5) M-1 and 1.6 x 10(6) M-1 respectively. During the simultaneous presence of L-tryptophan plus octanoate or diazepam plus octanoate, pronounced mutual reductions in binding were observed. Analysis of the data suggests that the reductions in binding represent competition for a common high-affinity binding site. Thus a region seems to exist that is capable of binding one molecule of these diverse ligands with a high affinity. The location of this region within the albumin molecule is discussed.
TL;DR: Examples are presented of the application of Divicell derivatives to the purification of biomacromolecules such as immunoglobulins and lectins by affinity chromatography and the results were comparable to those obtained using the corresponding Sepharose-derived absorbents.
TL;DR: It was concluded that chondroitin sulphate, human serum albumin and Tamm-Horsfall mucoprotein cannot account for the inhibitory effects of macromolecules with a relative mass greater than 10 kDa in spun and filtered urine.
Abstract: The effects of physiological concentrations of chondroitin sulphate, human serum albumin and Tamm-Horsfall mucoprotein on the crystallization of calcium oxalate in undiluted, ultrafiltered human urine were investigated using particle size analysis and scanning electron microscopy. Neither the amount of oxalate required to induce detectable calcium oxalate crystal nucleation nor crystal morphology was affected by the presence of any of these macromolecules. Chondroitin sulphate had no effect on the amount of crystalline material deposited or on the size of the particles precipitated in response to a standard oxalate load. Human serum albumin slightly reduced the size of the crystal aggregates and caused a small increase in the amount of crystal matter precipitated. By contrast, Tamm-Horsfall mucoprotein significantly inhibited crystal aggregation and markedly increased the volume of matter deposited, although this could not be attributed to a promotion of solute precipitation. It was concluded that chondroitin sulphate, human serum albumin and Tamm-Horsfall mucoprotein cannot account for the inhibitory effects of macromolecules with a relative mass greater than 10 kDa in spun and filtered urine. Nonetheless, Tamm-Horsfall mucoprotein is likely to inhibit crystal aggregation in whole urine in vivo and may therefore be instrumental in preventing calcium oxalate stone formation.
Journal Article•
TL;DR: Analysis of plasma samples from a clinical study showed that enantioselective irreversible binding of FEN to plasma proteins also occurs in vivo, and covalent binding of R- and S-FEN to Plasma proteins was measured in all subjects, indicating an esterase-like activity of the albumin molecule.
Abstract: Stereoselective degradation of fenoprofen (FEN) glucuronides and irreversible binding of FEN enantiomers to human serum albumin via their glucuronides were studied. At different pH values, 37 degrees C, and in the absence of albumin, degradation half-lives were diastereomeric, resulting mainly from a combination of hydrolysis and acyl migration. Lower pH enhanced FEN glucuronide stability and reduced the extent of irreversible binding. The degradation rate of R-FEN glucuronide was greater than that of the S-glucuronide (S-FEN). When human serum albumin was added to the medium, stability was decreased as compared to protein-free buffer. FEN glucuronides were readily hydrolyzed to parent drug, indicating an esterase-like activity of the albumin molecule. In vitro irreversible binding was higher for R-FEN (1.22% +/- 0.36) than for S-FEN glucuronide (0.76% +/- 0.12), when a 0.1 mM concentration of each conjugate enantiomer was incubated under physiological conditions (pH 7.4, 37 degrees C). Incubation with unconjugated FEN did not lead to measurable irreversible binding. Analysis of plasma samples from a clinical study showed that enantioselective irreversible binding of FEN to plasma proteins also occurs in vivo. After administration of a single 600-mg dose of racemic FEN to six healthy volunteers, covalent binding of R- and S-FEN to plasma proteins was measured in all subjects. The percentage of S-FEN protein adduct was greater than that of its R-enantiomer adduct. Total amounts of FEN irreversibly bound to plasma protein in vivo were also very low (1.02 +/- 0.32 and 3.23 +/- 0.85 mol/mol protein x 10(-4) for R- and S-FEN, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)
Journal Article•
TL;DR: It is concluded that albumins with an increased negative charge are potent and nontoxic anti-HIV-1 agents and interfere with a post-binding event, of which one of the potential mechanisms is an interaction with the gp41 fusion protein, which is necessary for syncytium formation but is not involved in initial virus binding.
