human uterus adenocarcinoma epithelial cell line hela  (ATCC)

Journal: bioRxiv

Article Title: Converting Lysosomes into Photothermal Organelles Enables Nanoparticle-Free Tumor Ablation via Intracellular Vapor Bubbles

doi: 10.64898/2026.01.23.698471

Figure Lengend Snippet: (a) Schematic of intracellular sunitinib accumulation and lysosomal sequestration. (b) Relative mean fluorescence intensity (rMFI) of cells accumulating sunitinib. (c) Relative increase in Lysotracker™ Deep Red (LDR) staining of HeLa cells as a function of sunitinib concentration and incubation time. Data are presented as mean ± SEM (N=3, n=3). (d) Sunitinib accumulation (5 µM, 24 h) and co-localization with LDR staining. (e) Quantification of LDR fluorescence area per cell (normalized). Data are presented as median ± interquartile range (N=3, n=10).

Article Snippet: Human cervical epithelial adenocarcinoma HeLa cells, lung epithelial A549 cells and T lymphoblast Jurkat E6-1 cells were obtained from American Type Culture Collection (ATCC, Manassas, USA).

Techniques: Fluorescence, Staining, Concentration Assay, Incubation

Journal: bioRxiv

Article Title: Converting Lysosomes into Photothermal Organelles Enables Nanoparticle-Free Tumor Ablation via Intracellular Vapor Bubbles

doi: 10.64898/2026.01.23.698471

Figure Lengend Snippet: (a) Schematic of laser irradiation of cancer cells with sunitinib-loaded lysosomes to produce intracellular VBs, illustrating the VB formation and collapse from sunitinib-loaded lysosomes. (b) Dark-field microscopy images of HeLa cells before and during laser irradiation for visual detection of intracellular VBs, and (c) the corresponding quantification of the intracellular VB formation threshold. (d) Dark-field microscopy images of Jurkat cells before and during laser irradiation and (e) the corresponding intracellular VB formation threshold. The thresholds were determined for HeLa and Jurkat cells treated with 10 µM sunitinib for 24 h. (f) Dark-field microscopy images of HeLa cells incubated with different concentrations of sunitinib (2.5-20 μM) and exposed to pulsed laser irradiation at a fluence of 1.0 J cm -2 . (g) Time-resolved dark-field microscopy images capturing the formation and collapse of VBs in HeLa cells incubated with 20 μM sunitinib following a single nanosecond laser pulse. The sequence illustrates the full dynamic process of intracellular VB generation and subsequent collapse.

Article Snippet: Human cervical epithelial adenocarcinoma HeLa cells, lung epithelial A549 cells and T lymphoblast Jurkat E6-1 cells were obtained from American Type Culture Collection (ATCC, Manassas, USA).

Techniques: Irradiation, Microscopy, Incubation, Sequencing

Journal: bioRxiv

Article Title: Converting Lysosomes into Photothermal Organelles Enables Nanoparticle-Free Tumor Ablation via Intracellular Vapor Bubbles

doi: 10.64898/2026.01.23.698471

Figure Lengend Snippet: (a) Decrease in cell viability after laser treatment (1.8 J cm -2 ) applied on HeLa cells incubated with sunitinib for 24 h at concentrations of 0-10 µM. (b) Cell viability of HeLa cells treated with sunitinib (5 and 10 µM) decreased with increasing laser fluence. (c) Transmission electron microscopy (TEM) images of HeLa cells incubated with 10 µM sunitinib for 24 h for sunitinib only control (no laser, sunitinib CTR), low-fluence irradiation (0.3 J cm -2 ), and VB-generating fluence (0.5 J cm -2 ). Low-magnification images (top) depict overall cellular morphology, while corresponding high-magnification views (bottom) focus on lysosomal structures. Yellow dashed boxes mark cytoplasmic voids observed after VB formation. (d) Quantification of average lysosome diameter in HeLa cells based on TEM analysis after different treatments. (e) Quantification of caspase-3/7 activation in HeLa cells incubated with 10 µM sunitinib for 24 h, followed by laser irradiation at increasing fluences (0.3-1.0 J cm -2 ). Caspase activity was measured using the Caspase-Glo® 3/7 assay and normalized to the non-treated control (NTC) group. (f) Assessment of intracellular reactive oxygen species (ROS) levels in Jurkat cells using the CellROX™ Deep Red assay following nanosecond pulsed laser irradiation. (g) Confocal microscopy images of HeLa cells treated with 10 µM sunitinib after scanning with different laser fluences in a pre-defined line pattern and (h) images of cells treated with different concentrations of sunitinib (5-20 µM) using a fixed laser fluence (1.1 J cm -2 ). (i) Confocal microscopy images of cells scanning in a pre-defined text pattern (10 µM, 1.1 J cm -2 ). Data are presented as mean ± SEM (N=3, n≥3).

