Therefore, we decided to investigate the anatomy of the pelvic organs of a group of human female foetuses, collected at autopsy. Methods We collected at autopsy 36 human female fetuses at different gestational ages, that did not displayed any visible alteration of the pelvic organs. The
characteristics of the fetuses are depicted in Table 1. Pelvic organs were collected en-block, fixed in paraphormaldeyde and included in paraffin. We performed histological analysis of the pelvic organs for each fetus, using Hematoxylin/Eosin and Hematoxylin/Van Gieson staining. For immunohistochemistry 5–7 μm specimen sections embedded in paraffin, were cut, mounted on glass and selleckchem dried overnight at 37°C. All sections were then deparaffinized in xylene, rehydrated through a graded alcohol series and washed in phosphate-buffered Savolitinib datasheet saline (PBS). PBS was used for all subsequent
washes and for antiserum dilution. AZD8931 concentration Tissue sections were quenched sequentially in 3% hydrogen peroxide in aqueous solution and blocked with PBS-6% non-fat dry milk (Biorad, Hercules, CA, U.S.A.) for 1 h at room temperature. Slides were then incubated at 4°C overnight at 1:100 dilution with the following antibodies: the affinity-purified rabbit antibody ERα for the oestrogen receptor (Santa Cruz, Santa Cruz, CA, USA; cat. # sc-542) and the mouse monoclonal antibody M11 for CA125(Dako Laboratories, Carpinteria, CA, USA).
After three washes in PBS to remove the excess of antiserum, the slides were incubated with Alectinib diluted goat anti-rabbit or anti-mouse biotinylated antibodies (Vector Laboratories, Burlingame, CA, U.S.A.) at 1:200 dilution in PBS-3% non-fat dry milk (Biorad) for 1 h. All the slides were then processed by the ABC method (Vector Laboratories) for 30 min at room temperature. Diaminobenzidine (Vector Laboratories) was used as the final chromogen and haematoxylin was used as the nuclear counterstaining. Negative controls for each tissue section were prepared by leaving out the primary antiserum. Positive controls constituted of tumour tissues expressing either the oestrogen receptor or CA125, were run at the same time. All samples were processed under the same conditions.