Function Of Albumin Concentration Research Articles

Measurement of free and bound fractions of extracellular ATP in biological solutions using bioluminescence

Measurement of extracellular ATP in biological solutions is complicated by protein-binding and rapid enzymatic degradation. We hypothesized that the concentration of extracellular ATP could be determined luminometrically by limiting degradation and measuring the free and protein-bound fractions. ATP was added (a) at constant concentration to solutions containing varying albumin concentrations; (b) at varying concentrations to a physiological albumin solution (4 gm/dL); (c) at varying concentrations to plasma. After centrifugation, a fraction of each supernatant was heated. ATP in heated and unheated samples was measured luminometrically. Blood was drawn into saline or an ATP-stabilizing solution and endogenous plasma ATP measured. ATP-albumin binding was a linear function of albumin concentration (3.5% ATP bound at 100 micromol/L to 33.2% ATP bound at 1000 micromol/L) but independent of ATP concentration (29.3%, 10-1000 nmol/L ATP in 602 micromol/L albumin). Heating released the majority of bound ATP from albumin-containing solutions (94.8 +/- 1.7%) and plasma (97.6 +/- 5.1%). Total endogenous plasma ATP comprised 93 +/- 27 nmol/L (free) and 150 +/- 40 nmol/L (total fraction). Without stabilizing solution, degradation of free endogenous plasma ATP occurred. Within a physiological range (10-1000 nmol/L), ATP binds albumin independently of ATP concentration. Heating releases bound ATP, enabling accurate luminometric measurement of total extracellular ATP (free and bound) in biological samples.

Penetration and loading of human serum albumin in porous silicon layers with different pore sizes and thicknesses

Human serum albumin was adsorbed into porous silicon layers with thickness up to 3 μm and with different mean pore radius in the range 4.5–10 nm. The adsorbed amount of protein was quantified by I 125 radioactive labeling techniques and ellipsometry. The results show that albumin penetrated into the pores when the mean pore radius was larger than 5.5 nm, but could not totally occupy the available surface area when the layer thickness was larger than 1 μm. Loading of albumin both into porous layers and onto plane silicon as a function of albumin concentration was also investigated. These measurements show that loading of protein increased with protein concentration at least up to 10 mg/ml for porous silicon and up to 1 mg/ml for plane silicon. The maximum deposition into the type of porous layers used here was 28 μg/cm 2, compared to 0.36 μg/cm 2 for plane silicon.

Conformation Change of Albumin Adsorbed on Polycarbonate Membranes as Revealed by ToF-SIMS

The adsorption of proteins at biomaterial surfaces depends on the properties of the substrate and can cause changes in protein conformation. Time-of-flight secondary ion mass spectroscopy (ToF-SIMS) was used in this study to characterize human serum albumin (HSA) adsorption on two different polycarbonate surfaces: a native membrane and a hydrophilic treated one. The amount adsorbed as a function of HSA concentration in solution was compared for the two substrates. The treated membrane was found to have a lower affinity for albumin than the native one. Principal component analysis was used to reveal changes in albumin conformation as a function of albumin concentration in solution and to compare the conformations adopted on the two substrates. The albumin conformation was different on the two substrates, and in every case, the protein lost its native structure. A correlation was found between the amount adsorbed on the hydrophilic surface and the albumin conformation on this surface.

Fluid-attenuated inversion recovery MR imaging: identification of protein concentration thresholds for CSF hyperintensity.

Using human volunteers and phantoms emulating CSF, we analyzed the effects of varying protein concentration on the signal intensity of saline solution. Also, for different fluid-attenuated inversion recovery (FLAIR) sequences, we compared protein concentration thresholds above which the signal from these solutions becomes hyperintense to that from brain parenchyma. Nine albumin solutions of varying concentrations (3.9 mg/dl to 2500 mg/dl) were imaged using fast FLAIR MR sequences (TR, 6000 msec: inversion time, 1730 msec: echo train length, 20) at different effective TEs (110, 150, 200, and 250 signal-to-noise ratios from the different albumin solutions versus albumin concentration were generated and correlated with average signal-to-noise ratios from brain parenchyma and CSF. We saw a gradual increase in signal-to-noise ratios from the albumin solutions as a function of albumin concentration. As the effective TE increased, the point of intersection between the plots and the average signal-to-noise ratio from brain parenchyma occurred at lower albumin concentrations. FLAIR MR imaging is potentially useful to evaluate pathologic conditions that increase CSF protein concentration. Using phantoms and healthy volunteers, we defined a protein concentration threshold above which the signal from saline solutions becomes hyperintense to that from brain parenchyma. This threshold depends on the effective TE used in the FLAIR sequence and is 250 mg/dl for an effective TE of 110 msec, 125 mg/dl for 150 msec. 110 mg/dl for 200 msec, and 95 mg/dl for 250 msec.

