Journal: bioRxiv

Article Title: Stiffness and Viscoelasticity of Adipose Tissue Decellularized Extracellular Matrix Hydrogels Influence Proliferation, Growth Pattern, Migration and Invasion of Breast Cancer Cells

doi: 10.64898/2025.12.11.693682

Figure Lengend Snippet: A) Confocal microscope images of MDA-MB-231 cells stained with DAPI (blue) and phalloidin (green) growing within 1% and 3% adipose tissue dECM-derived hydrogels after 8 and 15 days in culture. Experiments were repeated six times per condition. Scale bar: 50 μm. B) Histograms showing the growth angles of connected cells in 1% (blue) and 3% (red) adipose tissue dECM-derived hydrogels after 15 days in culture, quantified using Dragonfly software.

Article Snippet: For DAPI/Phalloidin staining, cells were fixed with 4% paraformaldehyde in PBS, permeabilized with 0.1% Triton X-100 in PBS and stained with a solution containing DAPI (4’,6-diamidino-2-phenylindole) (MBD0015, Merck) and Phalloidin conjugate (Alexa Fluor 488, #8878, Cell Signaling Technologies) at 1:1000 and 1:200, respectively, for 30 minutes in a rocker at room temperature to facilitate dye diffusion and covered with aluminum foil.

Techniques: Microscopy, Staining, Derivative Assay, Software

Journal: The Journal of Experimental Medicine

Article Title: The cell cycle restricts activation-induced cytidine deaminase activity to early G1

doi: 10.1084/jem.20161649

Figure Lengend Snippet: Uracil-qPCR detects uracil accumulation at the antibody 5′-Sμ region. (A) Diagram of how Pfu polymerase and its mutant Pfu-Cx act on DNA. Uracil-free DNA can be amplified by both Pfu and Pfu-Cx polymerases (left). The lack of uracil recognition by Pfu-Cx polymerase allows it to replicate uracil-containing DNA, whereas Pfu polymerase stalls (right). This differential ability to amplify uracil-containing DNA is harnessed to measure the relative level of uracil (see Materials and methods; ). (B) Experimental layout. In vitro–activated B lymphocytes were arrested with nocodazole 42 h after stimulation. 6 h later, nocodazole was removed and B cells were collected for analysis at different time points. (C) DAPI staining for DNA content at the indicated time points upon nocodazole release. Red, the sorted populations for experiments in and Fig. S2 (E and F). Upon release, B cells were still mainly in M phase at 0.5 h, whereas the released G1 population increased to ∼40% at 1.5 h (early G1) and 60% at 4 h (middle G1). At 6 h, some cells entered S phase, and 30% were still in G1 (late G1). (D) Relative uracil levels at the antibody 5′-Sμ region, as determined by Uracil-qPCR (see Materials and methods). AID-proficient and -deficient cells are compared. Error bars, SEM; *, P < 0.01, two-tailed Student t test. Three independent experiments are shown. (E) Diagram of the sorting strategy for the analysis of G1, S, G2, and M cells. In vitro–activated B cells at different cell cycle stages were sorted according to phospho-Histone H3 (Ser10) and DNA content (DAPI). (F) The relative uracil levels at 5′-Sμ at different cell cycle stages were determined by Uracil-qPCR in WT and UNG-deficient B cells. One experiment is shown.

Article Snippet: For sorting cells at different cell cycle stages , in vitro–activated B cells (48 h after isolation) were stained with Alexa Fluor 488–conjugated anti–phospho-Histone H3, Ser10 antibody (Cell Signaling Technology), and DAPI (0.5 µg/ml), before sorting with FACSAria (BD).

Techniques: Mutagenesis, Amplification, In Vitro, Staining, Two Tailed Test