Immuno-histochemical localization of cholesterol binding proteins in Schistocerca gregaria (Forskal)

Abstract

This manuscript aims to investigate immunocytochemical localization of cholesterol binding proteins (CBPs) in semi-thin sections of midgut of Schistocerca gregaria (Forskal). For this purpose, polyclonal antibody specific to CBPs were raised in albino mice and used in immuno-fluorescence and immuno-blotting to determine the cellular location of CBPs. Midgut tissue sections were incubated with pAbs anti-cholesterol binding protein (primary antibody) and finally associate them with HRP conjugated anti-rabbit immunoglobulin (secondary antibody). Semi thin tissue sections of midgut portion were stained with hematoxylin and eosin for establishing general morphology of epithelial cells in control sections. Positive control tissue sections were stained with Sudan Black-B for microscopic visualization of cholesterol binding sites. Further, cholesterol association in tissue sections was confirmed by using tetramethylrhodamine isothiocyanate (TRITC) labeled florescent antibodies and immuno-blotting of CBPs. Finally, CBPs or cholesterol-carrying proteins were detected intracellularly in midgut epithelial/ microvillus cells named as CBP+. Zymogene like dense granules localized were found scattered throughout the apical portion of microvillus cells. Further, presence of these CBPS was confirmed by SDS-PAGE gel electrophoresis and immuno-blotting. In treatments, dietary cholesterol was found to be internalized bound to complexed with CBPs before absorption. Further, same protein was also localized in other tissues like fat body, testis, and ovary of male and female insects of S. gregaria. However, present study done on immuno-cytochemical localization of cholesterol binding proteins in microvillus cells confirms that CBPS are main careers of cholesterol. These proteins also assist in transport of cholesterol to the various organs after its subsequent absorption.

Keywords: Cholesterol binding proteins (CBPs), cholesterol, lipoprotein, midgut, immunocytochemical localization, S. gregaria (Forskal).

African Journal of Biotechnology Vol 13(28) 2884-2896