a549-hace2-tmprss2 cells  (InvivoGen)

Journal: JID Innovations

Article Title: Human keratinocytes exhibit limited potential for SARS-CoV-2 infection despite ACE2 and mature cathepsin L expression

doi: 10.1016/j.xjidi.2025.100447

Figure Lengend Snippet: Quantification of ACE2 , dACE2 , TMPRSS2, NRP1, CTSL , BSG (CD147), DPP4 (CD26), AXL , KREMEN1 , and ASGR1 gene expression. mRNA levels normalized to GAPDH (2 -ΔCT ) in ( a ) NHDF, NHEKs, RHE, and human skin samples and in ( b ) Caco-2/TC-7, Calu-3, and HaCaT cell lines. Gene expression was quantified by RT-qPCR in triplicate. Data represent the mean ± SEM from ( a ) 3 to 14 independent experiments using biological samples from different individuals and ( b ) 3 or 4 independent experiments from cultured cells. NHDF, normal human dermal fibroblast; NHEK, normal human epidermal keratinocyte; RHE, reconstructed human epidermis.

Article Snippet: The following TaqMan MGB-FAM-labelled probes (Applied Biosystems, Thermo Fisher Scientific) were used for qPCR analysis: ACE2 (Hs01085340_m1), ACE2 + dACE2 (Hs01085333_m1), TMPRSS2 (Hs01120965_m1), NRP1 (Hs00826128_m1), CTSL (Hs00964650_m1), BSG (CD147) (Hs00936295_m1), DPP4 (Hs00897386_m1), AXL (Hs01064444_m1), KREMEN1 (Hs00230750_m1), ASGR1 (Hs01005019_m1), and reference gene GAPDH (Hs02758991_g1).

Techniques: Gene Expression, Quantitative RT-PCR, Cell Culture

Journal: JID Innovations

Article Title: Human keratinocytes exhibit limited potential for SARS-CoV-2 infection despite ACE2 and mature cathepsin L expression

doi: 10.1016/j.xjidi.2025.100447

Figure Lengend Snippet: Differential ACE2, TMPRSS2, NRP1, CTSL, CD147, AXL, and DPP4 protein expression. Whole-cell lysates of the indicated cells were analyzed by western blot using the following antibodies: ACE2 (mAb clone AC384), TMPRSS2 (mAb clone S20014A), NRP1 (mAb clone 14H4), CTSL (mAb clone 33/1), CD147 (mAb clone HIM6), AXL (polyclonal goat IgG, AF154), and DPP4 (polyclonal goat IgG, AF1180). ( a ) Representative immunoblots from 2 to 4 independent experiments. ( b ) Densitometric quantification normalized to actin, expressed as mean ± SEM from 2 to 4 independent experiments. CTSL, cathepsin L.

Article Snippet: The following TaqMan MGB-FAM-labelled probes (Applied Biosystems, Thermo Fisher Scientific) were used for qPCR analysis: ACE2 (Hs01085340_m1), ACE2 + dACE2 (Hs01085333_m1), TMPRSS2 (Hs01120965_m1), NRP1 (Hs00826128_m1), CTSL (Hs00964650_m1), BSG (CD147) (Hs00936295_m1), DPP4 (Hs00897386_m1), AXL (Hs01064444_m1), KREMEN1 (Hs00230750_m1), ASGR1 (Hs01005019_m1), and reference gene GAPDH (Hs02758991_g1).

Techniques: Expressing, Western Blot

Journal: JID Innovations

Article Title: Human keratinocytes exhibit limited potential for SARS-CoV-2 infection despite ACE2 and mature cathepsin L expression

doi: 10.1016/j.xjidi.2025.100447

Figure Lengend Snippet: Validation of flow cytometry protocols and anti-ACE2 antibodies. ( a ) Comparison of cell detachment protocols. ( b ) Comparison of anti-ACE2 antibody staining by flow cytometry. ( c, d ) Binding of 2019-nCoV Spike Protein S1 (RBD) protein and blocking with anti-ACE2 antibody (mAb clone AC384) assessed by ( c ) flow cytometry and ( d ) immunofluorescence (red). Cell nuclei are stained with DAPI (white). Bar = 10 μm. Data are representative of 3 independent experiments. ( e ) Cell surface expression of ACE2 (mAb clone Poly5036), TMPRSS2 (mAb clone S20014A), NRP1 (mAb clone U21-1283), CD147 (mAb clone HIM6), AXL (mAb clone 108724), and DPP4 (mAb clone BA5b) in A549 cells. Data are representative of 2 independent experiments.

Article Snippet: The following TaqMan MGB-FAM-labelled probes (Applied Biosystems, Thermo Fisher Scientific) were used for qPCR analysis: ACE2 (Hs01085340_m1), ACE2 + dACE2 (Hs01085333_m1), TMPRSS2 (Hs01120965_m1), NRP1 (Hs00826128_m1), CTSL (Hs00964650_m1), BSG (CD147) (Hs00936295_m1), DPP4 (Hs00897386_m1), AXL (Hs01064444_m1), KREMEN1 (Hs00230750_m1), ASGR1 (Hs01005019_m1), and reference gene GAPDH (Hs02758991_g1).