Article Snippet: Human cervical epithelial adenocarcinoma HeLa cells, lung epithelial A549 cells and T lymphoblast Jurkat E6-1 cells were obtained from American Type Culture Collection (ATCC, Manassas, USA).

Techniques: Incubation, Transmission Assay, Electron Microscopy, Control, Irradiation, Activation Assay, Activity Assay, Caspase-Glo Assay, Confocal Microscopy

Journal: bioRxiv

Article Title: Converting Lysosomes into Photothermal Organelles Enables Nanoparticle-Free Tumor Ablation via Intracellular Vapor Bubbles

doi: 10.64898/2026.01.23.698471

Figure Lengend Snippet: (a) Diameter and (b) viability of HeLa spheroids incubated with different concentrations of sunitinib. (c) Viability of spheroids before and after laser treatment. Data are presented as mean ± SEM (N≥5, n=3) (d) Optical microscopy images showing VBs at the spheroid’s edges as a function of localized laser irradiation. Yellow dashed circles indicate the irradiated region and highlight the structural disruption of the spheroid observed at the moment of VB collapse. Yellow arrows mark prominent VBs formed during laser exposure. (e) Dark-field microscopy images showing the photoablation effect of spheroids cultured with mounting concentrations of sunitinib. (f) Single time-point analysis of spheroids using bright-field microscopy for morphometric analysis. (g) Single time-point analysis of spheroids by confocal fluorescence microscopy. Fluorescent labeling with Hoechst (blue, indicating nuclei) and TO-PRO-3 (red, indicating dead cells) enabled evaluation of fluorescence intensity distribution within the spheroid area. Confocal fluorescence microscopy was performed using Z-stack imaging to further assess the spatial distribution of both markers following laser exposure, shown as maximum intensity projections. Sunitinib CTR, sunitinib only control.

Article Snippet: Human cervical epithelial adenocarcinoma HeLa cells, lung epithelial A549 cells and T lymphoblast Jurkat E6-1 cells were obtained from American Type Culture Collection (ATCC, Manassas, USA).

Techniques: Incubation, Microscopy, Irradiation, Disruption, Cell Culture, Fluorescence, Labeling, Imaging, Control

Journal: bioRxiv

Article Title: Converting Lysosomes into Photothermal Organelles Enables Nanoparticle-Free Tumor Ablation via Intracellular Vapor Bubbles

doi: 10.64898/2026.01.23.698471

Figure Lengend Snippet: (a) Schematic of pulsed laser irradiation using the LumiPore™ platform of lysosome-stained cells with Lysotracker™ Deep Red (LDR) to produce VBs, illustrating their formation and collapse. (b-e) Dark-field microscopy of HeLa and HT1080 cells before and during laser application and visual detection of VBs and VB threshold quantification for cells treated with 1 µM LDR for 24 h. (f) Confocal fluorescence microscopy of HeLa cells treated with 1 µM LDR before and after laser irradiation at 1.2 J cm -2 , illustrating intracellular morphological deformations following VB generation and collapse. (g) Cell viability of HT1080 cells treated with LDR (1 μM) with increasing laser fluence. (h) Quantification of HT1080 cell viability after incubation with different concentrations of LDR (0.1-1 µM), followed by pulsed laser irradiation at 0.5 J cm -2 and 1.2 J cm -2 . (i) Quantification of caspase-3/7 activation (Caspase-Glo® 3/7 assay) in HT1080 cells incubated with 1 µM LDR for 24 h, followed by laser irradiation at increasing fluences (0.3-1.0 J cm -2 ). Caspase activity was normalized to the non-treated control (NTC) group. Data are presented as mean ± SEM (N=3, n≥3). (j) Confocal fluorescence microscopy analysis of MC38 Gal8-GFP reporter cells incubated with 0.5 μM LDR for 24 h and subsequently irradiated with 532 nm nanosecond pulsed laser at varying fluences: LDR only control (LDR CTR, no laser), 0.3, 0.8, 1.6, and 2.0 J cm -2 . Representative images show Gal8-GFP puncta formation, indicative of lysosomal membrane permeabilization. GFP channel images were converted to grayscale using ImageJ for enhanced contrast. (k) Time-resolved dark-field microscopy images showing sequential frames of HeLa cell spheroids incubated with 1 µM LDR before and after exposure to a 532 nm nanosecond pulsed laser at a fluence of 1.2 J cm -2 . (l) Single time point analysis of spheroids using bright-field microscopy.

