Preparation of colony stimulating activity from large batches of human urine and production of antisera against it

Abstract

In vitro induction of myelopoetic colonies from mouse bone marrow has been used for measurement of leucopoetic colony stimulating activity (CSA) isolated from large batches of human urine. After high flow dialysis in artificial kidneys and immediate adsorption to DEAE-Cellulose, followed by purification on Con A-Sepharose, treatment with insoluble Papain and gelfiltration on Sephadex G 100, enrichment of CSA was about 6,000-fold. An important step of the enrichment procedure was the separation from a CSA-inhibiting protein, probably combining with CSA. Specific activity was further increased by preparative polyacrylamide gel electrophoresis to 5.3 X 10(6) units per mg protein. The total enrichment exceeded 25,000-fold. The final purification product consisted of a group of closely related proteins with high specific activity. Antisera raised with one of the electrophoretic fractions suppressed bioactivity in each of the different purification steps including the final CSA fractions differing in electrophoretic mobility. The antisera furthermore inhibited CSA in human lung and monocyte conditioned media but had only very little effect on partially purified CSA from stimulated human lymphocytes as well as CSA derived from mouse lung conditioned medium.

Occurrence of a monocyte/macrophage colony‐stimulating factor in the continuous ambulatory peritoneal dialysis fluids and its chromatographic behaviors and antigenicity compared with human urinary colony‐stimulating factor

“…According to Metcalf and Stanley (19691, the urinary CSF concentration varies over a range of 0-300 unitdm1 depending upon the total daily urine volume, while the mean was about 60 units/ml. Other workers also reported the urinary CSF concentration to be about 9 units/ml (Motoyoshi et al, 1978), 30 units/ml (Laukel et al, 1978), or 32 unitdm1 (Ishii et al, 198213). The peritoneal CSF concentration observed in the present study seems to be nearly comparable with the concentration of CSF in normal human urine.…”

“…It is also our aim to compare the peritoneal CSF with the CSF from normal human urine. The urinary CSF has been considerably been purified and characterized by many workers (Stanley et al, 1975;Laukel et al, 1978;Motoyoshi et al, 1978). The comparison of the peritoneal and urinary CSFs will disclose the difference between the freshly produced CSF and the CSF excreted ultimately in the urine.…”

A granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) produced by carrageenin‐induced inflammatory cells of mice

“…The factor is produced in various tissues as well as by various cultured cells, and displays an apparent heterogeneity of molecular properties principally due to its nature as a sialoglycoprotein (Stanley et al, 1971;Ayusawa et al, 1979;Tsuneoka and Shikita, 1980a). On the other hand, CSFs produced by various organs of endotoxin-treated mice are universally similar to each other in that they all produce an active principle of 23,000 daltons upon desialylation and gel-filtration chromatography in the presence of 6 M guanidine (Nicola et al, 1979), whereas there is another subclass CSF of higher molecular weight, such as those obtained from mouse L-cell conditioned medium (Stan-ley and Heard, 1977;Tsuneoka and Shikita, 1977;Waheed and Shadduck, 1979;Tsuneoka and Shikita, 1980b) or from human urine (Stanley et al, 1975;Laukel et al, 1978). Recently, we have reported that two types of CSF are inducible in the culture of mouse spleen cells in response to either endotoxin or x-rays (Onoda et al, 1980).…”

Humoral Factors Modulating Growth of Granulocyte Macrophage Progenitor Cells

“…This supposed dual action of CSA would there fore be similar to that of erythropoietin in stimulating erythropoiesis and hemoglobin synthesis as well. Our current knowledge on the regulation of neutrophil and macrophage production by CSA almost entirely originates from cultures of progenitor cells in semisolid agar or methylcellulose [4,12,14,20].…”