The CD4 Knockout THP-1 Polyclonal Cells product provides a CRISPR/Cas9-mediated gene-disrupted polyclonal population targeting the CD4 locus in the THP-1 human monocytic leukemia cell line. This pool of edited cells serves as a loss-of-function model for investigating CD4-dependent processes, including T-cell co-receptor signaling and HIV-1 entry mechanisms. By introducing targeted disruption into the CD4 gene, the polyclonal knockout population enables robust and reproducible studies without requiring monoclonal isolation, offering a maximally diverse genetic background for rapid phenotypic analysis and functional assays in hematological and immunological research contexts.
The THP-1 host cell line was originally isolated from the peripheral blood of a pediatric patient with acute monocytic leukemia and has since become a widely utilized suspension cell model for studying monocyte and macrophage biology, differentiation pathways, and innate immune responses. THP-1 cells retain many features of native monocytes, including the capacity to differentiate into macrophage-like or dendritic cell-like phenotypes upon stimulation with phorbol esters or cytokines, making them a versatile platform for probing cell-type-specific functions. Their robust growth characteristics and amenability to genetic manipulation further support high-throughput screening and detailed mechanistic dissection.
CD4 encodes a glycoprotein co-receptor that critically augments T-cell receptor (TCR) signaling and is the primary cellular receptor for HIV-1 entry. In T-lymphocytes, CD4 interacts with the TCR/CD3 complex and binds MHC class II molecules on antigen-presenting cells, recruiting and activating the Src-family kinase Lck. Upon TCR stimulation, Lck phosphorylates immunoreceptor tyrosine-based activation motifs (ITAMs) within CD3 chains, leading to ZAP70 recruitment, activation, and subsequent phosphorylation of the adaptor protein LAT. This nucleates a signalosome that propagates downstream through PLC-??1 to generate second messengers IP3 and DAG, triggering calcium mobilization and activation of transcription factors such as NFAT and NF-??B. Concurrently, MAP kinase cascades, including ERK, are engaged. CD4 activity is regulated by TCR stimulation, cytokines such as IL-16, and transcriptional regulators including RUNX1 and Ikaros. In the context of HIV-1 infection, CD4 directly binds the viral envelope glycoprotein gp120, facilitating membrane fusion and viral entry, a process that can be modeled in THP-1 cells because CD4 expression is retained and the cells are permissive to HIV-1 replication.
In THP-1 cells, CD4 knockout has particular significance given the monocytic lineage. Although THP-1 cells are not classical T lymphocytes, they express CD4 and can be efficiently infected by HIV-1 laboratory strains, making them a valuable surrogate system to study viral entry, post-entry events, and the interplay between viral proteins and myeloid cell signaling. Disruption of CD4 in this background allows researchers to dissect CD4-dependent versus -independent mechanisms of HIV-1 infection and to evaluate host factors involved in viral replication or latency. Additionally, because THP-1 cells model macrophage immune functions, the knockout provides insight into how CD4-mediated signals may modulate monocyte/macrophage activation, cytokine production, or antigen presentation capabilities, expanding the utility beyond lymphocyte-centric paradigms.
This knockout pool is ideally suited for a broad range of experimental applications. In HIV research, it enables pseudovirus infection assays and drug screening for entry inhibitors, where loss of the receptor eliminates background signal. For T-cell signaling pathway analysis, users can perform western blotting for CD4 downstream effectors such as phospho-Lck, ZAP70, or ERK, or employ NFAT reporter assays to quantify pathway activation. Co-immunoprecipitation studies can map CD4 interactions with Lck or MHC class II, while RT-qPCR validates gene expression changes upon CD4 disruption. Furthermore, the polyclonal format supports flow cytometric assessment of CD4 surface expression to confirm knockout, and the cells are suitable for functional immune assays examining monocyte/macrophage responses such as cytokine profiling or phagocytosis. For further details or technical support, please contact Ascent Research.
