Cell and Vaccine Production

36 cm.sup.2 sintered titanium plates are immersed in 40 ml of media consisting of 50% F 10, 40% 199, 5% tryptose phosphate broth and 5% calf serum. Trypsinized chick embryo fibroblasts are suspended in the media at a concentration of 5 .times. 10.sup. 5 cells per centimeter square of surface area. The unit is held at 36.5.degree.C in a CO.sub.2 incubator for 3 days during which time the initially plated cells spread over the surface forming a cell sheet.

A 3 l. tank operating with 50 circular sintered titanium plates is completely filled with a slurry of trypsinized chick embryo fibroblasts suspended in a medium consisting of 199 with 45ml of 2.8% NaHCO.sub.3 per liter and containing 10% fetal calf serum. The unit is held such that the plates are parallel to the work surface and the cells settle onto the surface for 3 hours. Following this the fluid is drained, mixed with fresh chick cells and returned to the tank. The tank is rotated through 180.degree. so that the lower side of the plates become the upper. The new cells are allowed to plate for 3 hours.

1.5 l. of fluid is removed, the unit is rotated through 90.degree. to bring the plate perpendicular to the work surface and the plates are rotated about the center axis so that the plated cells pass through the growth media and are aerated in a cyclical manner. The operation is continued for 72 hours until complete cell sheets are formed. The cells are recovered from the tank by draining the growth fluid, followed by the addition of a solution of trypsin which strips the cells from the plates. Draining this final fluid yields a usable cell slurry. The trypsin can be inactivated with calf serum to prevent cell destruction.

A 3 l. tank operating with 50 circular sputtered titanium plates is completely filled with a slurry of trypsinized chick embryo fibroblasts suspended in a medium consisting of 199 with 45ml of 2.8% NaHCO.sub.3 per liter and containing 10% fetal calf serum. The unit is held such that the plates are parallel to the work surface and the cells settle onto the surface for 3 hours. Following this the fluid is drained, mixed with fresh chick cells and returned to the tank. The tank is rotated through 180 degrees so that the lower side of the plates becomes the upper. The new cells are allowed to plate for 3 hours.

1.5 l. of fluid is removed, the unit is rotated through 90.degree. to bring the plate perpendicular to the work surface and the plates are rotated about the center axis so that the plated cells pass through the growth media and are aerated in a cyclical manner. The operation is continued for 72 hours until complete cell sheets are formed. The unit is then washed with Hank's solution and mumps virus in media 199 with 25% of SPGA is added to infect the cell sheet. After 96 hours this fluid is drained from the unit. The fluid obtained may be used as a mumps vaccine.

A 3 l. tank operating with 50 circular sintered glass plates is completely filled with a slurry of trypsinized duck embryo fibroblasts suspended in a medium consisting of media 199 with 45ml of 2.8% NaHCO.sub.3 per liter and containing 10% fetal calf serum. The unit is held such that the plates are parallel to the work surface and the cells settle onto the surface for 3 hours. Following this the fluid is drained, mixed with fresh duck cells and returned to the tank. The tank is rotated through 180.degree. so that the lower side of the plates becomes the upper. The new cells are allowed to plate for 3 hours.

1.5 l. of fluid is removed, the unit is rotated through 90.degree. to bring the plate perpendicular to the work surface and the plates are rotated about the center axis so that the plated cells pass through the growth media and are aerated in a cyclical manner. The operation is continued for 72 hours until complete cell sheets are formed. The unit is then washed with Hank's solution and rubella virus in media 199 with 25% of SPGA is added to infect the cell sheet. After 96 hours this fluid is drained from the unit. The fluid obtained may be used as a rubella vaccine.

A 3 l. tank operating with 50 circular sintered plastic plates is completely filled with a slurry of trypsinized green monkey kidney cells suspended in a medium consisting of media 199 with 45ml of 2.8% NaHCO.sub.3 per liter and containing 10% fetal calf serum. The unit is held such that the plates are parallel to the work surface and the cells settle onto the surface for 3 hours. Following this the fluid is drained, mixed with fresh green monkey kidney cells and returned to the tank. The tank is rotated through 180.degree. so that the lower side of the plates becomes the upper. The new cells are allowed to plate for 3 hours.

