Plasma proteins separation

Apart from its natural occurrence, Co may find its way into other proteins either adventitiously or deliberately. A study was undertaken where the blood, serum, and plasma of workers occupationally exposed to Co were analyzed for the element.1189 When separated by gel electrophoresis under denaturing conditions, the Co fractions in all blood, serum, and plasma samples showed a similar protein pattern. A variety of proteins of differing size were found to bind Co in fractions collected at pFl 5, whereas only hemoglobin was found in the pH 7 fractions. The conclusions were that in vivo Co is bound to plasma proteins, perhaps albumin and hemoglobin. 

The overall accuracy of the predictions, assessed as the mean-fold error of prediction of the test set was 2.03, making this approach one that would possess suitable accuracy for use in drug design and human pharmacokinetic predictions. Similar methods developed separately for acids and bases showed an improvement in accuracy. This investigation also included a prediction of unbound VD, which should represent a simpler parameter to predict since it would be based only on tissue binding and not plasma protein binding. However, it is interesting to note that this approach was less accurate for this parameter, which would be unexpected. 

Smithies vertical starch gel electrophoresis (S7) separates the plasma proteins more distinctly than any other method. If the Hp concentration is normal, the Hp type can generally be recognized directly after the staining for proteins, but sensitive and more specific staining for heme groups, e.g., benzidine, o-dianisidine (04), and malachite green (N5) are preferable. This technique consumes more hydrolyzed starch than the simpler original horizontal electrophoresis technique (S5). 

The complement system comprises twenty plasma proteins present in the blood and in most bodily fluids. They are normally present in an inactive form but become activated via two separate pathways the classical pathway, which requires antibody, and the alternative pathway, which does not. Once the initial components of complement are activated, a cascade reac-... 

Size Exclusion Chromatography (Chapter 31) has also been included as a means of analysis for substances that undergo separation more or less as per their molecular size, viz., insulin and human insulin—for proteins of higher molecular weight corticotrophin—for impurities of higher molecular weights and plasma-protein solution—for polymers and aggregates. 

Precipitation by ethanol in the cold was used effectively by J.Mellanby in 1908 to obtain diptheria antitoxin two years later Hardy and Gardiner reported the precipitation of plasma proteins by cold ethanol or acetone. The resulting proteins remained soluble in water, i.e. they were not denatured, and subsequent estimation of protein as nitrogen was helped by the use of nitrogen-free precipitants. Separations using organic solvents were considerably extended by Edsall,... 

Sulfur-containing amino acids such as cystine and homocystine tend to bind to plasma proteins. This binding is irreversible hence, these amino acids will be severely underestimated unless the plasma is deproteinized immediately following its separation from red cells. Blood should be left standing for as short a time as possible to avoid binding of cystine to proteins and hemolysis. 

FIGURE 23-22 The composition of blood. Whole blood can be separated into blood plasma and cells by centrifugation. About 10% of blood plasma is solutes, of which about 10% consists of inorganic salts, 20% small organic molecules, and 70% plasma proteins. The major dissolved components are listed. Blood contains many other substances, often in trace amounts. These include other metabolites, enzymes, hormones, vitamins, trace elements, and bile pigments. Measurements of the concentrations of components in blood plasma are important in the diagnosis and treatment of many diseases. 

Freeman, T. 1970. Techniques for protein separation. In Plasma Protein Metabolism. M.A. Rothschild and T. Waldman (Editors). Academic Press, New York. 

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