Comparative analysis of protein quantification methods for the rapid determination of protein loading in liposomal formulations

Maryam T. Hussain, Neil Forbes, Yvonne Perrie

Research output: Contribution to journalArticle

5 Citations (Scopus)
13 Downloads (Pure)

Abstract

Advances in manufacturing processes provide the ability for the high throughput production of liposomes containing a range of moieties, from small molecules to large biologicals (including proteins and nucleic acids for prophylactic and therapeutic applications). Whilst rapid quantification methods for small molecules are generally well established, the ability to rapidly quantify liposomal entrapment of proteins is limited. Indeed, most standard protein quantification techniques (including the BCA assay and Reverse phase-high performance liquid chromatography (RP-HPLC)) measure protein encapsulation indirectly, by measuring the amount of non-incorporated drug, and subtracting from the initial amount of protein added. However, this can give inaccurate and misrepresentative results. To address this, we have developed a range of methods to directly quantify protein entrapment within liposomes. The encapsulation efficiency within neutral, anionic and cationic liposome formulations was determined by three techniques; BCA assay, RP-HPLC and HPLC coupled to an evaporative light scattering detector, (HPLC-ELSD). All three methods are reliable for the quantification of protein, with linear responses and correlation coefficients of 0.99, and LOQ for all three methods being less than 10 µg/mL. Here within, we provide three methods for the rapid and robust quantification of protein loading within liposomal (and other bilayer) vesicle systems.

Original languageEnglish
Article number39
Number of pages16
JournalPharmaceutics
Volume11
Issue number1
DOIs
Publication statusPublished - 18 Jan 2019

Keywords

  • microfluidics
  • liposome
  • solubilisation
  • protein quantification

Projects

Research Output

  • 5 Citations
  • 1 Article

Rapid and scale-independent microfluidic manufacture of liposomes entrapping protein incorporating in-line purification and at-line size monitoring

Forbes, N., Hussain, M. T., Briuglia, M. L., Edwards, D. P., ter Horst, J. H., Szita, N. & Perrie, Y., 10 Feb 2019, In : International Journal of Pharmaceutics. 556, p. 68-81 14 p.

Research output: Contribution to journalArticle

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27 Citations (Scopus)
18 Downloads (Pure)

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