Abstract: A series of neoglycoproteins was synthesized by coupling of thiophosgene-activated p-aminophenyl derivatives [Biol. Cell. 47:95-110 (1983); J. Histochem. Cytochem. 32:1091-1094 (1984)] of various sugars to human serum albumin. The compounds were evaluated for their in vitro activity against human immunodeficiency virus (HIV). Neoglycoproteins with the highest sugar content were found to be the most potent inhibitors of HIV-1-induced cytopathogenicity. However, this was not due to the nature of the sugar used but, rather, was related to the extra negative charge of the neoglycoproteins. To investigate whether the antiviral activity of the neoglycoproteins exhibited sugar specificity, increased with increasing negative charge, or depended on both sugar specificity and negative charge, we synthesized albumins and neoglycoproteins with an enhanced negative charge, by treatment with formaldehyde or succinic anhydride. Succinylated human serum albumin had the most pronounced net negative charge and had an IC50 of about 1 microgram/ml. No cytotoxicity was observed at concentrations up to 1 mg/ml, implicating a selectivity index (CC50/IC50) of at least 10(3). To elucidate the mechanism of action of these anionic albumins, we investigated whether they interfered with HIV-1 adsorption to the cells, binding of anti-OKT4A monoclonal antibody (mAb) to the CD4 receptor, binding of anti-gp120 mAb to gp120, or inhibition of syncytium formation in co-cultures of HIV-1-infected HUT-78 cells with MOLT-4 cells. From these experiments, we conclude that albumins with an increased negative charge (a) are potent and nontoxic anti-HIV-1 agents, (b) cause a 50% reduction of syncytium formation in the same concentration range as their IC50 in the antiviral assay, and (c) do not bind to the OKT4A epitope of the CD4 receptor and only partly inhibit anti-gp120 mAb-gp120 interaction and virus-cell binding at concentrations that are 100 times higher than their IC50 in the antiviral assay. Therefore, we conclude that the modified albumins interfere with a post-binding event, of which one of the potential mechanisms is an interaction with the gp41 fusion protein, which is necessary for syncytium formation but is not involved in initial virus binding.
TL;DR: The results indicate that the rotational correlation times for the internal motions and the overall protein rotation in reverse micelles decrease with increasing water concentration.
Abstract: Lysozyme, human serum albumin (HSA), and liver alcohol dehydrogenase (LADH) have been studied in reverse micelles by frequency domain fluorescence spectroscopy. The emission of the tryptophanyl residues of the proteins was monitored. Fluorescence and anisotropy decays were measured from 2 to 350 MHz for each protein in reverse micelles and in aqueous solutions. The wide range of modulation frequencies available allowed direct monitoring of the internal motions of tryptophan residues, occurring in the subnanosecond time range, together with the whole protein rotational dynamics in the micelles. The results indicate that the rotational correlation times for the internal motions and the overall protein rotation in reverse micelles decrease with increasing water concentration. Lysozymes showed peculiar rotational dynamics which reflect denaturation occurring as the protein increases its water content in the reverse micelle. This effect was not observed for the other proteins. Dynamic measurements appear useful in understanding structural changes arising from the interactions between proteins and micellar systems. © 1991 American Chemical Society.
TL;DR: DAG was more labile to hydrolysis than any of its isomers but the latter, in particular the 4-isomer, were much better substrates for formation of covalent DF-HSA adducts.
TL;DR: A novel support based on poly(vinyl alcohol) carrying a blood group-specific trisaccharide ligand for the removal of blood group antibodies from human serum exhibits excellent antibody binding capacities when compared with supports carrying protein G and protein A ligands.
TL;DR: It is concluded that minor effects of glutamate toxicity could be considerably aggravated by the potentiating activity of serum albumin, also in pathological situations in vivo.
TL;DR: The periodate assay for glycated protein has been adapted for use with a microplate reader, allowing in vitro glycation of human serum albumin to be assayed on 0.1-mg aliquots.
TL;DR: HEC adhesion and spreading on TCPS resulted in a significant deposition of fibronectin irrespective of the serum concentration in the solution used for the pretreatment of TCPS, and this may explain why HEC normally fail to adhere on to prosthetic surfaces.
