Human Leukocytic Acid Phosphatase

The acid phosphatase activity of leukocytes was studied by Valentine and Beck (B8, VI) in 1951. There appear, however, to have been no significant attempts to purify the enzyme from this source, or to describe its characteristics. Recently, Szajd and Pajdak (S32) indicated the isoenzyme characteristics of leukocyte acid phosphatase, and Li and his associates (L7, L8) studied this problem in greater detail. They suspended a leukocyte preparation, carefully separated from blood, in 5% Triton X-100 to yield a final concentration of 10 X 10 cells per milliliter and subjected the suspension to six cycles of alternate freeze-thaw treatment. The suspension was then centrifuged at lOOOp for 15 minutes at 4°C, and the supernatant was used for electrophoretic studies. Specimens centrifuged at 100,000p for 15 minutes gave the same results. Electrophoresis was carried out at 4°C for 60 minutes on a 7.5% acrylamide gel matrix containing 0.5% Triton X-100 at pH 4.0 with a current of 4 mA per tube. The substrate was -naphthyl phosphate. 

The values for the normal leukocyte acid phosphatase activity and the normal isoenzyme pattern will be described in connection with the alteration of these in various hematologic and hematopoietic disorders. 

Liver acid phosphatase has been of particular interest since the demonstration by de Duve (D7, D8, D9, DIO) that acid phosphatase and other hydrolytic enzymes were enclosed in an intracellular structure, the lysosome, of the liver and played an important role in the intra- 

The purification of acid phosphatase from the human liver and the description of its properties do not appear to have been accomplished. Partly, this may be due to the inherent difiBculty of obtaining normal, fresh human material in amounts substantial enough for purification. However, because of the cellular and physiological importance of acid phosphatase, it is advisable to describe in the present section the purifications of the enzyme from rat and bovine liver. Moreover, since these purifications were accomplished with the awareness that acid phosphatase from this source might be present in multiple molecular forms, the descriptions will naturally involve a consideration of the isoenzymes and their properties. 

A 336-fold purification by column chromatography was achieved by Brightwell and Tappel (B32) from lysosomes obtained by differential and density sucrose gradients. The lysosomes were frozen and thawed several times, then centrifuged. The resulting soluble acid phosphatase fraction was dialyzed against suitable buffers and then applied to a DEAE-cellulose column or to a CM-cellulose column. Each column was eluted with a linear 0-1 M NaCl solution, the former at pH 7.2 and the latter at pH 5.6. Two peaks of acid phosphatase activity were ob- 

---

Smith GP, Peters TJ (1981) SubceUular localization and properties of pyridoxal phosphate phosphatases of human polymorphonuclear leukocytes and their relationship to acid and alkaline phosphatase. Biochim Biophys Acta 661 287-294... 

---