Calibration of Oxygen-Dependent Quenching of the Phosphorescence of Pd-meso-tetra (4-Carboxyphenyl) Porphine: A Phosphor with General Application for Measuring Oxygen Concentration in Biological Systems

Oxygen-dependent quenching of phosphorescence is a function of the frequency of collision between the phosphor and molecular oxygen and of the efficiency of energy transfer during these collisions. Thus, quenching is dependent on the rate of diffusion of the phosphor and its molecular environment. For measurements in biological samples, the Pd-porphyrin is bound to serum albumin, and this provides a uniform microenvironment for the phosphor which is relatively unaffected by changes in the pH and ionic composition of the medium. Calibration of the phosphor is of particular value because it is absolute, i.e., the calibration is valid independent of the laboratory and the time of measurement. This paper reports the calibration constants determined for Pd-meso-tetra (4-carboxyphenyl) porphine, as measured by two independent methods: by stoichiometric titration of the oxygen with ascorbate in the presence of ascorbate oxidase and by comparison with a high-accuracy oxygen electrode. The measurements were carried out in a specially designed thermostatted vessel in which the oxygen electrode and phosphorescence lifetime measurements of oxygen were made simultaneously. The calibration constants for the oxygen-dependent quenching of the phosphorescence of Pd-meso-tetra (4-carboxyphenyl) porphine were determined as a function of albumin concentration, ionic strength in medium, pH, and temperature.

Intestinal hydrolysis of pyridoxal 5′-phosphate in vitro and in vivo in the rat: Effect of protein binding and pH

Phosphatase-mediated hydrolysis is the first step in the intestinal absorption of pyridoxal 5′-phosphate (PLP). Studies presented here evaluated the effects of PLP-protein binding and pH on intestinal hydrolysis in the rat. Models included in vitro PLP decay and in vivo PLP disappearance from perfused jejunal segments. Pyridoxal 5′-phosphate binding to albumin was pH-, PLP concentration-, and albumin concentration-dependent. Binding occurred at higher pH levels (5–7.4) and markedly inhibited hydrolysis of 2 μM PLP both in vitro and in vivo. At low pH (3–4), alkaline phosphatase was still active with PLP as substrate; binding of PLP to albumin was negligible; and PLP hydrolysis was unaffected by even a high concentration of albumin. The pH required to prevent binding and thus in turn prevent inhibition of hydrolysis was a function of albumin concentration: the higher the concentration, the lower the pH necessary. The present studies suggest an important role for the low pH values resulting from gastric acid secretion in the normal absorption of dietary PLP, and raise questions about the implications of widespread use of acid-lowering therapeutic modalities on vitamin B6 nutritional status.

Protein A-bearing Staphylococcus aureus as the solid phase in an enzyme immunoassay and its application to determination of urinary albumin.

Protein A-bearing Staphylococcus aureus was used as a solid-phase matrix in a sandwich-type enzyme immunoassay for urinary albumin. Heat-inactivated, formalin-fixed bacteria were coated with affinity-purified goat anti-human albumin, exposed to solutions containing standard or unknown concentrations of albumin, then challenged with an alkaline phosphatase/anti-human albumin conjugate obtained by periodate oxidation. Alkaline phosphatase activity bound to the bacteria was a function of albumin concentration from 25 to 1000 micrograms/L. This assay was applied to determinations of urinary albumin concentrations between 1.25 and 1000 mg/L. Between-run CV was 2.55 (63.9 mg/L concentration). Within-run CVs for albumin concentrations of 1.9, 38.1, and 638.0 mg/L were 3.7, 3.7, and 2.4%, respectively. Analytical recovery was 95 to 107% across the full working range of the assay. Bence Jones proteins and hemoglobin had no significant effect on the assay. Nonspecific binding of the enzyme-antibody conjugate was 1.3% (SD = 0.7%). Values agreed well with those by radial immunodiffusion.

Pulmonary disposition of cortisol.

High-dose corticosteroid therapy is used in various lung diseases, yet it is not known if the drug enters the alveolar acinus. Cortisol levels, expressed as a function of albumin concentration, were measured in serum and bronchoalveolar lavage fluid in 12 patients with lung disease. There was a linear relation of cortisol concentration between blood level and bronchoalveolar lavage fluid level when expressed in relation to the albumin concentration (r = 0.93, p less than 0.001). Incremental doses of intravenous hydrocortisone reach alveolar structures in a concentration directly related to the circulating blood levels.