SYSTEMS AND METHODS FOR CONTINUOUS CULTIVATION AND RECOVERY OF COMESTIBLE NON-HUMAN ANIMAL CELLS
The present disclosure relates to systems, apparatuses, and methods for continuous cultivation and recovery of comestible non-human animal cells in a suspension bioreactor. For example, in one or more implementations, the disclosed methods include cultivating non-human animal cells in suspension culture within a bioreactor vessel, removing a measure of suspension culture from the bioreactor vessel while continuing to provide cell culture media thereto, and recovering a first portion of cultivated non-human animal cells from the removed measure of suspension culture while returning a second portion of cultivated non-human animal cells to the bioreactor vessel. Also, in some implementations, the disclosed systems include at least one sterile output module configured to prevent backflow contamination when recovering cultivated cells and removing spent media during continuous cultivation of non-human animal cells according to the disclosed methods.
1 . A method for producing animal cells, the method comprising:
cultivating a suspension culture comprising animal cells in a bioreactor vessel;
removing a measure of the suspension culture from the bioreactor vessel while providing cell culture media to the bioreactor vessel;
recovering a first portion of the measure of the suspension culture for harvest concurrent with returning a second portion of the measure of the suspension culture to the bioreactor vessel without returning the first portion to the bioreactor vessel; and
harvesting animal cells from the first portion of the suspension culture for further processing.
2 . The method of claim 1 , further comprising continuing cultivation of the suspension culture in the bioreactor vessel while recovering the first portion for harvest and returning the second portion to the bioreactor vessel.
3 . The method of claim 1 , further comprising:
monitoring at least one culture parameter within the bioreactor vessel; and
controlling, based on the monitored at least one culture parameter, at least one of:
an amount of the measure removed from the bioreactor vessel;
a fraction of the measure recovered as the first portion; or
a fraction of the measure returned as the second portion.
4 . The method of claim 1 , wherein removing the measure of suspension culture while providing the cell culture media comprises maintaining a culture volume within the bioreactor vessel within a target range by coordinating a media addition rate and a removal rate.
5 . The method of claim 1 , further comprising:
processing the removed measure of the suspension culture using a cell retention device to provide a concentrated cell culture having a higher cell concentration than the removed measure and a culture broth having a lower cell concentration than the removed measure; and
recovering the first portion concurrent with returning the second portion by recovering and returning respective portions of the concentrated cell culture.
6 . The method of claim 5 , wherein the cell retention device comprises a continuous flow centrifuge, a sterile filtration device, a gravity sedimentation device, or a magnetic separation device.
7 . The method of claim 5 , wherein the cell retention device comprises a sterile cell retention device disposed external to the bioreactor vessel and fluidly coupled to the bioreactor vessel by (i) a suspension culture line configured to convey the removed measure from the bioreactor vessel to the sterile cell retention device and (ii) a cell retention line configured to return at least some of the concentrated cell culture to the bioreactor vessel.
8 . The method of claim 1 , wherein removing the measure, recovering the first portion, and returning the second portion are performed without removing the measure or the second portion from a sterile environment comprising the bioreactor vessel and one or more additional system components.
9 . The method of claim 8 , further comprising transferring the first portion through a sterile output module that provides a boundary between the sterile environment and a non-sterile environment, wherein the sterile output module is configured to prevent backflow contamination into the sterile environment during recovery of the first portion.
10 . The method of claim 1 , further comprising directing the first portion to a cell concentration device configured to remove spent media from the first portion to recover a further concentrated cell mass for further processing.
11 . A method for producing animal cells, the method comprising:
cultivating a suspension culture comprising animal cells in a bioreactor vessel during a steady-state cultivation period by:
removing one or more measures of the suspension culture from the bioreactor vessel while providing cell culture media to the bioreactor vessel;
for each removed measure of the suspension culture, recovering a first portion of the removed measure for harvest concurrent with returning a second portion of the removed measure to the bioreactor vessel, wherein the first portion is not returned to the bioreactor vessel; and
transferring the first portion for further processing of cultivated animal cells contained in the first portion.
12 . The method of claim 11 , further comprising maintaining, during the steady-state cultivation period, a cell density within the bioreactor vessel at or near a target cell density by controlling at least one of: (i) a media addition rate, (ii) a removal rate, (iii) a fraction of each removed measure recovered as the first portion, or (iv) a fraction of each removed measure returned as the second portion.
13 . The method of claim 11 , wherein providing the cell culture media and removing the one or more measures during the steady-state cultivation period are performed according to a predetermined or controlled rate profile.
14 . The method of claim 11 , further comprising:
for each removed measure of the suspension culture, processing the removed measure using a cell retention device to provide (i) a concentrated cell culture having a higher cell concentration than the removed measure and (ii) a culture broth having a lower cell concentration than the removed measure; and
recovering the first portion concurrent with returning the second portion by recovering and returning respective portions of the concentrated cell culture.
15 . The method of claim 14 , wherein the cell retention device comprises a continuous flow centrifuge, a sterile filtration device, a gravity sedimentation device, or a magnetic separation device.
16 . The method of claim 14 , wherein the cell retention device comprises a sterile cell retention device disposed external to the bioreactor vessel by (i) a suspension culture line configured to convey each removed measure from the bioreactor vessel to the sterile cell retention device and (ii) a cell retention line configured to return at least some of the concentrated cell culture to the bioreactor vessel.
17 . The method of claim 11 , further comprising transferring the first portion through a sterile output module that provides a boundary between (i) a sterile environment comprising the bioreactor vessel and one or more additional system components and (ii) a non-sterile environment, wherein the sterile output module is configured to prevent backflow contamination into the sterile environment during transfer of the first portion.
18 . The method of claim 17 , wherein the sterile output module comprises a plurality of flow paths configured such that, during transfer of the first portion through a first parallel flow path, at least a second parallel flow path is isolated for cleaning or sterilization.
19 . The method of claim 11 , wherein cultivating during the steady-state cultivation period comprises flowing suspension culture through a suspension culture line including a bypass line that separates the suspension culture line into a first zone having a higher flow rate and a second zone having a lower flow rate to decouple at least one of: distance between the bioreactor vessel and a downstream device, flow rate through the downstream device, or residence time of cells within the suspension culture line.
20 . The method of claim 11 , further comprising:
monitoring, during the steady-state cultivation period, at least one culture parameter within the bioreactor vessel; and
adjusting, based on the at least one culture parameter, at least one of (i) a recovery fraction associated with recovering the first portion or (ii) a return fraction associated with returning the second portion.