An abnormal isozyme of alkaline phosphatase (ALP: orthophosphoric monoester phosphohydrolase, EC 3. 1. 3. 1) detected in the serum of four children with no malignancy is described.
The molecular weights of the abnormal enzyme in all four patients were about 1.7×10⁵ daltons, and the enzyme migrated to the front position of the liver ALP (α1∼α2 globulin region) irrespective of electrophoretic medium. Other features of the enzyme, i.e., Michaelis constants, molecular sizes, susceptibilities to amino acids and to the action of neuraminidase, and heat stability suggest that this enzyme is an anomalous bone ALP with respect to sugar chains.

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Enzyme activities and isozyme patterns of blood plasma were compared with those of heart muscles in the hypoxic states using Wistar male rats. The findings of myocardial damage during hypoxia were not so significant in plasma LDH-isozyme. Plasma LDH-isozyme did not show the pattern of heart muscles, but it was like that of the liver even in the hypoxic states. Pattern of plasma CK-isozyme during hypoxia resembled that of heart muscles except increasing CK-BB, and mitochondrial CK was also recognized in cathode side toward CK-MM in plasma. It could be said that hypoxia might play a important role as to the appearances of type 2 macro-CK in plasma. Plasma CK-isozyme indicated more quickly a findings of myocardial damage during hypoxia than plasma LDH-isozyme.

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The urinary proteins from 19 patients of renal disease were analyzed with micro two-dimensional electrophoresis in the absence of denaturing agents. The two-dimensional electrophoretic patterns of the urinary proteins resembled with those of plasma proteins, although the contents of high molecular weight plasma proteins were low. The patterns of the patients with nephrotic syndrome were characterized by the high content of α₁-acid glycoprotein. Comparing the urinary protein patterns, differences in the contents of G_C-globulin, prealbumin, α₂ HS-glycoprotein, IgG polymer and unidentified low molecular weight proteins were demonstrated.

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The cellular proteins from various human organs were analyzed by two-dimensional (isoelectric focusing-SDS PAGE) electrophoresis. More than one hundred of polypeptide spots were detected in each organ. Although a unique polypeptide map was obtained from each organ, the patterns of stomach and colon, both originating from the same germ layer and possessing columnar epithelium, were relatively similar. The patterns of heart muscle and skeletal muscle, both originating from the same germ layer and possessing the ability of contraction, were quite different from each other. Those of liver and kidney were similar, although the organs were originated from different germ layer and possessed different functions and cell types. Therefore, the patterns in various organs could not be classified according to their histology, functions and embryology.
There were three spots (molecular weight×10^-3/isoelectric point: 55/6.0, 43/7.6, 38/8.0), which were commonly found in all of the organs except for the blood. These spots might be species-associated cellular proteins.
There were several qualitatively or quantitatively characteristic spots in each organ. These might be organ-associated cellular proteins.

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An anti-acetylated lactate dehydrogenase (LD) B₄ anti-serum was prepared from a New Zealand white male rabbit. This anti-serum formed soluble LD and antibody complexes with non-acetylated LD B₄ molecules without inhibiting the LD activity.
This property was also found in other LD isoenzymes containing B subunit in their molecules. Changes of electrophoretic mobility towards the cathodic side were observed in these LD antibody complexes, and free and bound LD isoenzymes were completely separated and could be quantitated by densitometric analysis. These soluble complexes were completely precipitated by the addition of anti-rabbit immunoglobulin G and the free LD activity was clearly measured by the assay of remaining activity in the centrifugal supernatant.
Employing these two separating methods, i. e., the electrophoretic and the immunological method, the equilibrium constant (K) and the heterogeneity index (a) of the antibody for each LD isoenzyme were measured by Sips plot analysis. The antibody showed a high affinity to LD isoenzymes and a different heterogeneity index for each LD isoenzyme. These findings should be useful for the analysis of enzyme linked immunoglobulin appearing in human sera.

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Human colon cancer was transplanted to nude mouse and the changes in serum protein distribution were examined by micro two-dimensional electrophoresis in the absence of denaturing agents. Three serum proteins, Spot 1, 2 and 3, were increased in level by the cancer transplantation. The isoelectric point (pI) and molecular weight (MW) were: Spot 1, pI 4.7-5.2, MW 120, 000; Spot 2, pI 5.2, MW 110, 000; Spot 3, pI 5.1-5.8, MW 90, 000. Spot 1 was identified to be haptoglobin fully saturated with hemoglobin by benzidine staining. Using blotting-immunochemical staining, 17 mouse serum proteins were identified on the two-dimensional protein map and Spot 3 was identified to be hemopexin.

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