| Autor: |
Pearl Akamine, José A. González-Feliciano, Ruth Almodóvar, Gloriner Morell, Javier Rivera, Coral M. Capó-Vélez, Manuel Delgado-Vélez, Luis Prieto-Costas, Bismark Madera, Daniel Eichinger, Ignacio Pino, José H. Rivera, José Ortiz-Ubarri, José M. Rivera, Abel Baerga-Ortiz, José A. Lasalde-Dominicci |
| Jazyk: |
angličtina |
| Rok vydání: |
2023 |
| Předmět: |
|
| Zdroj: |
Vaccines, Vol 11, Iss 5, p 975 (2023) |
| Druh dokumentu: |
article |
| ISSN: |
2076-393X |
| DOI: |
10.3390/vaccines11050975 |
| Popis: |
We have developed a pipeline to express, purify, and characterize HIV envelope protein (Env) gp145 from Chinese hamster ovary cells, to accelerate the production of a promising vaccine candidate. First in shake flasks, then in bioreactors, we optimized the growth conditions. By adjusting the pH to 6.8, we increased expression levels to 101 mg/L in a 50 L bioreactor, nearly twice the previously reported titer value. A battery of analytical methods was developed in accordance with current good manufacturing practices to ensure a quality biopharmaceutical. Imaged capillary isoelectric focusing verified proper glycosylation of gp145; dynamic light scattering confirmed the trimeric arrangement; and bio-layer interferometry and circular dichroism analysis demonstrated native-like properties (i.e., antibody binding and secondary structure). MALDI-TOF mass spectrometry was used as a multi-attribute platform for accurate mass determination, glycans analysis, and protein identification. Our robust analysis demonstrates that our gp145 product is very similar to a reference standard and emphasizes the importance of accurate characterization of a highly heterogeneous immunogen for the development of an effective vaccine. Finally, we present a novel guanosine microparticle with gp145 encapsulated and displayed on its surface. The unique properties of our gp145 microparticle make it amenable to use in future preclinical and clinical trials. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
|
