Hollow fiber cell propagation system and method
First Claim
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1. A hollow fiber cell propagation system comprising:
- (i) a hollow fiber bioreactor including nutrient permeable hollow fibers positioned in an extracapillary space;
(ii) a first loop connected to a first inlet and a first outlet of said bioreactor so as to provide and change flow of fluids through lumens of said fibers, said first loop comprising;
(a) pump means for circulating fluids through said first loop;
(b) means for introducing gases into said first loop;
(c) means for determining and changing the temperature of the fluids in said first loop;
(d) means for determining the dissolved oxygen content of said fluids in said first loop upstream or downstream of said bioreactor, said dissolved oxygen determining means being positioned in flow block means with a sensing portion of said dissolved oxygen determining means positioned in said flow block means so as to be in contact with fluid flowing in said first loop, said flow block means including sterile filter means for passing calibrating gas into said flow block means so as to be in contact with said sensing portion for calibrating said dissolved oxygen determining means at a pressure equal to that of the fluid flowing in said first loop;
(e) means for determining and changing the flow rate of said fluids through said first loop;
(f) means for determining and changing pH of said fluids in said first loop;
(g) means for introducing fluids comprising reagents and basal medium containing nutrient into said first loop; and
(iii) a second loop connected to a second inlet and a second outlet of said bioreactor so as to provide and change flow of fluids through said extracapillary space.
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Abstract
A unique hollow fiber bioreactor system and method for the propagation of cells and the production of various cell propagation products is described. The system and method include the use of a flow block holder which allows a dissolved oxygen probe to be calibrated at the same pressure as the media in the bioreactor system.
88 Citations
55 Claims
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1. A hollow fiber cell propagation system comprising:
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(i) a hollow fiber bioreactor including nutrient permeable hollow fibers positioned in an extracapillary space; (ii) a first loop connected to a first inlet and a first outlet of said bioreactor so as to provide and change flow of fluids through lumens of said fibers, said first loop comprising; (a) pump means for circulating fluids through said first loop; (b) means for introducing gases into said first loop; (c) means for determining and changing the temperature of the fluids in said first loop; (d) means for determining the dissolved oxygen content of said fluids in said first loop upstream or downstream of said bioreactor, said dissolved oxygen determining means being positioned in flow block means with a sensing portion of said dissolved oxygen determining means positioned in said flow block means so as to be in contact with fluid flowing in said first loop, said flow block means including sterile filter means for passing calibrating gas into said flow block means so as to be in contact with said sensing portion for calibrating said dissolved oxygen determining means at a pressure equal to that of the fluid flowing in said first loop; (e) means for determining and changing the flow rate of said fluids through said first loop; (f) means for determining and changing pH of said fluids in said first loop; (g) means for introducing fluids comprising reagents and basal medium containing nutrient into said first loop; and (iii) a second loop connected to a second inlet and a second outlet of said bioreactor so as to provide and change flow of fluids through said extracapillary space. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27)
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28. A process for the conversion of a raw material to a useful product by cell metabolism which comprises
(i) inoculating the extracapillary space of a hollow fiber bioreactor with cells and a growth factor mixture for said cells; -
(ii) circulating basal medium comprising a nutrient into and through the lumens of the hollow fibers of said bioreactor; (iii) measuring dissolving oxygen content of said circulating basal medium before and after passage through said bioreactor with dissolved oxygen content determining means while said circulating basal medium is in a fluid flow loop including said lumens of said hollow fibers of said bioreactor; (iv) calibrating said dissolved oxygen determining means while said dissolving oxygen determining means is positioned in flow block means with a sensing portion of said dissolved oxygen determining means positioned in said flow block means so as to be in contact with fluid flowing in said fluid flow loop when said dissolved oxygen determining means is not being calibrated, passing calibrating gas into said flow block means so as to be in contact with said sensing portion for calibrating said dissolved oxygen determining means at a pressure equal to that of fluid flowing in said fluid flow loop; (v) introducing oxygen into said circulating basal medium to establish and maintain an oxygen content sufficient to support aerobic metabolism of said cells; (vi) utilizing the differential in dissolved oxygen content of the circulating basal medium before and after passage through the lumens of the hollow fibers of said bioreactor to determine aerobic efficiency and provide a real time verification and measurement of cell metabolism in said bioreactor. - View Dependent Claims (29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44)
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45. A hollow fiber process for the conversion of a raw material by cell metabolism which comprises
(i) inoculating the extracapillary space of a hollow fiber bioreactor with cells and a growth factor for said cells; -
