Human Immunodeficiency Virus (HIV) Separation and Enrichment via the Combination of Antiviral Lectin Recognition and a Thermoresponsive Reagent System.
Pharm Res
; 33(10): 2411-20, 2016 10.
Article
em En
| MEDLINE
| ID: mdl-27401412
ABSTRACT
PURPOSE:
In order to improve the detection limit of existing HIV diagnostic assays, we explored the use of a temperature-responsive magnetic nanoparticle reagent system in conjunction with cyanovirin-N for HIV recognition to rapidly and efficiently concentrate viral particles from larger sample volumes, ~ 1 ml.METHODS:
Cyanovirin-N (CVN) mutant, Q62C, was expressed, biotinylated, and then complexed with a thermally responsive polymer-streptavidin conjugate. Confirmation of protein expression/activity was performed using matrix assisted laser desorption/ionization (MALDI) and a TZM-bl HIV inhibition assay. Biotinylated CVN mutant recognition with gp120 was characterized using surface plasmon resonance (SPR). Virus separation and enrichment using a thermoresponsive magnetic nanoparticle reagent system were measured using RT-PCR.RESULTS:
Biotinylated Q62C exhibited a KD of 0.6 nM to gp120. The temperature-responsive binary reagent system achieved a maximum viral capture of nearly 100% HIV, 1 × 10(5) virus copies in 100 µl, using pNIPAAm-Q62C within 30 minutes. Additionally, the same reagent system achieved nearly 9-fold enrichment by processing a 10-times larger sample of 1000 µl (Fig. 3).CONCLUSION:
This work demonstrated a temperature-responsive reagent system that provides enrichment of HIV using antiviral lectin CVN for recognition, which is potentially amenable for use in point-of-care settings.Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Temas:
Geral
Base de dados:
MEDLINE
Assunto principal:
Antivirais
/
Proteínas de Bactérias
/
Proteínas de Transporte
/
HIV-1
/
Lectinas
Limite:
Humans
Idioma:
En
Revista:
Pharm Res
Ano de publicação:
2016
Tipo de documento:
Article
País de afiliação:
Estados Unidos