Anti cgas antibody

Tissues were fixed with 4% paraformaldehyde (Beyotime), embedded in paraffin, sliced into sections, and placed on adhesive microscope slides. Sections were stained with hematoxylin and eosin (H&E) for pathological analysis. Cardiac inflammation was scored on H&E sections as previously described.^{30 (link)} For immunohistochemical (IHC) staining, the anti-cGAS antibody (Cell Signaling Technology) was used.

Immunostaining for protein expression of cGAS or cdsDNA and image acquisition by fluorescence microscopy was performed as described previously [43 (link)]. Briefly, HNSCC cells were seeded on glass coverslips in 6-well plates and treated according to the CBMN assay. At 24 h after exposure, cells were fixed in 3.7% formaldehyde/PBS for 10 min, permeabilized in 5% bovine serum albumin/0.5% Triton-X-100/PBS for 30 min at room temperature, and incubated with an anti-cGAS antibody (Cell Signaling Technology, Danvers, MA, USA). For the immunostaining of cdsDNA with an anti-dsDNA antibody (Abcam, Cambridge, UK), a mild permeabilization protocol was applied using 0.1% Tween-20 and 0.01% Triton-X-100 in PBS for 3 × 5 min at room temperature not affecting the nuclear envelope. Incubation with the primary antibody was performed overnight at 4 °C. Cells were washed in PBS and incubated with a secondary antibody conjugated with Alexa Fluor 488^® or Cy5 (Thermo Fisher Scientific, Dreieich, Germany) for 1 h at room temperature followed by mounting in an HOECHST33258-containing antifade mountant. Fluorescence microscopic image acquisition was performed with an Axio Imager.A1 microscope (Zeiss, Jena, Germany).

Immunostaining for protein expression of cGAS or cdsDNA and image acquisition by fluorescence microscopy was performed as described previously [43 (link)]. Briefly, HNSCC cells were seeded on glass coverslips in 6-well plates and treated according to the CBMN assay. At 24 h after exposure, cells were fixed in 3.7% formaldehyde/PBS for 10 min, permeabilized in 5% bovine serum albumin/0.5% Triton-X-100/PBS for 30 min at room temperature, and incubated with an anti-cGAS antibody (Cell Signaling Technology, Danvers, MA, USA). For the immunostaining of cdsDNA with an anti-dsDNA antibody (Abcam, Cambridge, UK), a mild permeabilization protocol was applied using 0.1% Tween-20 and 0.01% Triton-X-100 in PBS for 3 × 5 min at room temperature not affecting the nuclear envelope. Incubation with the primary antibody was performed overnight at 4 °C. Cells were washed in PBS and incubated with a secondary antibody conjugated with Alexa Fluor 488^® or Cy5 (Thermo Fisher Scientific, Dreieich, Germany) for 1 h at room temperature followed by mounting in an HOECHST33258-containing antifade mountant. Fluorescence microscopic image acquisition was performed with an Axio Imager.A1 microscope (Zeiss, Jena, Germany).