Dako real envision hrp rabbit mouse secondary antibody

Ovary tissues were fixed by 10% neutral buffered formalin (BBC Chemical, Mount Vernon, WA, USA), embedded in paraffin, and sectioned with 6 μm of the ovaries. Sectioned ovary tissues were deparaffinized in a 60 °C dry oven by xylene and ethanol. Deparaffinized tissues underwent antigen retrieval by an EDTA (eLbio, Seongnam-Si, Korea) reaction. After removing the unbound primary antibody, the tissues in slides were incubated with DAKO Real EnVision HRP Rabbit/Mouse secondary antibody (Dako) at room temperature for 1 h. The slides were incubated with DAB and counterstained with hematoxylin (Dako). After the reaction, slides were rinsed with tap water. Slides underwent dehydration by ethanol and xylene. Tissues were analyzed by the 3D HISTECH program (The Digital Pathology Company, Budapest, Hungary).

Sectioned ovarian tissues were deparaffinized in a 60 °C dry oven and by xylene and ethanol. Deparaffinized tissues were subjected to antigen retrieval by EDTA (eLbio, Seongnam-si, Republic of Korea) reaction and slowly cooled with water. The ovarian tissues were washed with distilled water (D. W) and treated with peroxide blocking solution containing 3% H₂O₂ in methanol for 10 min. Next, the ovarian tissues were washed with D.W and treated with primary antibodies with diluent buffer (Dako) at 4 °C overnight. The rabbit anti-PDGF receptor β antibody (28E1; 3169S, Cell Signaling Technology) was diluted 1:250. After removal of the unbound primary antibody, the tissues in slides were incubated with Dako Real EnVision HRP Rabbit/Mouse secondary antibody (Dako, California, USA) at room temperature for 1 h. The slides were incubated with DAB and counterstained with hematoxylin (Dako). After reaction, slides were rinsed by tap water. Slides were dehydrated by ethanol and xylene. Tissues were analyzed by the 3D HISTECT program (The Digital Pathology Company).