Article Snippet: Human cervical epithelial adenocarcinoma HeLa cells, lung epithelial A549 cells and T lymphoblast Jurkat E6-1 cells were obtained from American Type Culture Collection (ATCC, Manassas, USA).

Techniques: Irradiation, Staining, Microscopy, Fluorescence, Incubation, Activation Assay, Caspase-Glo Assay, Activity Assay, Control, Membrane

Journal: Journal of Virology

Article Title: A novel antiviral role of ankyrin repeat and LEM domain-containing 2 (ANKLE2) in restricting vaccinia virus through barrier to autointegration factor (BAF)

doi: 10.1128/jvi.01549-25

Figure Lengend Snippet: Impact of ANKLE2 depletion on BAF phosphorylation and WT or ΔB1 vaccinia virus infection in HeLa cells. ( A ) HeLa cells were transfected with siCTRL or siANKLE2 for 3 days, followed by immunoblotting analysis of ANKLE2. An immunoblot of GAPDH was included as a loading control. ( B ) Cytoplasmic BAF from siANKLE2-transfected HeLa cells was obtained by subcellular fractionation and analyzed by immunoblotting. Closed arrow indicates phosphorylated BAF. The open arrow indicates unphosphorylated BAF. An immunoblot of GAPDH was included as a loading control. ( C ) Relative phospho-BAF protein levels were quantified using ImageLab software (Bio-Rad) from the four independent replicates. The values for siCTRL-transfected cells were normalized to 1. ( D ) DNA accumulation and ( E ) viral yield were measured for WT or ΔB1 vaccinia virus at 24 hpi in siRNA-transfected HeLa cells infected at an MOI of 3. Data are from more than three independent experimental replicates. *, P < 0.05; ***, P < 0.001; ****, P < 0.0001. Relative fold differences are given under each bracket.

Article Snippet: Human cervix epithelial adenocarcinoma HeLa cells were purchased and obtained directly from the ATCC.

Techniques: Phospho-proteomics, Virus, Infection, Transfection, Western Blot, Control, Fractionation, Software

Journal: Journal of Virology

Article Title: A novel antiviral role of ankyrin repeat and LEM domain-containing 2 (ANKLE2) in restricting vaccinia virus through barrier to autointegration factor (BAF)

doi: 10.1128/jvi.01549-25

Figure Lengend Snippet: Characterization of expression and localization of endogenous and mutant ANKLE2 in siRNA-treated transduced HeLa cells. ( A ) Schematic representation of human full-length ANKLE2 and its N-terminal truncation mutants used in this study, showing annotated domains and amino acid positions indicated by black arrows. ( B ) HeLa cells were transduced with lentivirus expressing various siRNA-resistant ANKLE2 truncations, followed by transfection with siCTRL or siANKLE2. Cells were then treated with 1 µg/mL doxycycline for 3 days to induce protein expression and analyzed by immunoblotting. ( C ) Immunofluorescence analysis of HeLa cells transduced with empty vector (CTRL), full-length ANKLE2-3XFLAG, and N-terminal truncation mutants Δ2–53, Δ2–117, and Δ2–252. ANKLE2-transduced HeLa cells were stained with anti-FLAG antibody (red) and DAPI (blue). The representative images shown were taken using an inverted confocal microscope at ×60 magnification. Scale bars represent 25 µm.

Article Snippet: Human cervix epithelial adenocarcinoma HeLa cells were purchased and obtained directly from the ATCC.

Techniques: Expressing, Mutagenesis, Transduction, Transfection, Western Blot, Immunofluorescence, Plasmid Preparation, Staining, Microscopy

Journal: Journal of Virology

Article Title: A novel antiviral role of ankyrin repeat and LEM domain-containing 2 (ANKLE2) in restricting vaccinia virus through barrier to autointegration factor (BAF)

doi: 10.1128/jvi.01549-25

Figure Lengend Snippet: DNA accumulation and viral yield analysis of WT and ΔB1 vaccinia virus in ANKLE2-transduced HeLa cells following endogenous ANKLE2 depletion. DNA accumulation (left panels) and viral yield (right panels) were measured for ( A ) WT and ( B ) ΔB1 vaccinia virus at a low MOI of 0.1 at 48 hpi, as well as for ( C ) ΔB1 vaccinia virus at a higher MOI of 3 with 24 hpi. Data are from more than three independent experimental replicates. Numbers below the significance bars represent the average fold-change value. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Article Snippet: Human cervix epithelial adenocarcinoma HeLa cells were purchased and obtained directly from the ATCC.

