Immunoelectrophoresis quantitative

FIG. 12.15 Crossed immunoelectrophoresis of human serum with rabbit antihuman serum. (Redrawn with permission from B. Weeke, in A Manual of Quantitative Immunoelectrophoresis Methods and Applications (N. H. Axelsen, J. Kroll, and B. Weeke, Eds.), Universitetsforlaget, Oslo, Norway, 1973.)... 

N. H. Axelsen, J. KroII, and B. Weeke, Eds., A Manual of Quantitative Immunoelectrophoresis, BioRad Laboratories, Richmond, Calif. 

H. G. Minchin and T. Freeman, Clin. Sci, 35 403 (1968). Quantitative Immunoelectrophoresis of Human Serum Proteins. 

The rocket technique or electroimmunodiffusion (EID), which allows a precise quantitation of antigen, is described in Chapter 21, the crossed immunoelectrophoresis (CIE), which is a qualitative and quantitative method, is dealt with in Chapter 22, and the crossed immunoaffinoelectrophoresis (CIAE), which is more specially designed for the detection of biospecific... 

Clarke H. G. M. and Freeman T. (1967) Quantitative Immunoelectrophoresis method (Laurell electrophoresis), in ProtieUs of the Biological Fluids, 14th Colloquium, 1966 (Peelers, H, ed ), Elsevier, Amsterdam, pp. 503-509. 

Emmett, M. and Crowle, A. J. (1982) Crossed Immunoelectrophoresis Qualitative and quantitative considerations./. ImmunoL Methods 50, R65-R83. 

Brandshund I, Siersted SE, Teisner B. Double-decker rocket immunoelectrophoresis for direct quantitation of complement split products with C3d specificities in plasma. J Immunol Methods 1981 44 63-71. 

Assays for insulin antibodies fall into three categories (1) quantitative radioimmunoelectrophoresis, which measures the binding of IgG antibody to radiolabeled insulin by rocket Immunoelectrophoresis into anti-IgG-containing agarose (2) RIAs with separation of bound and free insulin by precipitation with PEG or a second antibody and (3) solid phase immobilization of insulin to test tubes or Sepharose. These are discussed in more detail in Reeves. ... 

Kll. Kroll, J., Immunochemical identification of specific precipitin lines in quantitative immunoelectrophoresis patterns. Scand. J. Clin. Lab. Invest. 24, 55-60 (1969). 

Quantitative immunoelectrophoresis (Birkmeyer et al., 1981) and reverse quantitative immunoelectrophoresis (Birkmeyer et a)., 1982) allow the determination of average affinity of different antibody populations for an antigen. Plots of rocket area vs. the amount of antibody applied yield straight lines, the slope of which is indicative of affinity (steeper if the affinity is higher). Actual values of K are not obtained. Relative avidity indices can also be obtained with the test of Farr (1958) (Griswold and Nelson, 1984). 

Within certain limits, molecular morphology also comes in, as effects are observed in the agar network, according to the size and form of the protein molecule. Thus, immunoelectrophoresis is making use of three distinct properties of the protein molecule. It follows that the analytical consequences are considerable, even if results are not strictly quantitative. 

As stated in the introduction, immunoelectrophoresis (abbreviated ImEl) is a two-stage method. The electrophoretic run is comparatively simple and will be dealt with in a following chapter. The immunochemical reaction in agar gel is more complex, and this makes it necessary to examine the advantages of gel difEusion methods as well as their limitations. Many investigators use the Ouchterlony method, preceding or parallel to ImEl. This method will become still more important if present attempts at quantitation gain definite shape. 

Immunoelectrophoresis [123] can be characterized as two-dimensional agarose gel electrophoresis in which, in the second dimension, the advantage of immunoprecipi-tation reactions are utilized. There are five different arrangements of this technique in use today [124-127] (1) crossed Immunoelectrophoresis (2) fused rocket im-munoelectrophoresis (3) rocket immunoelectrophoresis (4) cross line immunoe-lectrophoresis (5) tandem crossed immunoelectrophoresis. For more detailed reviews see Refs. 128, 129. In all variations the important aspect is the possibility of obtaining quantitative data. 

Scherer, R., M. Abd-El-Fattah, and G. Ruthenstroth-Bauer. 1977. Some applications of quantitative two-dimensional immunoelectrophoresis in the study of the systemic acute-phase reaction of the rat. In Perspectives in inflammation, future trends and developments, ed. 

Immunoelectrophoresis is a specific method characterized by the presence of polyclonal monospecific antibodies against a specific protein in the agarose gel. The running buffer used in this method should have a pH corresponding to the isoelectric point of the antibody. The fixation of the latter in the gel leads to the formation of a precipitate comprised of the antigen and the specific antibody, which may be visualized by using a Coomassie blue stain or some other suitable technique (Westermeier and Scheibe, 2009). Immunoelectrophoresis has been used for the qualitative and quantitative analysis of individual proteins in complex mixture (Hansen and Larsen, 2008). 

Weeke, B. (1973) A manual of quantitative immunoelectrophoresis. Methods and applications. 1. General remarks and principles, equipment, reagents and procedures. Scand. ]. Immunol, 2 (Suppl. 1), 15-35... 

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