Decrease of intracellular fluorescein fluorescence polarization (IFFP) in human peripheral blood lymphocytes undergoing stimulation with phytohaemagglutinin (PHA), concanavalin A (ConA), pokeweed mitogen (PWM) and anti‐CD3 antibody

Abstract

In the present study we describe the induction of changes in intracellular fluorescein fluorescence polarization (IFFP) in lymphocytes undergoing activation with a variety of stimulants. These stimulants included the lectins phytohaemagglutinin (PHA), concanavalin (ConA), pokeweed mitogen (PWM) and anti-CD3 antibody. Changes in IFFP were detected in individual cells using the Cellscan apparatus. Our results show that by employing mitogenic concentrations of PHA, as revealed in a [3H]-thymidine incorporation assay, a decrease in the IFFP in human peripheral blood lymphocytes (PBL) occurred within 40 min. ConA and anti-CD3 affected similarly IFFP, whereas PWM, a B lymphocyte lectin, had no effect on IFFP at the concentrations employed. Kinetic analysis revealed that changes in IFFP occurred within 20-40 min after exposure to the stimulants and lasted for 24 h. Our results show that stimulants which activate CD3+ lymphocytes caused immediate changes in IFFP, in an enriched population of human PBL. The possible mechanisms involved in IFFP modulation following exposure to selected stimulants are discussed.

A Novel System to Diagnose Cutaneous Adverse Drug Reactions Employing the Cellscan—Comparison with Histamine Releasing Test and Inf‐γ Releasing Test

“…The CellScan is unique in its capability of repetitive measurements of individual cells in a cell population. It has been used to detect activation of lymphocytes both in physiological (Eisenthal et al 1996, Zurgil et al 1996 and pathological situations including: cancer (Ron et al 1995, Merimsky et al 1996, Rahmani et al 1996, Schiffenbauer et al 2002, Cohen et al 2003, autoimmune disorders (Zurgil et al 1997, and atherosclerosis ).…”

Monitoring of Intracellular Enzyme Kinetic Characteristics of Peripheral Mononuclear Cells in Breast Cancer Patients

“…The estimation of the kinetics of this process was conducted via the Michaelis-Menten constants. Our preliminary results have shown that early activation events, which occur upon cell incubation with and without either phytohemagglutinin or autologous tumor tissue, may be demonstrated by the general biochemical kinetic parameters (the Michaelis-Menten constant, K m , and the maximal velocity, V max ) that display the differences in intracellular processes in the lymphocytes, related to the enzymatic activity as well as to the cell size, the cytoplasm membrane permeability, and so forth (15,16,19).…”

Indication that Intracellular Fluorescence Polarization of T Lymphocytes is Cell Cycle Dependent.

“…Other environmental factors such as ion concentrations Ca^, K+and the concentration of the lectin, were shownto have markedinfluence on the fluorescence polarization and intensity (22). IFFP measurements of human peripheral blood lymphocytes have been shownto be a sensitive means of detecting early lymphocyte stimulation (8,9,13). High correlations were found between cell cycle parameters and mean IFFP values of the Jurkat T cell line.…”

“…Early activation responses of human peripheral lymphocytes as induced by mitogenic or antigenic stimulation can be detected by a characteristic decrease in IFFP (3,8,9,13,21). Changes in IFFP were also evident in fibroblasts following cytokine exposure (15).…”