Techniques: Virus

Journal: Journal of Virology

Article Title: A novel antiviral role of ankyrin repeat and LEM domain-containing 2 (ANKLE2) in restricting vaccinia virus through barrier to autointegration factor (BAF)

doi: 10.1128/jvi.01549-25

Figure Lengend Snippet: Abundance of viral proteins I3, A11, L4, and F17 during expression of ANKLE2 mutants. ANKLE2-transduced HeLa cells were transfected with siRNAs to deplete endogenous ANKLE2, followed by doxycycline inducement and infection with WT or ΔB1 vaccinia virus at an MOI of 3 for 24 hpi. Whole cell lysates were analyzed by immunoblot using antibodies specific to the proteins on the right. A representative immunoblot is shown with lane numbers shown below each column.

Article Snippet: Human cervix epithelial adenocarcinoma HeLa cells were purchased and obtained directly from the ATCC.

Techniques: Expressing, Transfection, Infection, Virus, Western Blot

Journal: Journal of Virology

Article Title: A novel antiviral role of ankyrin repeat and LEM domain-containing 2 (ANKLE2) in restricting vaccinia virus through barrier to autointegration factor (BAF)

doi: 10.1128/jvi.01549-25

Figure Lengend Snippet: Immunofluorescence analysis of doxycycline-treated ANKLE2-transduced HeLa cells infected with ( A ) WT or ( B ) ΔB1 vaccinia virus at an MOI of 5 for 6 hours at 37°C prior to fixation. Viral DNA replication factories were detected by I3 staining (green) using anti-I3 antibody, whereas transduced ANKLE2 was detected using anti-FLAG primary antibody (red). The representative images shown were taken using an inverted confocal microscope at ×60 magnification. Scale bars represent 25 µm.

Article Snippet: Human cervix epithelial adenocarcinoma HeLa cells were purchased and obtained directly from the ATCC.

Techniques: Immunofluorescence, Infection, Virus, Staining, Microscopy

Journal: Journal of Virology

Article Title: A novel antiviral role of ankyrin repeat and LEM domain-containing 2 (ANKLE2) in restricting vaccinia virus through barrier to autointegration factor (BAF)

doi: 10.1128/jvi.01549-25

Figure Lengend Snippet: Protein immunoblotting of cytoplasmic BAF in doxycycline-treated ANKLE2-transduced HeLa cells following subcellular fractionation. Cytoplasmic and membrane fractions were analyzed by immunoblot using antibodies specific to the proteins on the right. When using an antibody recognizing total BAF, the closed arrow indicates the migration position of phosphorylated BAF, and the open arrow indicates the unphosphorylated BAF. A representative immunoblot is shown with lane numbers shown below each column.

Article Snippet: Human cervix epithelial adenocarcinoma HeLa cells were purchased and obtained directly from the ATCC.

Techniques: Western Blot, Fractionation, Membrane, Migration

Journal: Journal of Virology

Article Title: A novel antiviral role of ankyrin repeat and LEM domain-containing 2 (ANKLE2) in restricting vaccinia virus through barrier to autointegration factor (BAF)

doi: 10.1128/jvi.01549-25

Figure Lengend Snippet: Immunoblotting of FLAG-tag immunoprecipitated lysates from doxycycline-treated ANKLE2-transduced HeLa cells. Confluent monolayers of cells in 10 cm 2 dishes were harvested, pelleted, and lysed with a lysis buffer. Complexes of transduced ANKLE2 containing 3XFLAG tag were immunoprecipitated overnight at 4°C and eluted by incubation at 37°C for 24 h. Samples were then analyzed by immunoblotting using antibodies against ANKLE2, FLAG, PP2A, and total BAF proteins. INPUT lysates were included to confirm the similar levels of ANKLE2, PP2A, and BAF proteins in each lysate. A representative immunoblot is shown with lane numbers shown below each column.

Article Snippet: Human cervix epithelial adenocarcinoma HeLa cells were purchased and obtained directly from the ATCC.

Techniques: Western Blot, FLAG-tag, Immunoprecipitation, Lysis, Incubation