Histomorphological Demonstration of Stromal Components in Palpable Female Breast Tumors, Counterstained with Annatto Seed Crude Extract
Charles Confidence Nkesichi *
Department of Histopathology/Cytology, Faculty of Medical Laboratory Science, Rivers State University, Nigeria.
Gideon Ihebuzo N. Ndubuka
Federal University of Technology Owerri, Nigeria.
Yibala Ibor Oboma
Niger Delta University, Wilberforce Island Bayelsa State, Nigeria.
Oraedu Ifeanyichukwu Felix
Rivers State University, Rivers State, Nigeria.
*Author to whom correspondence should be addressed.
Abstract
Background: Annatto seed, obtained from Bixa orellana, contains natural yellow-orange pigments that may be useful as biological staining agents.
Aims: This study evaluated the histomorphological demonstration of stromal components in palpable female breast tumours using crude annatto seed extracts as counterstains to haematoxylin.
Methods: A total of 100 female breast tissue blocks were examined, comprising fibroadenoma (40 blocks), fibrocystic change (30 blocks), and invasive ductal carcinoma (30 blocks). Dried annatto seeds were used for crude dye extraction. One hundred grams of seed material was extracted in 200 ml of each solvent, including 95% ethanol, 70% ethanol, distilled water, and acetone. Phytochemical assessment of the extract was performed. Tissue sections were stained using routine haematoxylin and eosin as the control method, while the annatto seed extracts were used as counterstains to Harris haematoxylin. Stained sections were evaluated photomicrographically for staining intensity, visibility of stromal components, and preservation of tissue architecture.
Results: The extracts produced varying colour intensity and staining performance. Acetone produced the highest colour yield and volume, followed by 95% ethanol, 70% ethanol, and distilled water. However, sections stained with acetone showed a yellowish background that reduced cellular clarity. Sections counterstained with 95% ethanolic extract showed clearer stromal visualisation and better preservation of tissue morphology across fibroadenoma, fibrocystic change, and invasive ductal carcinoma when compared with the other crude extracts. The appearance was closer to that observed in routine haematoxylin and eosin-stained control sections.
Conclusion: Crude annatto seed extract, particularly the 95% ethanolic preparation, demonstrated potential as a counterstain for stromal components in selected palpable female breast tumours. Further optimisation and quantitative validation are required before routine diagnostic use can be recommended.
Keywords: Annatto seed extract, Bixa orellana, breast tumours, stromal components, counterstaining, haematoxylin and eosin, fibroadenoma, fibrocystic change, invasive ductal carcinoma, histomorphology