Open AccessStandardization of a flow cytometric method for measurement of low‐density lipoprotein receptor activity on blood mononuclear cells
Author(s) -
Løhne Kari,
Urdal Petter,
Leren Trond P.,
Tonstad Serena,
Ose Leiv
Publication year - 1995
Publication title -
cytometry
Language(s) - English
Resource type - Journals
eISSN - 1097-0320
pISSN - 0196-4763
DOI - 10.1002/cyto.990200404
Subject(s) - peripheral blood mononuclear cell , flow cytometry , receptor , coefficient of variation , ldl receptor , lipoprotein , biology , in vivo , immunology , endocrinology , medicine , microbiology and biotechnology , in vitro , cholesterol , chemistry , biochemistry , genetics , chromatography
Flow cytometric methods for measurement of low‐density lipoprotein (LDL) receptor activity on peripheral blood mononuclear cells (PBMC) may be used to identify patients with familial hypercho‐lesterolemia (FH). However, cellular LDL receptor activities measured in FH heterozygotes may over‐lap with those of healthy subjects. Analytical variation is probably responsible for some of this over‐lap. We have examined several technical details that may affect analytical variation. In each analysis, we included one standard and two control cell preparations. These were cells isolated from three donors and stored in aliquots at –135°C. Use of standard cells reduced between‐series analytical variation of the controls by approximately 50%. Preincubation‐conditions used to induce the maximum number of receptors, the concentration of fluorochrome 1,1′‐dioctadecyl‐3,3,3′,3′‐tetramethylindocarbocyanine‐perchlorate (DiI)‐LDL, labelling time, and conditions during storage of labelled cells before flow cytometry were also examined in order to reduce analytical variation. Having standardized the assay, we found among 20 healthy subjects a median receptor activity of 100% vs. 51% among 26 patients who fulfilled clinical criteria for FH. However, four of the patients showed distinctly normal receptor activities, which may suggest either the presence of some other biochemical defect or that in vivo dysfunctional receptors may be measured as normal in some patients with our assay. © 1995 Wiley‐Liss, Inc.