1.5 l. of fluid is removed, the unit is rotated through 90.degree. to bring the plate perpendicular to the work surface and the plates are rotated about the center axis so that the plated cells pass through the growth media and are aerated in a cyclical manner. The operation is continued for 72 hours until complete cell sheets are formed. The cells are recovered from the tank by draining the growth fluid followed by the addition of a solution of trypsin which strips the cells from the plates. Draining this final fluid yields a usable cell slurry. The trypsin can be inactivated with calf serum to prevent cell destruction.

A 3 l. tank operating with 50 circular sintered stainless steel plates is completely filled with a slurry of VERO cells suspended in a medium consisting of 199 with 45ml of 2.8% NaHCO.sub.3 per liter and containing 10% fetal calf serum. The unit is held such that the plates are parallel to the work surface and the cells settle onto the surface for 3 hours. Following this the fluid is drained, mixed with fresh VERO cells and returned to the tank. The tank is rotated through 180.degree. so that the lower side of the plates becomes the upper. The new cells are allowed to plate for 3 hours.

1.5 l. of fluid is removed, the unit is rotated through 90.degree. to bring the plate perpendicular to the work surface and the plates are rotated about the center axis so that the plated cells pass through the growth media and are aerated in a cyclical manner. The operation is continued for 72 hours until complete cell sheets are formed. The cells are recovered from the tank by draining the growth fluid, followed by the addition of a solution of trypsin which strips the cells from the plates. Draining this final fluid yields a usable cell slurry. The trypsin can be inactivated with calf serum to prevent cell destruction.

A 3 l. tank operating with 50 circular sintered titanium plates is completely filled with a slurry of WI-38 cells suspended in a medium consisting of EBME with 45ml of 2.8% NaHCO.sub.3 per liter and containing 10% fetal calf serum. The unit is held such that the plates are parallel to the work surface and the cells settle onto the surface for 3 hours. Following this the fluid is drained, mixed with fresh WI-38 cells and returned to the tank. The tank is rotated through 180.degree. so that the lower side of the plates becomes the upper. The new cells are allowed to plate for 3 hours.

1.5 l. of fluid is removed, the unit is rotated through 90.degree. to bring the plate perpendicular to the work surface and the plates are rotated about the center axis so that the plated cells pass through the growth media and are aerated in a cyclical manner. The operation is continued for 72 hours until complete cell sheets are formed. The cells are recovered from the tank by draining the growth fluid, followed by the addition of a solution of trypsin which strips the cells from the plates. Draining this final fluid yields a usable cell slurry. The trypsin can be inactivated with calf serum to prevent cell destruction.

A 3 l. tank operating with 50 circular sintered titanium plates is completely filled with a slurry of green monkey kidney cells suspended in a medium consisting of 199 with 45ml of 2.8% NaHCO.sub.3 per liter and containing 10% fetal calf serum. The unit is held such that the plates are parallel to the work surface and the cells settle onto the surface for 3 hours. Following this the fluid is drained, mixed with fresh green monkey kidney cells and returned to the tank. The tank is rotated through 180.degree. so that the lower side of the plates becomes the upper. The new cells are allowed to plate for 3 hours.

Approximately 1.5 l. of fluid is removed, the unit is rotated through 90.degree. plate bring the plater perpendicular to the work surface and the plates are rotated about the center axis so that the plated cells pass through the growth media and are aerated in a cyclical manner. The operation is continued for 72 hours until complete cell sheets are formed. The unit is then washed with Hank's solution and parainfluenza virus in media 199 with 25% of SPGA is added to infect the cell sheet. After 96 hours this fluid is drained from the unit. The fluid obtained may be used as a parainfluenza vaccine.