toward the development of a single-round infection assay based on egfp reporting for anti-hiv-1 drug discovery
نویسندگان
چکیده
background: the rapid increase of hiv-1 strains resistant to current antiretroviral drugs is a challenge for successful aids therapy. this necessitates the development of novel drugs, and to this end, availability of screening systems for in vitro drug discovery is a priority. herein, we report the modification of a previously developed system for increased sensitivity, ease of use, and cost-efficiency, based on the application of the egfp marker. methods: a pcr-amplified gfp gene (gfp) was cloned into pmznl4-3, the plasmid already designed to produce single-cycle replicable virions, in frame with the reverse-transcriptase gene to construct the pmznl4-3/gfp plasmid. gfp-mznl4-3 pseudo-typed virions, as the first progeny viruses, were recovered from the culture supernatant of hek293t cells co-transfected with pmznl4-3/gfp and the helper plasmids pspax2 and pmd2g, which respectively encode hiv-1 gag-pol and vesicular stomatitis virus glycoprotein. single-cycle replication and virion production were assessed by syncytia formation, p24 antigen assays, and electron and fluorescence microscopy. results: the incorporation of egfp into the viral particles allowed their quantification by fluorometry, flow-cytometry, and fluorescence microscopy; however, this modification did not affect the single-round infectivity or production rate of the gfp fluorescence-emitting virions. conclusions: our results certify the development of a rapid, inexpensive, and safe gfp-reporting single-cycle replicable system for anti-hiv drug discovery. further experiments are needed to measure the validity and robustness of the assay.
منابع مشابه
Toward the Development of a Single-Round Infection Assay Based on EGFP Reporting for Anti-HIV-1 Drug Discovery
Background: The rapid increase of HIV-1 strains resistant to current antiretroviral drugs is a challenge for successful AIDS therapy. This necessitates the development of novel drugs, and to this end, availability of screening systems for in vitro drug discovery is a priority. Herein, we report the modification of a previously developed system for increased sensitivity, ease of use, and cost-ef...
متن کامل***Removed from the Issue*** Application of Cell-Based Assay Systems for the Early Screening of Human Drug Hepatotoxicity in the Discovery Phase of Drug Development
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***Removed from the Issue*** Application of Cell-Based Assay Systems for the Early Screening of Human Drug Hepatotoxicity in the Discovery Phase of Drug Development
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Withdrawing of Article ''Application of Cell-Based Assay Systems for the Early Screening of Human Drug Hepatotoxicity in the Discovery Phase of Drug Development''
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A novel medium-throughput biological assay system for HTLV-1 infectivity and drug discovery
Objective(s): Here, a reporter cell line containing two reporter vectors were developed, in order to monitor the Human T-Lymphotropic Virus type1(HTLV-1) infectivity and the cell viability simultaneously. Materials and Methods: The reporter cell line was constructed by stably transfected baby hamster's kidney cell line (BHK-21), with the genomes expressing two different reporters in separate pl...
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عنوان ژورنال:
reports of biochemistry and molecular biologyجلد ۴، شماره ۱، صفحات ۱-۹
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