(ii) passing basal medium and nutrients from a storage vessel into and through the lumens of said hollow fibers; (a) the rate of passage of said medium and reagents through said lumens being controlled to provide a preselected bioreactor glucose utilization rate; (iii) measuring dissolved oxygen content of said circulating basal medium before and after passage through said bioreactor with dissolved oxygen content determining means while said circulating basal medium is in a fluid flow loop including said lumens of said hollow fibers of said bioreactor, said dissolved oxygen content determining means including means for calibrating said dissolved oxygen content determining means at the same pressure as fluid flowing in said fluid flow loop; (iv) calibrating said dissolved oxygen determining means while said dissolved oxygen determining means is positioned in flow block means with a sensing portion of said dissolved oxygen determining means positioned in said flow block means so as to be in contact with fluid flowing in said fluid flow loop when said dissolved oxygen determining means is not being calibrated, passing calibrating gas into said flow block means so as to be in contact with said sensing portion for calibrating said dissolved oxygen determining means at a pressure equal to that of fluid flowing in said fluid flow loop; (v) introducing oxygen into said basal medium and nutrients in a predetermined amount sufficient to support aerobic metabolism of said cells; (vi) determining the differential in the dissolved oxygen content of said basal medium and nutrients upstream and downstream of said bioreactor; (vii) utilizing said dissolved oxygen differential to determine the aerobic efficiency of the cell metabolism in the bioreactor and to provide a real time verification and measurement of said cell metabolism; and (viii) simultaneously introducing growth factor into and removing product from said extracapillary space at rates which do not differ by more than about 5%. - View Dependent Claims (46, 47, 48)
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49. A process for conversion of a raw material into a useful product by cell metabolism which comprises:
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(i) inoculating the extracapillary space of a hollow fiber bioreactor with cells and a growth factor for said cells; (ii) circulating basal medium comprising a nutrient into and through the lumens of the hollow fibers of said bioreactor; (iii) measuring dissolved oxygen content of said circulating basal medium before and after passage through said bioreactor with dissolved oxygen content determining means while said lumens of said hollow fibers of said bioreactor; (iv) calibrating said dissolved oxygen determining means while said dissolved oxygen determining means is positioned in flow block means with a sensing portion of said dissolved oxygen determining means positioned in said flow block means so as to be in contact with fluid flowing in said fluid flow loop when said dissolved oxygen determining means is not being calibrated, passing calibrating gas into said flow block means so as to be in contact with said sensing portion for calibrating said dissolved oxygen determining means at a pressure equal to that of fluid flowing in said fluid flow loop; (v) introducing oxygen into said circulating basal medium to establish and maintain an oxygen content sufficient to support aerobic metabolism of said cells; and (vi) simultaneously measuring the glucose utilization rate (GUR) of said bioreactor and the dissolved oxygen (Δ
DO) differential across said bioreactor and using the GUR/Δ
DO ratio to assess the bioreactor cell production efficiency of said bioreactor. - View Dependent Claims (50, 51, 52)
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53. A process for the conversion of a raw material into a useful product by cell metabolism which comprises:
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(i) inoculating the extracapillary space of a hollow fiber bioreactor with cells and a growth factor for said cells; (ii) introducing a glucose free basal medium into a means for circulating fluids through the lumens of the hollow fibers of said bioreactor; (iii) separately introducing glucose into said means for circulating fluids through the lumens of said hollow fibers of said bioreactor; (iv) measuring dissolved oxygen content of said circulating basal medium before and after passage through said bioreactor with dissolved oxygen content determining means while said circulating basal medium is in a fluid flow loop including said lumens of said hollow fibers of said bioreactor; (v) calibrating said dissolved oxygen determining means while said dissolved oxygen determining means is positioned in flow block means with a sensing portion of said dissolved oxygen determining means positioned in said flow block means so as to be in contact with fluid flowing in said fluid flow loop when said dissolved oxygen determining means is not being calibrated, passing calibrating gas into said flow block means so as to be in contact with said sensing portion for calibrating said dissolved oxygen determining means at a pressure equal to that of fluid flowing in said fluid flow loop; (vi) introducing oxygen into said circulating basal medium to establish and maintain an oxygen content sufficient to support aerobic metabolism of said cells; and (vii) simultaneously measuring the glucose utilization rate (GUR) of said bioreactor and the dissolved oxygen (Δ
DO) differential across said bioreactor and using the GUR/Δ
DO ratio to assess the bioreactor cell production efficiency of said bioreactor. - View Dependent Claims (54)
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55. A hollow fiber process for the conversion of raw material by cell metabolism which comprises
(i) inoculating the extracapillary space of a hollow fiber bioreactor will cells and a growth factor for said cells; -
(ii) passing basal medium and nutrients from a storage vessel into and through the lumens of said hollow fibers; (a) the rate of passage of said medium and reagents through said lumens being controlled to provide a selected bioreactor glucose utilization rate; (iii) measuring dissolved oxygen content of said circulating basal medium before and after passage through said bioreactor with dissolved oxygen content determining means while said circulating basal medium is in a fluid flow loop including said lumens of said hollow fibers of said bioreactor; (iv) calibrating said dissolved oxygen determining means while said dissolved oxygen determining means is positioned in flow block means with a sensing portion of said dissolved oxygen determining means positioned in said flow block means so as to be in contact with fluid flowing in said fluid flow loop when said dissolved oxygen determining means is not being calibrated, passing calibrating gas into said flow block means so as to be in contact with said sensing portion for calibrating said dissolved oxygen determining means at a pressure equal to that of fluid flowing in said fluid flow loop; (v) introducing oxygen into said basal medium and nutrients in a predetermined amount sufficient to support aerobic metabolism of said cells; (vi) simultaneously measuring the glucose utilization rate (GUR) of said bioreactor and the dissolved oxygen (Δ
DO) differential across said bioreactor and using the GUR/Δ
DO ratio to assess the bioreactor cell production efficiency of said bioreactor; and(vii) monitoring and controlling one or more of steps (i) through (vi) with a programmable controller.
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Specification