Process for culturing altered human secretory pituitary cells

Claim 1

PROCESS FOR THE PRODUCTION OF ALTERED HUMAN SECRETORY PITUITARY CELLS HAVING A NUCLEUS CONTAINING FROM ABOUT 76 TO ABOUT 81 CHROMOSOMES INCLUDING A SMALL NUMBER OF TELOCENTRIC AND ACROCENTRIC CHROMOSOMES, SAID CELLS BEING HETERPLIOD AND POSSESSING PITUITARY HORMONE PRODUCING ACTIVITY WHICH COMPRISES THE STEPS OF MAINTAINING FRESHLY EXCISED HUMAN PITUITARY TISSUE AT ABOUT 37* C. IN A BASAL NUTRIENT MEDIUM COMPRISING AMINO ACIDS, VITAMINS, INORGANIC SALTS AND ANTIBIOTICS, CUTTING SAID TISSUES INTO SMALL FRAGMENTS WHILE IN CONTACT WITH SAID MEDIUM, CULTURING SAID FRAGMENTS FOR INTITIAL GROWTH IN A NUTRIENT MEDIUM COMPRISING SAID BASAL MEDIUM AND CONTAINING ABOUT 20% HUMAN SERUM BY VOLUME TOGETHER WITH CELLS NUTRIENTS UNTIL VIABLE TISSUE FRAGMENTS ARE OBTAINED, REPEATEDLY CULTURING SAID VIABLE TISSUE FRAGMENTS IN STATIONARY CULTURE MEDIA IN GLASS VESSELS AT ABOUT 37* C. AND AT A PH BELOW ABOUT 7.6 IN A NUTRIENT MEDIUM CONTAINING ABOUT 10% BY VOLUME OF ANIMAL SERUM AND CONTAINING AMINO ACIDS VITAMINS, INORGANIC SALTS, AND ANTIBIOTICS, TRANSFERRING SELECTED EPITHELIAL-LIKE CELLS ADHERING TO THE GLASS SURFACES OF SAID VESSELS BY GENTLE MECHANICAL MOVEMENT OF SAID CELLS AND INTRODUCTION THEREOF INTO SUCCESSIVE PORTIONS OF CULTURE MEDIUM, SAID STATIONARY CULTURE BEING CONTINUED UNTIL A CELL POPULATION DENSITY OF ABOUT 1 TO 100 CELLS PER SQ. CM. OF VESSEL FLOOR AREA IS ATTAINED TRANSFERRING SAID CELLS AT A CONCENTRATION OF ABOUT 2000,000 TO 3000,000 CELLS PER ML. TO A SUSPENSION CULTURE SYSTEM, REPEATEDLY CULTURING SAID CELLS IN SAID SUSPENSION SYSTEM AT A CELL POPULATION DENSITY OF ABOUT 50,000 TO ABOUT 5 MILLION OR MORE CELLS PER ML. IN A NUTRIENT MEDIUM CONTAINING A HIGHER CONCENTRATION OF AMINO ACIDS THAN THE STATIONARY CULTURE MEDIUM UNTIL ALTERED CELLS APPEAR WHICH EXHIBIT INCREASED CHROMOSOME COUNT, HETERPLOID CHARACTERISTICS, AND HORMONE PRODUCING ACTIVITY, AND SEPARATING SAID ALTERED CELLS.