A_2AR activity affected UUO (unilateral ureteral obstruction)-induced deposition of collagen III

(A) Representative immunohistochemistry of Collagen III) (Col III) from the A_2AR KO and WT mice, at day 3, 7 and 14 post-UUO or Sham, following treatment of CGS21680 (CGS) or vehicle (Veh). Scale bar  = 50 µm, 400×. 

Collagen III in rat kidney tissue is detected ab7778 at 1/400 dilution. Pressure cooker mediated antigen retrieval was performed.

(From Figure 3 of Xiao et al)

ab7778 staining Collagen III in human testicle tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections).
Tissue was fixed with paraformaldehyde and a heat mediated antigen retrieval step was performed using TE buffer pH 9.0. Samples were then incubated with ab7778 at a 1/200 dilution for 30 minutes at 20°C. The secondary used was an undiluted, HRP-conjugated goat anti-rabbit polyclonal.

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Anti-Collagen III antibody (ab7778) at 1/1000 dilution + Human collagen III at 0.1 µg

Predicted band size: 138 kDa



Western Blot produced under denaturing and reducing conditions.

ab7778 at 1:400 (45 min RT) showed strong staining in FFPE sections of human skin(left, dermis) with moderate to strong red staining and testis (right) where strong staining was observed within connective tissue between seminiferous tubules. The antibody showed strong extracellular staining within connective tissues across many organs with minimal background staining. Slides were steamed in 0.01 M sodium citrate buffer, pH 6.0 (ab64214) at 99-100°C - 20 minutes for antigen retrieval.

Immunohistochemistry of ab7778. Tissue: right lobe of the liver section. A:Central Vein (CV) fibrosis, B: Non-fibrotic CV, C: Perisinusodial fibrosis, D: Non-fibrotic area, E: Protat tract fibrosis, F: Septal fibrosis (arrow). Fixation: formalin fixed paraffin embedded. Antigen retrieval: not required. Primary antibody: Anti-collagen type I at 1:500 for 4°C for 24hr. Secondary antibody: Peroxidase biotin-streptavidin rabbit secondary antibody at 1:10,000 for 45 min at RT. Localization: Anti-collagen type III is intra and extracellular. Staining: 3.3’-diaminobenzidine tetrahydrochloride was used as the chromogen. Nuclei were counterstained purple with hematoxylin.

Ab53088 staining Human skin (ab30166). Staining is localised to the extracellular matrix.
Left panel: with primary antibody at 1 ug/ml. Right panel: isotype control.
Sections were stained using an automated system DAKO Autostainer Plus , at room temperature. Sections were rehydrated and antigen retrieved with the DAKO 3-in-1 antigen retrieval buffer citrate pH 6.0 (ab64214) in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 minutes and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.