[Immunochemical characterization of alpha-amylases in wheat seeds at different ontogenical steps (author's transl)].

Protein extracts of wheat seeds taken at an early stage of development, at maturation and after seven days of germination were investigated by using immunochemical techniques with immune sera prepared against alpha-amylases purified from developing seeds and alpha-amylases purified from germinated seeds. After immunochemical analyses carried out in agarose gel, alpha-amylase characterization was performed by using beta-limit dextrin followed by iodine staining. Detection of three antigenic alpha-amylases separated by agarose immunoelectrophoretic analysis at pH 8.6 and called I, II, III from the anode to the cathode, as well as an antigenical relationship between the anodic enzymes I and II were confirmed. Three constituents in I and four in II were further distinguished by using long duration electrophoresis in agarose gel. The immune sera reacted with all of these constituents. Thus with these immune sera a quantitative determination of all anodic alpha-amylase proteins can be attempted as quantitation of two antigens. During this identification consitutents with a apparent activity on beta-limit dextrin but delivering incomplete unstained substrate were detected. These constituents found in developing seeds have electrophoretic mobility close to that of constituents I but differ antigenically from alpha-amylases I and II. Combination of rocket-line-immunoelectrophoresis and alpha-amylase characterization reaction on precipitin bands was developed for comparing amounts of each of three alpha-amylase antigens in different seed extracts. The use of this technique for quantitating at the same time two antigen groups having a certain cross reactivity is discussed. Preliminary results of quantitative study on each of these antigen in developing, mature and germinating seeds are reported.