Techniques: Biomarker Discovery, Flow Cytometry, Comparison, Staining, Binding Assay, Blocking Assay, Immunofluorescence, Expressing

Journal: JID Innovations

Article Title: Human keratinocytes exhibit limited potential for SARS-CoV-2 infection despite ACE2 and mature cathepsin L expression

doi: 10.1016/j.xjidi.2025.100447

Figure Lengend Snippet: Cell surface expression of ACE2, TMPRSS2, NRP1, CD147, AXL, and DPP4 receptors is associated with exclusive binding of SARS-CoV-2 Spike Protein S1 (RBD) to ACE2-expressing cells. ( a ) Flow cytometry analysis of cell surface expression of ACE2 (mAb clone Poly5036), TMPRSS2 (mAb clone S20014A), NRP1 (mAb clone U21-1283), CD147 (mAb clone HIM6), AXL (mAb clone 108724), and DPP4 (mAb clone BA5b) in the indicated cell lines and primary skin cells. Data are representative of 2–4 independent experiments. ( b, c ) Representative immunofluorescence microscopy images showing ( b ) ACE2 expression (mAb clone Poly5036) and ( c ) binding of 2019-nCoV Spike Protein S1 (RBD) in red. Cell nuclei are stained with DAPI (white). Bar = 10 μm. The proportions of cells expressing ACE2 and binding to spike are shown. Data are representative of at least 2 independent experiments. SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.

Article Snippet: The following TaqMan MGB-FAM-labelled probes (Applied Biosystems, Thermo Fisher Scientific) were used for qPCR analysis: ACE2 (Hs01085340_m1), ACE2 + dACE2 (Hs01085333_m1), TMPRSS2 (Hs01120965_m1), NRP1 (Hs00826128_m1), CTSL (Hs00964650_m1), BSG (CD147) (Hs00936295_m1), DPP4 (Hs00897386_m1), AXL (Hs01064444_m1), KREMEN1 (Hs00230750_m1), ASGR1 (Hs01005019_m1), and reference gene GAPDH (Hs02758991_g1).

Techniques: Expressing, Binding Assay, Flow Cytometry, Immunofluorescence, Microscopy, Staining

Journal: JID Innovations

Article Title: Human keratinocytes exhibit limited potential for SARS-CoV-2 infection despite ACE2 and mature cathepsin L expression

doi: 10.1016/j.xjidi.2025.100447

Figure Lengend Snippet: TLR3-mediated activation upregulates ACE2 expression in human epidermal keratinocytes. NHEK and N/TERT-2G cells were stimulated with Poly (I:C) and IFN-α + β for 24 hours. ( a ) ACE2 and CTSL mRNA levels relative to control (2 -ΔΔCT ), measured by RT-qPCR in triplicate and normalized to GAPDH. Data represent the mean ± SEM from 3 independent experiments. Data were analyzed using the Wilcoxon signed-rank test, comparing each stimulation condition with the unstimulated control. Only significant P -values ( P < .05) are indicated in the figure. ( b, c ) Western blot analysis of ACE2 (mAb clone AC384) and CTSL (mAb clone 33/1). Shown are representative immunoblots and densitometric quantification, normalized to actin and expressed as mean ± SEM from 3 to 6 independent experiments. Data were analyzed using Wilcoxon signed-rank test, comparing each stimulation condition with the unstimulated control. Only significant P -values ( P < .05) are indicated in the figure. ∗ P < .05. ( d ) Western blot analysis of TMPRSS2 (mAb clone S20014A) expression. Shown is immunoblot representative of 2 independent experiments. ( e ) Representative immunofluorescence microscopy images showing ACE2 expression (mAb clone Poly5036) (red). Cell nuclei are stained with DAPI (white). Bar = 10 μm. Data are representative of at least 2 independent experiments. CTSL, cathepsin L; NHEK, normal human epidermal keratinocyte; Poly (I:C), polyinosinic:polycytidylic acid; TLR3, toll-like receptor 3.

Article Snippet: The following TaqMan MGB-FAM-labelled probes (Applied Biosystems, Thermo Fisher Scientific) were used for qPCR analysis: ACE2 (Hs01085340_m1), ACE2 + dACE2 (Hs01085333_m1), TMPRSS2 (Hs01120965_m1), NRP1 (Hs00826128_m1), CTSL (Hs00964650_m1), BSG (CD147) (Hs00936295_m1), DPP4 (Hs00897386_m1), AXL (Hs01064444_m1), KREMEN1 (Hs00230750_m1), ASGR1 (Hs01005019_m1), and reference gene GAPDH (Hs02758991_g1).

Techniques: Activation Assay, Expressing, Control, Quantitative RT-PCR, Western Blot, Immunofluorescence, Microscopy, Staining

Journal: ACS Omega

Article Title: Enzymatic Glucosylation Enhances the Solubility of Niclosamide but Abrogates Its Therapeutic Efficacy

doi: 10.1021/acsomega.5c12185

Figure Lengend Snippet: Antiviral and cytotoxicity properties of Nic and Nic-Glc against SARS-CoV-2 in (A) Vero E6 cells, (B) A549-hACE2 cells, and (C) Huh7.5 cells. Percent viral infectivity is shown on left y -axis and percent cell viability is shown on the right y -axis (mean ± SD; n = 4). All values are normalized to the nontreated controls. We also confirmed that Remdesivir showed activity as expected and that DMSO alone had no significant antiviral activity ( Figures S3 and S4 , Table S1 ).

Article Snippet: For SARS-CoV-2 experiments in Vero E6 and A549-hACE2 cells (InvivoGen), we used the previously described SARS-CoV-2/human/Denmark/DK-AHH1/2020 isolate (GenBank accession no. MZ049597 ).

Techniques: Infection, Activity Assay