Tegretol Xr

posted on 27 Aug 2012 08:04 by tegretolxrdu
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ence of an equivalence zone or faulty technique. Precipitation in gels. The precipitation of antigen and antibody in a gel has been used for the analysis of antigen mixtures by Oudin, Ouchter- lony, and others (see Oudin, 1952). In the one-dimensional simple diffu- sion procedure, one reagent, usually antibody, is mixed with agar and 214 MANUAL OF MICROBIOLOGICAL METHODS placed in a small test tube. The other reagent (antigen) is layered over the agar, and the test allowed to incubate 4 to 7 days. If only one antigen is present, there will be only one zone of precipitation, but if Tegretol Xr addi- tional antigens are present, additional rings will be seen, assuming, of course, that the serum contained antibodies directed against the other antigens and also assuming that two rings do not coincide because of diffusion rates and concentration. In the double-diffusion procedure, agar which contains neither reactant is poured into a petri dish. The antigen and antibody are placed in wells and allowed to diffuse toward each other with the resultant formation of lines of precipitation. Since a complete discussion of these procedures is available in the review by Oudin (1952), the details Tegretol Xr have been omitted here. The quantitative measurement of precipitin. The procedures described elsewhere in this section are entirely adequate for some applications involving microbial antigens. In other instances, it may be more profit- able to employ a technic that permits the precise estimation of antibody contained in a precipitate. This may be done with a variation of the quantitative precipitin test as developed by Heidelberger and Kendall (1929, 1933, 1935). When microbial antigens are being studied, the quantitative precipitin test is useful for closely related (cross reacting) antigens, for studjdng the contamination of one antigen by another, for estimating concentration of an antigen present in a solution, or for any other application where the less quantitative technics are insufficiently sensitive. In the usual form of the quantitative test, increasing amounts of anti- gen are added to constant volumes of antiserum and the tubes stored at some constant temperature (0-4C) for several days until equilibrium has been reached. The precipitates are then carefully removed by centrifugation, and extraneous serum proteins are removed by rinsing the precipitates with ice-cold sahne solution. The precipitates are analyzed for nitrogen by a micro-Kjeldahl procedure. In the hands of a careful investigator this method permits accuracy that is comparable to that obtained in quantitative analytical chemistry. In addition to nitrogen analysis, antibody has been measured quantitatively in other ways. A procedure that is useful for serums with low concentrations of antibody is the Folin Ciocalteau analysis for tyrosine (Heidelberger and MacPherson, 1943). While the quantitative estimation of precipitins is a relatively simple operation, its proper evaluation may require more information than it is possible to impart in a chapter of this magnitude. Since other sources of material (Kabat and Mayer, 1948; Cohn, 1952) are available, further description of the procedures has not been included here. SEROLOGICAL METHODS 215 COMPLEMENT-FIXATION TEST FOR MICROBIAL ANTIGENS In the normal serum of numerous animal species there occurs a sub- stance known as alexin or complement. It is a mixture of at least four fractions, chiefly globulin in nature. Although it does not increase in amount Tegretol Xr following immunization, it has a role in serological reactions, since it is able to combine with many antigen-antibody complexes. This combination may be spoken of as the ''fixation" of complement. When erythrocytes are mixed with antierythrocyte serum and comple- ment is added, lysis of the cells will occur. Such Tegretol Xr lysis requires both anti- body and complement. Practical application is made of this lytic phenomenon in the complement-fixation test where sheep erythrocytes and their homologous antibody (hemolysin) are used as an indicator system. In the complement-fixation test, two separate antigen-antibody sys- tems are allowed to compete for complement. When the test is per- formed the antigen and antiserum under study are mixed and a carefully measured amount of complement is added. If the serum contains anti- bodies that react with the antigen, complement is absorbed by the anti- gen-antibody complex and no free complement remains. If, at this point, sheep erythrocytes and their homologous antibody (hemolysin) are added, the sheep cells will not lyse, since there is no free complement. If, on the other hand, the first antigen-antibody reaction had failed to occur owing to deficiency of either antibody or antigen, free complement would be available and the sheep cell indicator system would lyse. The complement-fixation test is not necessary for some systems where other antigen-antibody reactions are available; however, it sometimes yields information that cannot be obtained by other procedures. The test described here is a modification of the Kolmer method for syphilis (Kolmer 62^ aL, 1951). Reagents Tegretol Xr needed for the test include 0.9 per cent sodium chloride solu- tion (''saline"), a suspension of sheep erythrocytes, hemolysin, comple- ment, antigen, and antiserum. Preparation of sheep erythrocytes. Blood is collected from the jugular vein of the sheep. The erythrocytes may be preserved for several weeks by mixing the blood immediately with an equal volume of sterile modified Alsever solution (Bukantz et at., 1946) of the following composition: Glucose 20.5 g Sodium citrate 8.0 g Sodium chloride 4.2 g Tegretol Xr Citric acid . 55 Tegretol Xr g Distilled water 1 liter 216 MANUAL OF MICKOBIOLOGICAL METHODS If a sheep is not available, Tegretol Xr sheep blood may be obtained commercially; however, the erythrocytes are likely not Tegretol Xr to survive shipment through the mail. Freshly collected blood should be stored 48 hr before use.
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posted on 26 Aug 2012 08:02 by tegretolxrdu

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