Ouchterlony technique

Ouchterlony technique, 9 754 Ouricouri wax, 26 210—211 Ouricury, in mascara, 7 862 Outer approximations (OA), discrete optimization via, 26 1024 Outer—Helmholtz plane (OHP), 3 419 Outer-sphere (OS) mechanism,... 

By utilizing the Ouchterlony technique of two-dimensional, agar-gel diffusion, Slodki and his colleagues investigated the interaction of con A with O-phosphonomannans.363 They suggested a possible correlation between the extent of precipitate formation with con A and the content of a-D-(l— 2)-mannosidic linkages. 

Another antibody was experimentally produced in rabbits by injecting pepsin (G5). It was detected by the Ouchterlony technique and precipitation reactions, and may have some clinical significance in producing atrophy of chief cells. Repeated injection of this immune serum into rats also caused decrease in the volume of gastric juice, with lower concentrations of acid and pepsin. The controls injected with antibodies to egg albumin, nonspecific globulins, and saline did not show similar changes (see Fig. ). 

In an extension of these studies, Wada et al. (Wl) reported the incidence of positive precipitation reactions for serum proteins by the Ouchterlony technique to be 77.0% in gastric cancer (26 cases) and 64.5% in chronic gastritis (31 cases). However, this held true in only 15-20% of peptic ulcer cases. In atrophic gastritis, 6-8 precipitation lines were obtained with antisera to human serum and these included intensive lines of albumin and yglobulin and also faint lines of oro-somucoid, a2-globulin, ceruloplasmin, transferrin, and (3-globulin. 

Figure 9-4 Double immunodiffusion in two dimensions by the Ouchterlony technique. A, Reaction of identity B, reaction of nonidentity C, reaction of partial identity D, scheme for spur formation. Ag, Antigen Ab, antibody.

Ouchterlony technique. Double-radial immunodiffusion for the detection of precipitating autoantibodies against extractable nuclear antigens . Method of high diagnostic specificity but low sensitivity for diagnosis of autoimmune rheumatic diseases. 

Rl. Rhodes, J. M., The Ouchterlony technique using antigen or antiserum labelled with iodine. Nature 187, 793 (1960). 

Major progress in the analytical use of antibodies occurred with the development of several fundamental techniques the ability to detect cell-bound antibody (Coombs Test, 1945) immunoprecipitation in gels (Ouchterlony, 1953) radioimmunoassay (Yalow and Berson, 1960) and monoclonal antibodies (Kohler and Milstein, 1975). These, together with a con-... 

The titre of antibodies in laccase antisera was 1 16 as measured by the immunodiffusion technique of Ouchterlony (21). These antibodies were effective inhibitors of catechol oxidase activity, with 100/il of antisera reducing activity by 95%, in an assay mixture of 3ml of 0.1M catechol in... 

Fig. 2. Ouchterlony double-diffusion technique. The antigen is placed in the center well, cut in an agarose gel, and different antisera in a range of dilutions are placed in the sunounding wells. Antigen and antiserum diffuse toward each other and form a white precipitin line where an antibody recognizes the antigen.

This can be done by the Ouchterlony diffusion technique (see Fig. 2 and ref. 6). by enzyme-linked immunosorbent assay (see Chapter 15), or by Western blot, either using the purified protein or a more complex mixture of proteins containing the antigen of interest separated on a sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel (see Chapter 20). 

The most often used procedures for evaluating the specificity of antibody preparations are double diffusion methods developed by Ouchterlony and immunoelectrophoretic methods developed by Grabor and Williams. For a discussion of variations of these techniques as well as other useful procedures the reader is referred elsewhere (7-9). 

In 3 cases of Barta and Tichy (B3) with protein leakage into the gastric juice, serum proteins were identified by the Ouchterlony method. Hollander and Horowitz (H13), using this diffusion technique in canine gastric juice, demonstrated presence of serum albumin in canine gastric juice. 

Ouchterlony, 0., Interpretation of comparative immune precipitation patterns obtained by diffusion-in-gel techniques. In Immunochemical Approaches to Problems in Microbiology (M. Heidelberger and 0. J. Plescia, eds.), pp. 5-19. Rutgers Univ. Press, New Brunswick, New Jersey, 1960. 

Identification may be helped by changing the precipitating capacity of the antiserum. If the latter is absorbed with pure fractions of plasma proteins until no precipitation lines are formed in the Ouchterlony test, the pattern of ImEl will lack the corresponding lines. If the unknown antigen mixture contains fractions of tissues or tumors, the antiserum has to be absorbed first with pooled lyophilized human plasma. Another technique absorbs the antiserum specifically with a considerable excess of the unknown antigen a control ImEl shows the effect of absorption, thereby revealing the active antigen. Heremans (HI) has used this... 

The specificity and sensitivity of the antibody were verified using Immunodot, ouchterlony and ELISA techniques. The complete, detailed protocol for production of the antibody is available upon request and a brief version follows. 

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