Peripheral type benzodiazepine receptor in T lymphocyte rich preparation

Abstract

Some types of mood disorders and drugs are suggested to affect peripheral type benzodiazepine receptors (PBR), but their mechanisms are unclear. The isolation of pure lymphocytes is requisite for the investigation of the function of PBR on lymphocytes, since platelets and monocytes also have many PBR. The objective of this study was to establish a method of binding assay for PBR using pure T lymphocytes. Mononuclear cells and T lymphocytes were prepared by using a density gradient material and magnetic beads, respectively. The cells were analyzed using flow cytometry and a counting chamber. Binding studies were performed using T lymphocytes from 10 normal volunteers. The T lymphocytes were incubated with [ 3H]PK11195, harvested on glass fiber filters, and counted with a plate scintillation counter. The binding data were analyzed by the Scatchard method. With the magnetic bead technique, pure T cells were selected that contained only 1.5% monocytes and platelet cell ratio of 1.4. The Scatchard plot of the data indicated that only one type of specific binding site was involved in the binding. The dissociation constant (Kd) was 3.8 ± 1.3 nM (mean ± SD), and the Bmax was 379 ± 124 fmol 10 6 cells (mean ± SD). The density gradient- magnetic beads technique can be used as an appropriate method of preparation of T cells for PBR binding assay.