Development of a sensitive size exclusion HPLC method with fluorescence detection for the quantitation of recombinant human erythropoietin (r-HuEPO) aggregates

Gunturi, Srinivas R.; Ghobrial, I. A.; Sharma, Basant

doi:10.1016/j.jpba.2006.06.006

Cited by 32 publications

(14 citation statements)

References 16 publications

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“…[17,86–88] Diress et al [86] demonstrated the utility of fluorescence detection for improved sensitivity in cases where excipients may elute near or with the protein of interest. Gunturi et al [87] also showed the sensitivity of fluorescence detection for recombinant human growth hormone, also in the presence of excipients. These studies confirm the applicability of fluorescence detectors to measure low level of aggregates.…”

Section: Detectorsmentioning

confidence: 99%

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A Review Size-Exclusion Chromatography for the Analysis of Protein Biotherapeutics and Their Aggregates

Hong

Koza

Bouvier

2012

Journal of Liquid Chromatography & Related Technologies

459

291

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In recent years, the use and number of biotherapeutics has increased significantly. For these largely protein-based therapies, the quantitation of aggregates is of particular concern given their potential effect on efficacy and immunogenicity. This need has renewed interest in size-exclusion chromatography (SEC). In the following review we will outline the history and background of SEC for the analysis of proteins. We will also discuss the instrumentation for these analyses, including the use of different types of detectors. Method development for protein analysis by SEC will also be outlined, including the effect of mobile phase and column parameters (column length, pore size). We will also review some of the applications of this mode of separation that are of particular importance to protein biopharmaceutical development and highlight some considerations in their implementation.

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Section: Detectorsmentioning

confidence: 99%

“…To address this limitation alternative UV absorbance wavelengths or fluorescence can be used. [19,87] …”

Section: Applicationsmentioning

confidence: 99%

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A Review Size-Exclusion Chromatography for the Analysis of Protein Biotherapeutics and Their Aggregates

Hong

Koza

Bouvier

2012

Journal of Liquid Chromatography & Related Technologies

459

291

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“…These include external stabilization by influencing the properties of the surrounding solvent through the use of stabilizing excipients (e.g., amino acids, sugars, polyols) and internal stabilization by altering the structural characteristics of the protein through chemical modifications (e.g., mutations, glycosylation, pegylation) 2, 53, 58. While many protein pharmaceuticals have been successfully formulated by employing stabilizing mutations, excipients, and pegylation, their use can sometimes be problematic due to limitations, such as, predicting the stabilizing nature of amino acid substitutions, the occurrence of protein and excipient dependant nongeneralized stabilization effects, protein/excipient phase separation upon freezing, cross‐reactions between some excipients and the multiple chemical functionalities present in proteins, acceleration of certain chemical (e.g., aspartate isomerization) and physical (e.g., aggregation) instabilities by some excipients (e.g., sorbitol, glycerol, sucrose), detection interferences caused by some sugar excipients during various protein analysis methods, and safety concerns regarding the long‐term use of pegylated proteins in vivo due to possible PEG induced immunogenecity and chronic accumulation toxicity resulting from its reduced degradation and clearance rates 2, 4, 33, 48, 66, 78–95…”

Section: Introductionmentioning

confidence: 99%

Long‐distance corticocortical GABAergic neurons in the adult monkey white and gray matter

Tomioka

Rockland

2007

J of Comparative Neurology

122

125

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A subgroup of GABAergic neurons has been reported to project over long distances in several species. Here we demonstrate that long-distance cortically projecting nonpyramidal neurons occur in monkeys in both white and gray matter. Nonpyramidal neurons were first identified morphologically. Visualization of Golgi-like details was achieved by retrograde infection from injections of an adenovirus vector, producing enhanced green fluorescent protein (EGFP) under control of a neuron-specific promoter. Injections in areas V1, V4, TEO, and posterior TE resulted in EGFP-expressing nonpyramidal neurons up to 1.5 cm distant from the injections, mainly in the white matter. Some neurons occurred in the gray matter, mainly in layer 3, but also in layers 5 and 6, and, very occasionally, layer 1. As control, we injected cholera toxin subunit B, a standard retrograde tracer, in V4, and observed a similarly wide distribution of neurons in the white matter. Second, the GABAergic identity of EGFP-expressing nonpyramidal neurons was established by colabeling for EGFP and GAD67 in selected tissue sections. Most neurons positive for EGFP and GAD67 were positive for somatostatin (SS; 90%). Of those neurons positive for EGFP and SS, almost all were also positive for neuronal nitric oxide synthase or m2 muscarinic receptor, but only 23% were also positive for calretinin. None were positive for parvalbumin. We conclude that long-distance projecting GABAergic neurons 1) are phylogenetically conserved, although in monkeys most gray matter neurons are in the upper layers, and 2) are heterogeneous in terms of their neurochemistry, location, and potentially function.

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“…Detection of protein aggregates can be accomplished by traditional methods such as SEC (Size Exclusion Chromatography) [17] or ELISA (Enzyme Linked Immuno Sorbent Assay) [2]. Analytical ultra centrifugation, although technically challenging, has been described as the 'gold standard' [32] for the analysis of aggregates.…”

Section: Analytical Methods To Promote Quality and Prevent Immunogenimentioning

confidence: 99%

Quality and safety considerations for recombinant biological medicines: A regulatory perspective

Barnes¹,

Ragnarsson²,

Alván³

2009

International Journal of Risk and Safety in Medicine

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Biological medicines have introduced a new set of quality and safety issues because these medicinal products are large molecules synthesized in complex biological processes and may have pharmacodynamic properties that need extra consideration and study for safe administration to humans. A complex manufacturing process and molecular structure means that the exact identity of the material cannot be guaranteed by analytical technology alone but requires a well understood and reproducible manufacturing process. Furthermore, the production systems utilizing recombinant DNA technology in living cell systems are influenced by a multitude of factors that are not easily controlled. The result is that identity between manufactured lots of one product or between similar products (i.e., biosimilars) is difficult to confirm. Purification and analytical methods such as HPLC, CE, UPLC, SEC, ELISA, ELISpot, SPR and the transgenic mouse immunogenicity model are briefly reviewed as means of achieving the highest possible degree of safety for patients using biological medicines.

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Development of a sensitive size exclusion HPLC method with fluorescence detection for the quantitation of recombinant human erythropoietin (r-HuEPO) aggregates

Cited by 32 publications

References 16 publications

A Review Size-Exclusion Chromatography for the Analysis of Protein Biotherapeutics and Their Aggregates

A Review Size-Exclusion Chromatography for the Analysis of Protein Biotherapeutics and Their Aggregates

Long‐distance corticocortical GABAergic neurons in the adult monkey white and gray matter

Quality and safety considerations for recombinant biological medicines: A regulatory perspective

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