Globulins, serum separation

Figure 25 shows the result of ImEl of a protein mixture of seven isolated and highly purified serum protein fractions. Heide et al. (H5) controlled the purity of a y-globulin (sao = 7) with a low carbohydrate content. The group of y-globulins was separated beforehand by zone electrophoresis in polyvinyl chloride the Longsworth buffer had a pH of 8.6 t/2 = 0.1, and the field strength was 4.4 volts/cm. 

The best results in the starch block separation are achieved with 2 cc. or less of serum. However, fair separation can be obtained with a quantity up to 5 cc. Above this the band of origin becomes too broad and thicker blocks are necessary. In general, blocks more than 2 cm. in thickness have proved unsatisfactory, due in part to poorer dissipation of heat and increased gravity effects. Distortion of the bands with a marked curvature in the albumin component may be observed. The best results have been obtained with serum diluted with buffer to two-thirds concentration or less viscosity effects are prominent when whole serum is employed. The electroosmotic flow of water is only slightly greater than that observed with the usual filter papers. In a serum separation, most of the y-globulin falls behind the origin on the cathode side at pH 8.6. A wide application of the starch system for small-scale preparative electrophoresis, particularly of serum, has recently developed. Early observations indicated a... 

The third test of protein homogeneity, developments from which remain in common use, was that of electrophoresis. Arne Tiselius had been a research assistant in Svedberg s laboratory. From 1925 he pioneered the application of electrophoresis to the analysis and separation of protein mixtures, showing with dialyzed serum differences in mobility of the protein components and the presence of three classes of globulins, a, B, and y. 

The characteristic hypergammaglobulinemia of tropical and subtropical populations have been chiefly ascribed to malarial infection up until the 1950 s quantitation of the y-globulin in the serum of patients was wholly by the electrophoretic method, which does not separate the y-globulin into its different immunoglobulin components (D5, T2). 

Immunodeficiency 100-200 mg/kg/mo IV at 0.01-0.04 mL/kg/min to 400 mg/kg/dose max UP 400 mg/kg/dose IV daily x 5 d BMP 500 mg/kg/wk X in renal insuff Caution [C, ] Separate administration of live vaccines by 3 mo Contra IgA deficiency w/ Abs to IgA, severe thrombocytopenia or coagulation disorders Disp Inj SE Associated mostly w/ inf rate GI upset Interactions X Effects OF live virus vaccines EMS May cause anaphylactic Rxn OD Unlikely Immune Globulin, Subcutaneous (Vivaglobin) [Immune Serum] Uses Primary immunodeficiency Action IgG supl Dose 100—200 mg/kg BW subq wkly abd, thighs, upp arms, or lat al hip Caution [C, ] Contra Hx anaphylaxis to immune globulin some IGA deficiency Disp Inj SE Inj site Rxns, HA, GI complaint, fevCT, N, D, rash, sore throat EMS May be self administered at home may cause anaphylactic Rxn OD Unlikely to cause life-threatening Sxs... 

Zone electrophoresis is mostly used for biological applications. Peptide separation and the measurement of protein fractions from blood serum (proteinogram of albumin and o-, (3- and 7-globulins) are among the better known applications. This TLC for biochemists is useful for the separation of polysaccharides, nucleic acids (for DNA sequencing), proteins and other colloidal species. 

In a follow-up study, the same authors examined the applicability of the same device for relevant protein samples and investigated the main contributions to band broadening [82]. As a consequence of the small depth of the beds, zone spreading caused by Joule heating was shown to be negligible (see Sect. 3.1.1). Cross fields of up to 100 V/cm were applied for the separation of human serum albumin, ribonuclease A and bradykinin. The feasibility of fraction collection was demonstrated with four collected fractions of a whole rat plasma sample. Off-line analysis of these four isolated fractions by CE indicated the separation of serum albumins and globulins. 

El-FFF is a technique devoted to the fractionation of proteins which is reflected in the number of papers applying this technique to protein separations. The possibilities of El-FFF were first demonstrated by Caldwell for the separation of albumin, lysozyme, hemoglobin, and y-globulin in two different buffer solutions (pH 4.5 and 8.0) [35]. Later, the performance of an El-FFF channel with flexible membranes [36], a channel with rigid membranes [256], or a circular channel [260] for the separation of proteins were described. In these studies, human and bovine serum albumin, y-globulin (bovine), cytochrome C (horse heart), lysozyme (egg white) and soluble ribonucleic acid (t-RNA), as well as denaturated proteins, were successfully separated. 

A great number of proteins have been purified on Sephadex G-75 (Bjork and Porath, 1959 Hanson and Johansson, 1960 Pr aux and Lontie, 1961 Bjork, 1961 Palmstierna, 1961 Ames et al., 1961 Pettersson et al., 1962). Considerably improved gel types have been introduced recently thanks to pioneering work by Flodin. These gel substances, named Sephadex G-lOO and G-200, are produced in the form of spherical beads. They can be packed in beds with excellent filtering properties this is particularly true for G-100. The behavior of serum proteins has been studied by Flodin and Killander (1962). As is seen in Fig. 6, 7 S and 19 S 7-globulins can be separated on Sephadex G-200. 

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