Immunohistochemistry kit

Immunohistochemistry with a monoclonal p62 antibody was performed on formalin-fixed paraffin-embedded TMA sections mounted on poly L-lysine-coated slides. The sections were deparaffinized in xylene and rehydrated through graded alcohols into distilled water. Antigen retrieval was in 0.1 M citrate buffer using a pressure cooker at 95°C for 15 min. Immunostaining was performed by using an immunohistochemistry kit according to the manufacturer’s instructions (Invitrogen) after incubation with primary antibody to p62 (Santa Cruz Biotechnologies) for 1 h. Non-immune pooled mouse immunoglobulin was used as a negative control. Sections were incubated in secondary antibody for 1 h followed by colorimetric detection using chromogen in accordance with the manufacturers’ protocols (Dako). Slides were then counterstained with haematoxylin, rinsed, and dehydrated through graded alcohols into non-aqueous solution and cover-slipped with mounting media.

TRIzol lysate was purchased from the Invitrogen Company (Frederick, USA), The revertAid first stand cDNA synthesis kit was purchased from the Thermo Company (Lithuania, EU), the Fluorescence quantitative RT-PCR kit was purchased from the Takara Company (Dalian, China), the HO-1 primers were synthesized by the Invitrogen Company (Shanghai, China), the anti-heme oxygenase 1 antibody was purchased from the Abcam Company (Cambridge, MA, USA), the immunohistochemistry kit was purchased from the Invitrogen Company (Frederick, USA).