dapi  (Bio-Rad)

Journal: bioRxiv

Article Title: HIF-1α and RhoA Drive Enhanced Motility and Aerotaxis of Polyaneuploid Prostate Cancer Cells in Hypoxia

doi: 10.1101/2025.09.03.673887

Figure Lengend Snippet: (A-C) Representative image of PC3-GFP cells under normoxic conditions. (D-F) Representative image of PC3-GFP cells after 16 hours of hypoxia, with PACC cell (outlined) surrounded by non-PACC cells. The enlarged nucleus of the PACC cell is indicated by an arrow in (E). (G, H) Fluorescence-Activated Cell Sorting (FACS) analysis of PC3-GFP cells in normoxia (G) and hypoxia (H). PACCs were defined as cells with DNA content >4N. FSC-H: forward scatter height (proxy of cell size); UV- 450-A, DAPI fluorescence intensity (proxy for genomic content). (I) Quantification of PACCs from FACS analysis in normoxic and hypoxic conditions. Data are shown as mean ± SD. Significance determined in (I) as p ≤ 0.01 ( ** ) via Unpaired Student’s t test (see Material and Methods). Scale bars for A-F are in A and represent 20 μm.

Article Snippet: Nuclear staining was performed using a 1X solution of PureBlue DAPI Nuclear Staining Dye (Bio-Rad, #1351303) for 15 minutes, followed by three final washes (5 minutes each).

Techniques: Fluorescence, FACS

Journal: bioRxiv

Article Title: HIF-1α and RhoA Drive Enhanced Motility and Aerotaxis of Polyaneuploid Prostate Cancer Cells in Hypoxia

doi: 10.1101/2025.09.03.673887

Figure Lengend Snippet: (A-J) Immunofluorescence microscopy of PC3 cells under normoxia (A-E) and hypoxia for 16 hours (F-J) , visualizing GFP (A, F) , DAPI (B, G) , HIF-1α ( C, H) , and RhoA (D, I). Merged images (E, J) show GFP, DAPI, HIF-1α, and RhoA. PACCs under hypoxia (outlined in F-J) were identified by nuclear area ≥2.5x that of the normoxic parental population, as computed in Figure S2. (K , L) Quantification of HIF-1α intensity by Integrated Density (K) and Mean Fluorescence Intensity (MFI) (L) of PACC, Non-PACC, and Normoxic cells. (M, N) RhoA expression quantified by Integrated Density (M) and MFI (N) of PACC, Non-PACC, and Normoxic cells. Data represent mean ± SEM. Significance determined as p < 0.01 ( ** ), and p < 0.001 ( *** ) via One-way ANOVA with Tukey-Kramer post hoc test (see Materials and Methods). Scale bars for A-J are in A and represent 20 μm.

Article Snippet: Nuclear staining was performed using a 1X solution of PureBlue DAPI Nuclear Staining Dye (Bio-Rad, #1351303) for 15 minutes, followed by three final washes (5 minutes each).

Techniques: Immunofluorescence, Microscopy, Fluorescence, Expressing

Journal: bioRxiv

Article Title: HIF-1α and RhoA Drive Enhanced Motility and Aerotaxis of Polyaneuploid Prostate Cancer Cells in Hypoxia

doi: 10.1101/2025.09.03.673887

Figure Lengend Snippet: (A-O) Immunofluorescence microscopy of PC3-GFP cells either mock transfected (A-E) , transfected with non-targeting control siRNA (siNTC, F-J ), or transfected with HIF-1α siRNA ( siHIF-1α, K-O) and exposed to hypoxia for 16 hours. Panels show GFP (A, F, K) , DAPI (B, G, L) , HIF-1α (C, H, M) , and RhoA (D, I, N ). Merged images of all channels per condition are displayed in (E, J, O). PACCs (outlined in A-O ) were identified by nuclear area ≥2.5x that of the normoxic parental population (see Fig. S2). (P–Q) Quantification of RhoA expression by MFI (P) and Integrated Density (Q) in PACC and non-PACC cells transfected with non-targeting control siRNA (siNTC), HIF-1α siRNA (siHIF), or mock transfected (Mock). Data represent mean ± SEM. Significance determined as p < 0.05 (*), p < 0.001 ( *** ), p <0.0001 ( **** ), and ns = not significant via One-way ANOVA with Tukey-Kramer post hoc test (see Materials and Methods). Scale bar in A represents 20 μm and applies to A-O .

Article Snippet: Nuclear staining was performed using a 1X solution of PureBlue DAPI Nuclear Staining Dye (Bio-Rad, #1351303) for 15 minutes, followed by three final washes (5 minutes each).

Techniques: Immunofluorescence, Microscopy, Transfection, Control, Expressing

Journal: bioRxiv

Article Title: HIF-1α and RhoA Drive Enhanced Motility and Aerotaxis of Polyaneuploid Prostate Cancer Cells in Hypoxia

doi: 10.1101/2025.09.03.673887

Figure Lengend Snippet: (A-O) Immunofluorescence microscopy of PC3-GFP cells either mock transfected (A-E) , transfected with non-targeting control siRNA ( siNTC, F-J) , or transfected with HIF-1α siRNA ( siHIF-1α, K-O) under normoxic conditions. Panels show GFP (A, F, K) , DAPI (B, G, L) , HIF-1α (C, H, M) , and RhoA (D, I, N ). Merged images of all channels per condition are displayed in (E, J, O). (P–Q) Quantification of RhoA expression by MFI (P) and Integrated Density (Q) in normoxic cells transfected with non-targeting control siRNA (siNTC), HIF-1α siRNA (siHIF), or mock transfected (Mock). Data represent mean ± SEM. Significance determined as ns = not significant via One-way ANOVA with Tukey-Kramer post hoc test (see Materials and Methods). Scale bar in A represents 20 μm and applies to A-O .

Article Snippet: Nuclear staining was performed using a 1X solution of PureBlue DAPI Nuclear Staining Dye (Bio-Rad, #1351303) for 15 minutes, followed by three final washes (5 minutes each).

Techniques: Immunofluorescence, Microscopy, Transfection, Control, Expressing