Gel filtration chromatography as a method for removing bacterial endotoxin from antibody preparations

London, Anne Serdakowski; Mackay, Kasey; Lihon, Michael; He, Yaqin; Alabi, Busola Ruth

doi:10.1002/btpr.1961

Cited by 20 publications

(15 citation statements)

References 18 publications

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“…Size-exclusion chromatography (SEC) is a method that could be applied to eliminate endotoxin based on its size and molecular weight. SEC has been used for separating endotoxin from proteins and plasmid DNA [94,95]. However, as mentioned above, the wide range of the endotoxin size and molecular weight makes it difficult to remove all the endotoxin with this method (0.8 EU/ mg still present in the purified sample; see [95]).…”

Section: Gel Filtration Chromatographymentioning

confidence: 99%

Endotoxin Contamination: A Key Element in the Interpretation of Nanosafety Studies

Boraschi

2016

Nanomedicine (Lond.)

168

147

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The study of toxicity and potential risks of engineered nanoparticles is of particular importance in nanomedicine. Endotoxin, a common contaminant of bacterial origin, has biological effects that can mask the true biological effects of nanoparticles, if its presence is overlooked. In this review, we report the features of nanoparticle contamination by endotoxin, and the different biological effects of endotoxin-contaminated nanoparticles. We will describe different methods for endotoxin detection applied to nanoparticles, and discuss their pros and cons. Eventually, we describe various methods for eliminating endotoxin contamination in nanoparticles. Although there is no universal technique for efficiently removing endotoxin from nanoparticles, specific solutions can be found case by case, which can allow us to perform nanosafety studies in biologically relevant conditions.

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Section: Gel Filtration Chromatographymentioning

confidence: 99%

Endotoxin Contamination: A Key Element in the Interpretation of Nanosafety Studies

Boraschi

2016

Nanomedicine (Lond.)

168

147

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“…Having a molecular mass greater than 10 kDa, lipopolysaccharides (LPS) resulting from the biotechnological processes are large molecules, which sediment during the ultracentrifugation step. In the less probable situation when they are still present in the clarified lysate, SEC is a known method for their elimination 33 , 34 , and that is the reason for the lower endotoxin level detected in our NMN sample compared with the HPLC solvents used. Evidently, an even lower endotoxin level can be obtained by using endotoxin free solvents.…”

Section: Discussionmentioning

confidence: 93%

Size Exclusion Chromatography Method for Purification of Nicotinamide Mononucleotide (NMN) from Bacterial Cells

Marinescu

Popescu

Dinischiotu

2018

Sci Rep

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Over 12% of the world’s health resources are spent on treating diabetes, as high blood glucose is the third cause of mortality worldwide. Insulin resistance is the basis of the most common form of diabetes: type 2 diabetes. Recent animal studies report successful attempts at reversing type 2 diabetes by the administering of the NAD+ precursor nicotinamide mononucleotide (NMN). However, the current high price of this molecule urges for more efficient and cost-effective production methods. This work proposes a method for purifying NMN by Size Exclusion Chromatography (SEC) on silica with a covalently attached coating of poly(2-hydroxyethyl aspartamide) (PolyHEA) stationary phase using an isocratic elution with a denaturing mobile phase (50 mM formic acid) from a complex molecular mixture such as a fermentation broth. The eluted peaks were identified by UV-Vis analysis and confirmed with ESI+ mass spectrometry and a HPLC reversed-phase method. The proposed SEC method is simple, patent-free, directly applicable for industrial production with a minimum scale up effort. The need for multiple chromatographic steps is eliminated and the lysate filtration and clarification steps are simplified. Substantial reduction in NMN production costs and increased purity of NMN to the level suitable for usage in humans are expected.

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“…The purified protein was collected and gel filtration chromatography (cat no. 17014901; GE Healthcare Life Sciences) was used to further purify Anti-SEMA-7A, as described previously ( 21 ).…”

Section: Methodsmentioning

confidence: 99%

Anti-Semaphorin-7A single chain antibody demonstrates beneficial effects on pulmonary inflammation during acute lung injury

et al. 2018

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Pulmonary inflammation is a primary characteristic of lung injury initiated by the accession of immune cells into the alveolar space. Neutrophil migration serves an important role in pulmonary inflammation mediated by the migration of neutrophils into hypoxic tissue sites. The elimination of pulmonary inflammation is directly associated with rehabilitation in patients with lung injury. Anti-inflammatory treatment is essential following lung injury and ultimately determines patient outcomes. Semaphorin-7A (SEMA-7A) is a member of the Semaphorin family that influences the migration of neutrophils into hypoxic tissue sites, thus promoting inflammation. However, understanding of the role of SEMA-7A serves during lung injury is limited and the immunological function of SEMA-7A during the migration of neutrophils into acute injury sites remains unknown. The present study investigated SEMA-7A expression and constructed a single chain antibody for SEMA-7A (Anti-SEMA-7A) to study its therapeutic efficacy against pulmonary inflammation in a mouse model of acute injury sites. The data indicated that the expression of SEMA-7A was upregulated due to induction by pro-inflammatory cytokines and demonstrated that Anti-SEMA-7A inhibited SEMA-7A expression in vitro and in vivo. The current study also indicated that the production of pro-inflammatory cytokines induced by SEMA-7A in endothelial and epithelial cells enhanced pulmonary inflammation. Anti-SEMA-7A suppressed the transendothelial migration of neutrophils mediated by SEMA-7A. Anti-SEMA-7A treatment neutralized SEMA-7A expression and reduced signs of pulmonary inflammation, leading to the elimination of pulmonary inflammation in rat with acute lung injury. The current study identified Anti-SEMA-7A as a potential agent to interfere with the inflammatory pathway during acute lung injury, which may be the basis for anti-inflammatory strategies to treat lung injuries in the future.

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Gel filtration chromatography as a method for removing bacterial endotoxin from antibody preparations

Cited by 20 publications

References 18 publications

Endotoxin Contamination: A Key Element in the Interpretation of Nanosafety Studies

Endotoxin Contamination: A Key Element in the Interpretation of Nanosafety Studies

Size Exclusion Chromatography Method for Purification of Nicotinamide Mononucleotide (NMN) from Bacterial Cells

Anti-Semaphorin-7A single chain antibody demonstrates beneficial effects on pulmonary inflammation during acute lung injury

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