Use of depth filtration in series with continuous centrifugation to clarify mammalian cell cultures
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Publication Number
US-7759117-B2
Publication Date
2010-07-20
Expiration Date
Abstract
Methods for clarification of cell samples using centrifugation in combination with depth filtration.
Core Innovation
The invention relates to separating therapeutic proteins from an industrial scale cell sample by clarifying large-scale mammalian and bacterial cell cultures. The process includes centrifuging the cell sample using a continuous disk-stack centrifugation under a controlled gravitational force and a Q/Σ ratio so that the centrifugation separates cells and cellular debris from a centrate comprising the therapeutic proteins.
After centrifugation, the clarified centrate is applied to a depth filtration means to recover a filtrate comprising the therapeutic proteins. The depth filtration means separates remaining particulate matter from the filtrate containing the therapeutic proteins, and clarification is achieved by coupling continuous disk-stack centrifugation with subsequent depth filtration/polishing.
The description characterizes the centrifugation and filtration train by balancing throughput versus shear-induced fines generation, using Q/Σ and gravitational force considerations. Studies evaluate solids removal efficiency, centrate turbidity trends, and filter capacity impacts, supporting low-g/low-Q/Σ operating windows and downstream filter configurations.
Claims Coverage
The independent claims cover 4 inventive features centered on centrifugation of an industrial scale cell sample under specified gravitational force and Q/Σ conditions, followed by depth filtration of the resulting centrate to recover therapeutic proteins while separating remaining particulate matter.
Centrifugation at specified gravitational force and Q/Σ ratio to form a therapeutic-protein centrate
Centrifuging the cell sample at a gravitational force of from about 8,000×g to about 15,000×g and a Q/Σ ratio of about 1×10^-8 to 28×10^-8 m/s, wherein the centrifugation separates the cells and cellular debris from the centrate which comprises the therapeutic proteins.
Depth filtration of the therapeutic-protein centrate to separate remaining particulate matter
Applying the centrate to a depth filtration means so as to recover a filtrate which comprises the therapeutic proteins, wherein the depth filtration means is effective to separate remaining particulate matter from the filtrate comprising the therapeutic proteins.
Centrifugation at gravitational force and constrained Q/Σ ratio to form a therapeutic-protein centrate
Centrifuging the cell sample at a gravitational force of from 8,000×g to 12,000×g and a Q/Σ ratio in the range of from 0.9×10^-28 to 2.8×10^-28 m/s, wherein the centrifugation separates the cells and cellular debris from the centrate which comprises the therapeutic proteins.
Centrifugation at higher gravitational force and constrained Q/Σ ratio to form a therapeutic-protein centrate
Centrifuging the cell sample at a gravitational force of from 9,600×g to 15,000×g and a Q/Σ ratio in the range of from 0.9×10^-28 to 2.8×10^-28 m/s, wherein the centrifugation separates the cells and cellular debris from the centrate which comprises the therapeutic proteins.
Across the independent claims, the claim coverage focuses on centrifugation of an industrial scale cell sample to produce a centrate comprising the therapeutic proteins, with specified gravitational force and Q/Σ ratio ranges, followed by depth filtration effective to separate remaining particulate matter while recovering a filtrate comprising the therapeutic proteins.
Stated Advantages
Clarification of large-scale mammalian and bacterial cell cultures by producing a filterable centrate.
Depth filtration effective to separate remaining particulate matter from filtrate comprising the therapeutic proteins.
Solids removal efficiency as reported in the description (about 95–99.8%+).
Filter capacity impact and turbidity trends that support selecting low-g/low-Q/Σ operating windows and downstream filter configurations.
Economic comparisons indicating reduced filter area when centrifugation is used upstream of final 0.1 μm or 0.2 μm filtration.
Documented Applications
Separating therapeutic proteins from an industrial scale cell sample derived from large-scale mammalian cell cultures.
Separating therapeutic proteins from an industrial scale cell sample derived from large-scale bacterial cell cultures.
Application to CHO cell culture and NSO cell culture, including IDEC-114 CHO and NSO cultures.
Integration in a process for clarification using a centrifugation and depth filtration train with downstream final 0.1 μm or 0.2 μm filtration for secreted therapeutic proteins and secreted antibodies.
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