Sections (3–4 µm) were prepared from paraffin blocks and mounted on Thermo Super-Frost poly-L-lysine coated slides (Thermo Fisher Scientific GmbH, Schwerte, Germany). The slides were placed in Tris pH 9.0 retrieval buffer. Antigen retrieval was performed using a microwave oven, without boiling the buffer solution. After cooling of the slides at room temperature, endogenous peroxidase was blocked with 5% hydrogen peroxide. A 5% goat serum was used for blocking of the nonspecific antibody binding. Sections were washed by phosphate buffer solution (pH 7.4) after every step of staining. The following primary antibodies were added: FoxP3 (1:100; Abcam, Bristol, UK), CD56 (ready-to-use; Diagnostic Biosystems, URM) and PGRMC1 (1:150; Abcam) and the slides were incubated in a moist chamber at room temperature. Next, these slides were rinsed in wash buffer and incubated with secondary anti-mouse/rabbit antibodies. The visualization of product reaction was performed using 3,3-diaminobenzidine (DAB) chromogen, followed by Mayer’s hematoxylin counterstaining [23 (link)].
Pgrmc1
Manufactured by Abcam
Sourced in United Kingdom
PGRMC1 is a heme-binding protein that plays a role in the regulation of various cellular processes. It is involved in the modulation of progesterone signaling, cell proliferation, and survival. The core function of PGRMC1 is to serve as a membrane-associated progesterone receptor.
Automatically generated - may contain errors
2 protocols using pgrmc1
1
Immunohistochemical Staining of FoxP3, CD56, and PGRMC1
2
Immunohistochemical Analysis of PCOS Ovaries
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Ten PCOS and ten normal ovarian tissue samples were selected for IHC analysis. Ovarian tissues were embedded in optimal cutting temperature medium for sectioning. The sections (7 μm each) were fixed with 4% paraformaldehyde and permeabilized with 1% Triton X-100. Subsequently, the sections were washed twice with phosphate-buffered saline (PBS) and incubated with antibodies against S phase–related PCNA (1:500; Santa Cruz Biotechnology), PGRMC1 (1:900; Abcam), RBP1 (1:200; Abcam), HSP90 (1:200; Cell Signaling Technology), CALM1, ANXA6, and TPM2 (1:200; Abcam) for 2 h. The sections were washed twice with PBS and incubated with secondary horseradish peroxidase–conjugated antibodies for 1 h. Finally, the sections were washed thrice with PBS and developed using 3,3-diaminobenzidine (DAB). The positively stained cells were visualized under a contrast light microscope. Images were captured using an Eclipse TE 2000-U fluorescence microscope (Nikon, Japan). The protein expression was analyzed quantitatively by cell counting, and the data presented as the mean ratio of the cell counts of positive cells to the nuclei ± standard error (SE).
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