Chick embryonic plasma proteins and binding capacity for corticosterone

Plasma was collected from White Leghorn embryos on alternate days from Day 4 of incubation to Day 22 (day after hatch). The plasma-binding capacity for corticosteroid was essentially zero before Day 10, but rose rapidly between Days 10 and 12. Binding capacity remained high until Day 16 and then declined before hatch. The increase after Day 10 was concurrent with the appearance of α-globulin whose rate of migration on acrylamide gel electrophoresis was similar to that of purified chicken corticosteroid binding globulin. Corticosterone, which was present in the plasma of the 4-day embryo, rose to its highest level on Day 20. The relatively high corticosterone concentration, with low plasma-binding capacity, suggests that in the chick embryo levels of free or active corticosterone are highest before Day 10 and just prior to hatching.     

Evaluation of a competitive binding assay for cortisol using horse transcortin

A non-chromatographic competitive binding assay (CBA) using horse transcortin has been employed in the routine measurement of cortisol in plasma, urine and amniotic fluids. Comparing the values with those of a radioimmunoassay (RIA) or a fluorimetric method (FM) an excellent correlation between the three methods both in plasma and urine has been calculated in normal subjects and in patients with various endocrine disorders. In amniotic fluids, however, there were discrepancies between CBA and RIA. Whereas CBA showed no differences, RIA gave significantly higher values in amniotic fluids of female than of male fetuses. Elevated free plasma cortisol levels observed in patients with prostatic cancer after diethyl stilboestrol diphosphate therapy did not correlate with unconjugated urinary cortisol concentration as measured with CBA and FM. In newborns, a relatively high plasma level found 12 hours after birth was followed by a nadir on the 2nd and 3rd day of life and by an increase until levels of adults on the 5th day of life were reached.     

A rapid method for determining dynamic binding capacity of resins for the purification of proteins

Biolayer interferometry is a novel method for quantifying macromolecules, such as proteins, in solution. The presence of other, non-binding molecules does not interfere with quantification, which allows one to measure the concentration of the molecule of interest in a crude mixture. Here we apply this method to determining the dynamic binding capacity of affinity resins.     

Changes in cortisol binding to soluble receptor proteins in rat liver and thymus during development and ageing.

Changes in labeled cortisol binding to soluble cytoplasmic proteins of rat liver and thymus during development and ageing of animals have been observed.The relationship of age and cortisol binding to a fraction of soluble liver proteins precipitated at 50% saturation with ammonium sulfate displayed two maxima: The first one on the seventh day of postnatal life and the second one at 2.5 months of age. Binding of hormone to macromolecules of the same fraction prepared from older animals was less efficient. Almost the same picture has been obtained when total bound activity to macromolecular cytosol fraction was expressed per milligram of dry weight of liver.Binding of labeled cortisol to total cytosol macromolecules of liver and thymus was most efficient in 3-month-old animals and decreased after that, which is of interest in view of totally opposite effects of glucocorticoids upon various biosynthetic processes of these organs.     

The heat capacity paradox of ligand binding proteins: reconciling the microscopic and macroscopic world

Differential scanning microcalorimetry (DSC) is a superb method for the analysis of protein energetics. However, the relative simplicity of application has led astray many to assume that a proper analysis of the data was possible without a sound knowledge of the underlying statistical thermodynamic principles. In this study, the question is addressed of how to calculate properly the heat capacity signal of a protein in the presence of high affinity ligands. It is shown that the signal corresponds neither to grand canonic nor to canonic heat capacity. Statistical thermodynamic model calculations result only in the observed macroscopic heat capacity signal, if the protein in the calorimetric cell is assumed to form a grand canonic ensemble (T, p, mu controlled) which is, however, heated under constraints typical for a canonic ensemble (T, p, N controlled). As a consequence, the microscopic statistical thermodynamic heat capacity must be carefully distinguished from the macroscopically observable thermodynamic heat capacity in those cases where proteins unfold in the presence of high affinity ligands.