Protocol for high-throughput reservoir quantification across global HIV subtypes using a cross-subtype intact proviral DNA assay

Summary: The cross-subtype intact proviral DNA assay (CS-IPDA) is a high-throughput method to quantify HIV reservoir size in populations infected with any of the dominant global HIV-1 subtypes. Our protocol includes genomic DNA isolation optimized to minimize DNA shearing, a reference droplet digita...

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Bibliographic Details
Main Authors: Carolyn S. Fish, Noah A.J. Cassidy, Claire N. Levy, Sean M. Hughes, Keith R. Jerome, Julie Overbaugh, Florian Hladik, Dara A. Lehman
Format: Article
Language:English
Published: Elsevier 2022-12-01
Series:STAR Protocols
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Online Access:http://www.sciencedirect.com/science/article/pii/S2666166722005615
Description
Summary:Summary: The cross-subtype intact proviral DNA assay (CS-IPDA) is a high-throughput method to quantify HIV reservoir size in populations infected with any of the dominant global HIV-1 subtypes. Our protocol includes genomic DNA isolation optimized to minimize DNA shearing, a reference droplet digital PCR (ddPCR) assay to quantify T cells and assess DNA shearing, and a multiplex ddPCR targeting three distinct regions across the HIV genome to quantify intact proviruses as an estimate of replication-competent proviruses in the reservoir.For complete details on the use and execution of this protocol, please refer to Cassidy et al. (2022). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
ISSN:2666-1667