Underivatized Controlled Pore Glass for Immunoglobulin Aggregate Removal
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Solution Overview
Problem
The downstream processing of immunoglobulins often results in the formation of dimers and oligomers, which lack antigen-binding activity and are difficult to remove, complicating the purification process and requiring additional steps to achieve low-volume, high-concentration solutions for pharmaceutical applications.
Innovation Solution
The use of underivatized controlled pore glass (uCPG) selectively binds dimeric and oligomeric immunoglobulins at pH values between 5 and 7.5, allowing for the recovery of monomeric immunoglobulins with up to 95% efficiency by adsorption and subsequent removal of aggregates through centrifugation or filtration, without altering the protein structure.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional downstream processing steps (affinity chromatography, ion exchange chromatography) are used to purify immunoglobulins, then non-immunoglobulin molecules and contaminants are removed, but dimers and oligomers form and are difficult to remove
Solution Approach 1:
The patent employs controlled pore glass (CPG) with specific pore sizes (50-200 nm) to separate monomeric immunoglobulins from aggregated forms. The porous structure allows size-based discrimination where monomers pass through while larger aggregates are retained, effectively removing harmful aggregates without requiring additional purification steps that might induce further aggregation.
Solution Approach 2:
The patent utilizes pH gradient changes to control the binding and elution of immunoglobulins on CPG. By adjusting pH from acidic (binding condition) to neutral/alkaline (elution condition), the method selectively elutes monomeric immunoglobulins while retaining aggregates, thereby improving purification quality and reducing aggregate formation in the final product.
2Quantity of substance
If concentration steps are applied to achieve low volumes for economic handling, then storage and handling efficiency improve, but aggregate formation increases
Solution Approach 1:
The CPG filtration step is integrated into the concentration process, allowing the system to achieve high concentrations while continuously removing aggregates that form during concentration. The porous structure physically separates aggregates from monomers even at high concentrations, preventing the harmful effect of aggregate accumulation.
Solution Approach 2:
The patent applies CPG filtration before final concentration steps to pre-remove aggregates. This preliminary action prevents aggregates from forming during subsequent concentration operations, maintaining solution quality even as concentration increases for economic handling and storage.
3Reliability
If additional purification steps are added to remove aggregates, then product quality improves, but process complexity and time increase
Solution Approach 1:
The patent combines aggregate removal with existing purification steps by integrating CPG filtration into the flow-through mode of ion exchange chromatography. This merging eliminates the need for separate aggregate removal steps, maintaining product quality while avoiding additional process complexity and time consumption.
Solution Approach 2:
The CPG material serves multiple functions: it acts as both a filtration medium for aggregate removal and a chromatography support for purification. This multi-functionality allows a single step to achieve both aggregate removal and purification, improving product quality without adding process complexity.
4Reliability
If additional purification steps are added to remove aggregates, then product quality improves, but processing time increases
Solution Approach 1:
The patent merges aggregate removal with ion exchange chromatography by using CPG as the chromatography support. Aggregates are removed in the flow-through while monomers are retained and eluted, achieving both purification and aggregate removal in a single operation that does not increase processing time.
Solution Approach 2:
The method allows aggregates to be rapidly skipped through the CPG column in the flow-through fraction, separating them quickly from monomers. This rapid separation mechanism minimizes processing time while effectively improving product quality by removing aggregates.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This method effectively separates monomeric from aggregated immunoglobulins, maintaining the antigen-binding activity of the monomeric form and simplifying the purification process by achieving high yields of monomeric immunoglobulin while minimizing the formation of aggregates during incubation with uCPG.
Implementation Method 1
underivatized controlled pore glass (uCPG) surfaces selectively bind dimeric and oligomeric, i.e. aggregated, immunoglobulin of class G (IgG) present in a solution
Data Source
Figure 1A~1B
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AI summary
For the removal of high molecular weight compounds from recombinantly produced polypeptides generally chromatographic methods are employed. It has been found that underivatized controlled pore glass (uCPG) selectively binds high molecular weight compounds present in a solution. The purified polypeptide can be recovered e.g. from the flow through of a chromatography column containing uCPG as chromatography material. It has been found that this effect is pronounced at a pH value of about 4 to 6 in buffered solutions. With approximately 100 m2 to 150 m2 uCPG surface per g of polypeptide almost 80 % to 95 % of the high molecular weight compounds are removed with a yield of 80 % to 90 % of polypeptide.