No abstract

No abstract

The recent success in the separation of amino acids and peptides by filter paper chromatography has led numerous investigators to attempt to separate proteins in an analogous manner. These efforts have thus far not met with definite success. However the possibility of employing filter paper as a framework to support the liquid medium used for the separation of proteins through electrophoresis has proven useful. The observations of Durrum (1), Cremer and Tiselins (2), Wieland (3, 4), and Turba and Enenkel (5) have indicated that the serum proteins would separate on filter paper into components similar to those found in free electrophoresis. Furthermore by cutting the paper into segments and eluting the dye used to stain the separated serum proteins a curve could be obtained which resembled the usual electrophoretic pattern (2).Filter paper electrophoresis in addition to being extremely simple possesses advantages in some respects over the classical methods in free solution. First, absolute separation of components takes place rather than just separation into concentration gradients. Second, smaller amounts of material at lower concentrations can be investigated. Third, isolation of all the components which have been separated is possible. Fourth, the paper support makes it possible to develop techniques of two dimensional electrophoresis and devices for continuous flow preparative work (6, 7). Whether it is possible to achieve the accuracy obtained with the sensitive optical methods employed in free electrophoresis for localizing and quantitating components, is doubtful. In the present report a description is given of the separation and isolation of a large number of different proteins employing a method of paper electrophoresis between glass plates in which disturbing factors such as evaporation, heating, and buffer concentration gradients were reduced to a minimum. An attempt was made to employ the method for determining the mobilities of certain proteins. Material and MethodsIn a previous report from this laboratory (2) a method was described for filter paper electrophoresis in which the paper strip, immersed in buffer and containing the specimen to be analyzed, was clamped between glass plates and immersed under a solution of chlorobenzene and connected to electrode vessels. This apparatus was

Studies on the eleetrophoresis of serum have shown that besides the albumin normal sera possess three separate globulin components differing in mobility and designated as cz, /3, and ~' (1). The relationship of antibodies to these components is of considerable importance. Differences in the antibody globulin formed in various animal species have already been found by ultracentrifugal studies (2, 3) and in electrophoresis (4) as well as by immunological means (5). It has also been shown (1) that in an antiserum to crystalline egg albumin, the antibody migrated with the slowest (3') component and could be isolated in one of the cells of the apparatus and analyzed for antibody content. The work reported in this communication is an attempt to study and compare the electrochemical properties of the antibody in the original sera with the purified antibody preparations described in (6) and to measure the isoelectric points of these purified preparations. It is also of importance to make use of and correlate the results obtained using both the mutually independent electrophoretic and ultracentrifugal methods since molecular weight homogeneity does not necessarily mean electrochemical homogeneity and vice versa. Since the methods previously described for determining the concentration of the various components are not sufficiently precise, especially in systems of several components, the Lamm scale method (7) has been used with the electrophoresis apparatus (8) and has provided a quantitative method of following changes in the various serum components on removal of antibody.

No abstract