Flow-cytometric determination of high-density-lipoprotein binding sites on human leukocytes

Journal Article · Tue Dec 01 04:00:00 EST 1987 · Clin. Chem. (Winston-Salem, N.C.); (United States)

OSTI ID:5562614

Schmitz, G; Wulf, G; Bruening, T A; Assmann, G

In this method, leukocytes were isolated from 6 mL of EDTA-blood by density-gradient centrifugation and subsequently incubated with rhodamine isothiocyanate (RITC)-conjugated high-density lipoproteins (HDL). The receptor-bound conjugate particles were determined by fluorescent flow cytometry and compared with /sup 125/I-labeled HDL binding data for the same cells. Human granulocytes express the highest number of HDL binding sites (9.4 x 10(4)/cell), followed by monocytes (7.3 x 10(4)/cell) and lymphocytes (4.0 x 10(4)/cell). Compared with conventional analysis of binding of /sup 125/I-labeled HDL in tissue-culture dishes, the present determination revealed significantly lower values for nonspecific binding. In competition studies, the conjugate competes for the same binding sites as /sup 125/I-labeled HDL. With the use of tetranitromethane-treated HDL3, which fails to compete for the HDL receptor sites while nonspecific binding is not affected, we could clearly distinguish between 37 degrees C surface binding and specific 37 degrees C uptake of RITC-HDL3, confirming that the HDL receptor leads bound HDL particles into an intracellular pathway rather than acting as a docking type of receptor. Patients with familial dysbetalipoproteinemia showed a significantly higher number of HDL binding sites in the granulocyte population but normal in lymphocytes and monocytes, indicating increased uptake of cholesterol-containing lipoproteins. In patients with familial hypercholesterolemia, HDL binding was increased in all three cell types, indicating increased cholesterol uptake and increased cholesterol synthesis. The present method allows rapid determination of HDL binding sites in leukocytes from patients with various forms of hyper- and dyslipoproteinemias.

• Research Organization: Medizinischen Einrichtungen der Westfaelischen-Wilhelms-Universitaet, Muenster, Germany, F.R.
• OSTI ID: 5562614
• Journal Information: Clin. Chem. (Winston-Salem, N.C.); (United States), Journal Name: Clin. Chem. (Winston-Salem, N.C.); (United States) Vol. 33:12; ISSN CLCHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ANIMAL CELLS
ANIMALS
BETA DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
BIOLOGICAL MATERIALS
BIOSYNTHESIS
BLOOD
BLOOD CELLS
BODY FLUIDS
CELL FLOW SYSTEMS
CENTRIFUGATION
CHOLESTEROL
CONNECTIVE TISSUE CELLS
DAYS LIVING RADIOISOTOPES
DISEASES
ELECTRON CAPTURE RADIOISOTOPES
HYDROXY COMPOUNDS
INTERMEDIATE MASS NUCLEI
IODINE 125
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LEUKOCYTES
LIPIDS
LIPOPROTEINS
LYMPHOCYTES
MAMMALS
MAN
MATERIALS
MEMBRANE PROTEINS
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
PATIENTS
PRIMATES
PROTEINS
RADIOISOTOPES
REACTION KINETICS
RECEPTORS
SEPARATION PROCESSES
SOMATIC CELLS
STEROIDS
STEROLS
SYNTHESIS
TRACER TECHNIQUES
ULTRACENTRIFUGATION
VASCULAR DISEASES
VERTEBRATES