Study of chain branching in natural rubber using size-exclusion chromatography coupled with a multi-angle light scattering detector (SEC-MALS)

ABSTRACT Mechanical or thermal mastication experiments were performed on three commercial natural rubber (NR) samples of TSR10 grade made from latex of three different clones (GT1, PB235, and RRIM600). The mesostructure (different gel or aggregate fractions, structure of random coils of cis-1,4-polyisoprene) of all the NR samples was fully characterized by size exclusion chromatography coupled with multiangle light scattering (SEC-MALS), using pretreated SEC columns. This method was used to quantify and investigate the structure of the little-studied smaller microaggregates, constituting the microgel fraction smaller than 1 μm (Microgel<1μ) of NR. The three unmasticated NR samples showed no difference in terms of microaggregate structure. Conversely, microaggregates appeared denser after mastication. This phenomenon was found to depend on the mastication conditions, as mechanically masticated NR samples had smaller (lower radius of gyration) and more compact microaggregates than thermally masticated samples. Macrogel also behaved differently depending on the mastication conditions. Mechanical mastication conditions allowed a higher degradation of the macrogel compared with thermal mastication conditions.

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Comprehensive characterization and quantification of adeno associated vectors by size exclusion chromatography and multi angle light scattering

Scientific Reports ◽

10.1038/s41598-021-82599-1 ◽

2021 ◽

Vol 11 (1) ◽

Cited By ~ 1

Author(s):

Nicole L. McIntosh ◽

Geoffrey Y. Berguig ◽

Omair A. Karim ◽

Christa L. Cortesio ◽

Rolando De Angelis ◽

...

Keyword(s):

Light Scattering ◽

Size Exclusion Chromatography ◽

Molar Mass ◽

Size Exclusion ◽

Minimal Sample ◽

Accuracy And Precision ◽

Vector Genome ◽

Exclusion Chromatography ◽

Novel Applications

AbstractAdeno associated virus (AAV) capsids are a leading modality for in vivo gene delivery. Complete and precise characterization of capsid particles, including capsid and vector genome concentration, is necessary to safely and efficaciously dose patients. Size exclusion chromatography (SEC) coupled to multiangle light scattering (MALS) offers a straightforward approach to comprehensively characterize AAV capsids. The current study demonstrates that this method provides detailed AAV characterization information, including but not limited to aggregation profile, size-distribution, capsid content, capsid molar mass, encapsidated DNA molar mass, and total capsid and vector genome titer. Currently, multiple techniques are required to generate this information, with varying accuracy and precision. In the current study, a new series of equations for SEC-MALS are used in tandem with intrinsic properties of the capsids and encapsidated DNA to quantify multiple physical AAV attributes in one 20-min run with minimal sample manipulation, high accuracy, and high precision. These novel applications designate this well-established method as a powerful tool for product development and process analytics in future gene therapy programs.

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