The ALPI Knockout HeLa Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population designed for loss-of-function investigation of the human ALPI gene. This product comprises a heterogenous pool of edited cells with targeted disruption of the ALPI locus, eliminating intestinal alkaline phosphatase expression. As a polyclonal knockout, it captures a range of genetic modifications, enabling robust and reproducible phenotypic analysis without the clonal variability often associated with single-cell-derived lines.
The parental cell line, HeLa, is an immortalized human cervical adenocarcinoma cell line widely utilized in biomedical research. Originally derived from an epithelial carcinoma, HeLa cells exhibit robust growth characteristics and are amenable to a variety of genetic manipulations. Their well-characterized signaling pathways and responsiveness to extracellular stimuli make them a versatile host for studying gene function in a non-intestinal epithelial context.
ALPI encodes intestinal alkaline phosphatase (IAP), a glycosylphosphatidylinositol-anchored enzyme that hydrolyzes phosphate monoesters from bacterial lipopolysaccharide (LPS) and flagellin, thereby detoxifying these pathogen-associated molecular patterns and maintaining intestinal barrier homeostasis. IAP activity is transcriptionally regulated by butyrate, retinoic acid, vitamin D, and transcription factors CDX2 and HNF4A. By dephosphorylating LPS, IAP prevents excessive activation of the Toll-like receptor 4 (TLR4) signaling cascade, which includes the adaptor MyD88, kinases IRAK and TRAF6, and the IKK complex, ultimately suppressing NF-??B-mediated transcription of pro-inflammatory cytokines such as IL-6 and TNF-??. Thus, ALPI knockout abolishes IAP activity, enhancing TLR4-driven NF-??B activation and inflammatory cytokine production.
In the HeLa cell background, which does not endogenously express intestinal alkaline phosphatase, ALPI knockout provides a clean loss-of-function model free from confounding IAP activity. This is particularly advantageous for dissecting LPS-induced TLR4 signaling and NF-??B activation in a non-intestinal epithelial environment. The absence of ALPI in HeLa cells allows researchers to directly assess the impact of LPS stimulation on downstream pathways, including I??B degradation and NF-??B nuclear translocation, without interference from endogenous phosphatase-mediated detoxification.
These polyclonal knockout cells are suitable for a wide array of applications, including functional characterization of human ALPI, evaluation of LPS detoxification in non-intestinal cells, and screening of ALPI modulators. They also serve as a valuable tool for studying inflammation-related signaling pathways and for generating an ALPI-deficient background in complementation studies. Representative assays include alkaline phosphatase enzymatic activity measurements using pNPP, western blotting for ALPI, phospho-NF-??B, and I??B, cytokine ELISA for IL-6 and TNF-??, RT-qPCR for inflammatory markers, LPS detoxification assays (Limulus amebocyte lysate), and immunofluorescence for NF-??B nuclear translocation. For further inquiries, please contact Ascent Research.