Abstract: More insight into the mechanism of adhesion of human endothelial cells (HEC) on to polymeric surfaces may lead to the development of improved small-diameter vascular grafts HEC suspended in 20% human serum-containing culture medium adhere and spread well on moderately water-wettable polymers such as tissue culture polystyrene (TCPS) Earlier it was demonstrated that during adhesion and spreading of HEC on TCPS, cellular fibronectin is deposited on to this surface It was postulated that fibronectin deposition is accompanied by desorption of adsorbed serum proteins, eg human serum albumin (HSA) The amounts of adsorbed (cellular) fibronectin and HSA on TCPS surfaces pretreated for 1 h with solutions of human serum (ranging from 001%–20%), were determined after incubation of these surfaces for 6 h with HEC in culture medium and after incubation with culture medium without cells Protein adsorption was determined by means of a two-step enzyme-immunoassay (EIA) HEC adhesion and spreading on TCPS resulted in a significant deposition of fibronectin irrespective of the serum concentration in the solution used for the pretreatment of TCPS The deposition of cellular fibronectin on to TCPS, pretreated with human serum, was accompanied by displacement of adsorbed HSA Desorption of HSA from TCPS was only detectable with the EIA at serum concentrations ranging from 001%–1% Using131-l-labelled HSA as tracer protein; it could, however, be demonstrated that HSA was also displaced from TCPS, pretreated with solutions of higher serum concentrations Pretreatment of the hydrophobic vascular graft material PET (poly(ethylene terephthalate); Dacron) and of FEP (fluoroethylenepropylene copolymer; a Teflon-like polymer) with a solution containing 20% human serum resulted in a reduced adhesion of HEC compared to uncoated surfaces We suggest that this may be caused by a poor displacement of adsorbed serum proteins from these hydrophobic surfaces by cellular fibronectin This may explain why HEC normally fail to adhere on to prosthetic surfaces
TL;DR: The decrease in negative charge observed on incubation with sialidase suggested the presence of a carbohydrate moiety and the normalization of molecular weight following treatment with Endo-F indicated that this was an N-linked oligosaccharide.
TL;DR: A rapid and sensitive screening sandwich enzyme-linked immunosorbent assay (ELISA) was developed for the detection of staphylococcal enterotoxin B in cheese by using a highly avid anti-SEB antibody (Ab) as the capture Ab (CAb) and as the biotinylated Ab conjugate.
Abstract: A rapid and sensitive screening sandwich enzyme-linked immunosorbent assay (ELISA) was developed for the detection of staphylococcal enterotoxin B (SEB) in cheese by using a highly avid anti-SEB antibody (Ab) as the capture Ab (CAb) and as the biotinylated Ab conjugate. The glutaraldehyde fixation method for the immobilization of CAb on polystyrene dipsticks was superior to the adsorption fixation and the adsorption-glutaraldehyde fixation methods. The glutaraldehyde fixation method resulted in a higher surface-saturating CAb concentration as evaluated by the peroxidase saturation technique and by the ability of the CAb-coated dipstick to discriminate between positive and negative controls (index of discrimination). Of nine blocking agents used alone or in pairs, lysine-human serum albumin, bovine serum albumin, human serum albumin, and gelatin effectively saturated available sites on the CAb-coated dipsticks without causing interference with the antigen-Ab reactions. The addition of 1% polyethylene glycol to the diluent of the biotinylated anti-SEB Ab conjugate improved the detection of SEB. A concentration of 4% polyethylene glycol allowed a 5-min reaction time for the streptavidin-biotin-horseradish peroxidase conjugate. Cheddar cheese homogenate reduced the sensitivity of the SEB assay; however, the sensitivity was restored when 1.6% (wt/vol) of either a nonionic detergent (Mega-9) or two zwitterionic detergents (Zwittergent 3-10 and 3-12 detergent) was added to the diluent. By using the rapid sandwich ELISA, a minimum of 0.5 to 1.0 ng of SEB per ml was detected within 45 min. The whole procedure for the analysis of the cheddar cheese samples was completed within 1 h.(ABSTRACT TRUNCATED AT 250 WORDS)
TL;DR: Cis-DDP bound to multiple sites in addition to the lone sulfhydryl group (Cys-34), suggesting that massive conformational changes of the protein took place.
Abstract: Irreversible bindings of cis-diamminedichloroplatinum(II) (cis-DDP) to human serum albumin (HSA) were investigated in a pH 7.4 buffer containing 0.1 M NaCl at various molar ratios (cis-DDP/HSA) up to 60 over a 14 d period (37°C). The metal binding seemed to reach a plateau when incubated at less than 10 times excess of cis-DDP. As the molar ration increased, the reaction rate was relatively fast within the first day, followed by a moderate increase in the metal binding. When incubated at 60 times excess of cis-DDP, the metal bound as mush as 20 mol per mol of HSA in 14 d. Fluorescence quenching of the metal-bound protein suggested that the tryptophan residue was gradually exposed to a hydrophilic environment as the metal binding increased. Furthermore, cis-DDP cleaved disulfide bonds at the ratio of 1 mol of disulfide bond per 5.3 mol of the metal binding. It was therefore suggested that the metal binding also occurred at several sites other than the disulfide bond. Warfarin binding to the metal-bound protein, examined by fluorescence changes, also decreased with increasing metal binding or cleavage of the disulfide bonds. Thus, cis-DDP bound to multiple sites in addition to the lone sulfhydryl group (Cys-34), suggesting that massive conformational changes of the protein took place.
