Monolith affinity chromatography for the rapid quantification of a single-chain variable fragment immunotoxin
(2018) In Journal of Separation Science 41(15). p.3051-3059- Abstract
We developed a novel analytical method for concentration determination of tandem single-chain antibody diphtheria toxin (immunotoxin). The method is based on polymethacrylate monoliths with Protein L ligands as the binding moiety. Different buffers were tested for elution of the Protein L-bound immunotoxin and 4.5 M guanidinium hydrochloride performed best. We optimized the elution conditions and the method sequence resulting in a fast and robust method with a runtime <10 min. Fast determination of immunotoxin is critical if any process decisions rely on this data. We determined method performance and a lower limit of detection of 27 μg/mL and a lower limit of quantification of 90 μg/mL was achieved. The validity of the method in... (More)
We developed a novel analytical method for concentration determination of tandem single-chain antibody diphtheria toxin (immunotoxin). The method is based on polymethacrylate monoliths with Protein L ligands as the binding moiety. Different buffers were tested for elution of the Protein L-bound immunotoxin and 4.5 M guanidinium hydrochloride performed best. We optimized the elution conditions and the method sequence resulting in a fast and robust method with a runtime <10 min. Fast determination of immunotoxin is critical if any process decisions rely on this data. We determined method performance and a lower limit of detection of 27 μg/mL and a lower limit of quantification of 90 μg/mL was achieved. The validity of the method in terms of residual analysis, precision, and repeatability was proven in a range from 100 to 375 μg/mL. The short runtime and ease of use of a high-performance liquid chromatography method is especially useful for a process analytical tool approach. Bioprocesses related to immunotoxin where fermentation or other process parameters can be adjusted in accordance to the immunotoxin levels will be benefited from this method to achieve the highest possible purity and productivity.
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- author
- Satzer, Peter ; Sommer, Ralf ; Paulsson, Johanna LU ; Rodler, Agnes ; Zehetner, Romana ; Hofstädter, Klaus ; Klade, Christoph and Jungbauer, Alois
- organization
- publishing date
- 2018-08-01
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- affinity chromatography, antibody-drug conjugates, immunotoxins, monoliths, process analytical tools
- in
- Journal of Separation Science
- volume
- 41
- issue
- 15
- pages
- 9 pages
- publisher
- John Wiley & Sons Inc.
- external identifiers
-
- pmid:29873445
- scopus:85050611247
- ISSN
- 1615-9306
- DOI
- 10.1002/jssc.201800257
- language
- English
- LU publication?
- yes
- id
- a7de60ec-7def-491a-9597-c6315e18263f
- date added to LUP
- 2018-10-01 09:51:47
- date last changed
- 2024-06-24 20:09:12
@article{a7de60ec-7def-491a-9597-c6315e18263f,
abstract = {{<p>We developed a novel analytical method for concentration determination of tandem single-chain antibody diphtheria toxin (immunotoxin). The method is based on polymethacrylate monoliths with Protein L ligands as the binding moiety. Different buffers were tested for elution of the Protein L-bound immunotoxin and 4.5 M guanidinium hydrochloride performed best. We optimized the elution conditions and the method sequence resulting in a fast and robust method with a runtime <10 min. Fast determination of immunotoxin is critical if any process decisions rely on this data. We determined method performance and a lower limit of detection of 27 μg/mL and a lower limit of quantification of 90 μg/mL was achieved. The validity of the method in terms of residual analysis, precision, and repeatability was proven in a range from 100 to 375 μg/mL. The short runtime and ease of use of a high-performance liquid chromatography method is especially useful for a process analytical tool approach. Bioprocesses related to immunotoxin where fermentation or other process parameters can be adjusted in accordance to the immunotoxin levels will be benefited from this method to achieve the highest possible purity and productivity.</p>}},
author = {{Satzer, Peter and Sommer, Ralf and Paulsson, Johanna and Rodler, Agnes and Zehetner, Romana and Hofstädter, Klaus and Klade, Christoph and Jungbauer, Alois}},
issn = {{1615-9306}},
keywords = {{affinity chromatography; antibody-drug conjugates; immunotoxins; monoliths; process analytical tools}},
language = {{eng}},
month = {{08}},
number = {{15}},
pages = {{3051--3059}},
publisher = {{John Wiley & Sons Inc.}},
series = {{Journal of Separation Science}},
title = {{Monolith affinity chromatography for the rapid quantification of a single-chain variable fragment immunotoxin}},
url = {{http://dx.doi.org/10.1002/jssc.201800257}},
doi = {{10.1002/jssc.201800257}},
volume = {{41}},
year = {{2018}},
}