Methods for ligand-receptor assays in clinical chemistry.

Abstract

This is intended to be a practical review for the clinical chemist of the laboratory procedures most commonly used to quantitate hormone receptors in various cellular fractions. These procedures include use of charcoal adsorption and hydroxylapatite for intracellular receptors and of centrifugation and filtration for membrane receptors. We discuss the use of the Scatchard analysis to establish the steroid-receptor affinity and the quantity of steroid-receptor binding sites. Both pre- and post-labeled sucrose density gradient methods are outlined. One section is devoted to the direct and indirect methods used in nuclear “exchange” assays. Basic theory underlying each assay is presented, but, more importantly, we assess the advantages and disadvantages of each procedure. On the basis of this information, one may decide which assay is best suited for a particular laboratory and (or) specimen.

Development of a LytE‐based high‐density surface display system in Bacillus subtilis

“…Wall‐bound protein complexes were separated from the unbound proteins by centrifugation. Data were analysed using Scatchard plots (Smith and Sestili, 1980) to gain information concerning the binding affinity and the number of cell wall binding sites (B max ) recognized by CWBM LytE . Interestingly, CWBM LytE and its fusions consistently recognized two classes of binding sites in cell wall (Fig.…”

Bio‐nanocapsule‐based scaffold improves the sensitivity and ligand‐binding capacity of mammalian receptors on the sensor chip

“…This set‐up enables the detection of ligand‐receptor interactions in a physiological context but can also lead to high background signals by non‐specific binding of the ligand to the cell surface or by autonomous receptor internalization. Alternatively, membrane fractions were isolated from the cells expressing receptors endogenously or exogenously , and basically showed higher binding affinities by eliminating the cell‐derived background, but sometimes receptor denaturation during membrane extraction could lead to reduced binding affinities . Consequently, receptors are expressed in large amounts by in vitro translation systems .…”