TL;DR: Citric acid was shown to have no significant effect, compared with water alone and is unlikely to be the component in B. pinnatum extracts that are responsible for reducing the size of calcium oxalate calculi.
Abstract: www.PosterPresentations.com Calcium oxalate renal calculi are one of the most common types of kidney stones. Research has shown that citric acid can prevent calcium oxalate crystallization and that extracts of Bryophyllum pinnatum may be able to reduce the size of existing calcium oxalate crystals in vivo. Bryophyllum pinnatum extracts contain a high concentration of citric acid, which may contribute to its ability to dissolve renal calculi. The purpose of this study was to determine the efficacy of citric acid for dissolving calcium oxalate. Calcium oxalate powder, contained in dialysis tubing, was incubated in solutions of water, citric acid or EDTA for ten days and the change in mass was measured. Citric acid was shown to have no significant effect, compared with water alone and is unlikely to be the component in B. pinnatum extracts that are responsible for reducing the size of calcium oxalate calculi. Abstract
01 Jan 1991-International Journal of Radiation Applications and Instrumentation. Part A. Applied Radiation and Isotopes
TL;DR: The binding of the 45Ti with plasma transferrin in vitro and in vivo suggested that these compounds can be used for estimating the blood volume, and potential as an indicator representing the breakdown of the blood-brain barrier in the rat was demonstrated by autoradiography.
TL;DR: The polymer type appeared to be the primary determinant of platelet shape-change with adsorbed albumin producing a secondary effect, and somewhat different effects on spreading were observed with HSA and BSA, suggesting qualitatively different interactions between human platelets and HSA, than with B SA, which is commonly used in platelet preparations.
Abstract: Adsorbed albumin appears to passivate nearly all materials, minimizing platelet adhesion and thrombus formation. Since in vitro platelet spreading can be an indicator of in vivo reactivity leading to thrombosis, and as in vitro platelet adhesion investigations are routinely done in the presence of bovine or human serum albumin (BSA or HSA), we examined the influence of albumin on platelet reactivity to material substrates. Platelet spreading was examined subsequent to adherence onto several related polyurethanes, and to Formvar, in the presence of bulk albumin concentrations sufficient to form an adsorbed monolayer or a multilayer. No other exogenous proteins were present. The spreading behavior of adherent platelets was analyzed using generalized linear interactive modeling (GLIM). The models showed that the polymer type always influenced platelet responses, irrespective of the albumin concentration. In many experiments, platelet behavior could be adequately modeled without including the effects of album...
TL;DR: Covalent conjugates of AFP and Tf with horseradish peroxidase (HRP) were used to follow the endocytosis and intracellular pathway of these serum proteins by electron microscopy and suggested the presence of 3 types of receptor site with different affinities and capacities on cells of both lines.
Abstract: The kinetics of iodinated human alpha-fetoprotein (AFP) binding and uptake by 2 human neoplastic lymphoid cell lines (CEM and RAJI) have been studied. Three saturation plateaus were obtained by incubating CEM and RAJI cells at 4 degrees C with 125I-AFP at different concentrations. Scatchard analysis suggested the presence of 3 types of receptor site with different affinities and capacities on cells of both lines. AFP binding was inhibited by unlabelled human and bovine AFP, and to a lesser extent by human serum albumin (SAH); no significant competition was observed with human transferrin (Tf) or ovalbumin (Ova). Pulse-chase experiments showed that 125I-AFP was released practically undegraded from the cells. Covalent conjugates of AFP and Tf with horseradish peroxidase (HRP) were used to follow the endocytosis and intracellular pathway of these serum proteins by electron microscopy. Both proteins were observed in coated vesicles, endosomes and a tubular vesicular network localized in the Golgi-centrosphere region. SAH-HRP was internalized to a much lesser extent. Ova-HRP was poorly internalized and was observed in lysosome-like organelles.