Use of the fluorescence activated cell sorter for studying uptake of fluorescent fatty acids into cultured cells.
A procedure is described for the continuous monitoring and recording of fatty acid transport into cultured cells. Uptake of the fluorescent fatty acid derivative, 12-(1-pyrene)dodecanoic acid [P12] by a suspension of human promyelocytic leukemia (HL-60) cells was analyzed and recorded by the Fluorescence Activated Cell Sorter (FACS). A major advantage of the FACS is its ability for measuring the fluorescence of the cells only, without interference by that of the fatty acids dissolved or dispersed in the medium or complexed with serum albumin. The time-dependent fluorescence increase due to uptake of the acid by the cells could thus be monitored and recorded directly without a need for sedimenting, washing and extracting the cellular lipids. This procedure provided an accurate measurement of the kinetics of uptake of the fatty acid into the cells and permitted study of the effect of various parameters such as temperature, glucose, albumin or serum. The results indicated a biphasic uptake of P12 into the cells. The first, a rapid, energy-independent phase, lasted 3-4 min. This was followed by a slower uptake which was directly related to the metabolic utilization of the acid in the cells. This second phase represents the overall process of translocation across the cell membrane, activation to acyl coenzyme A and incorporation into the cellular neutral lipids and phosphoglycerides. When compared to HL-60, murine erythroleukemia (MEL) cells took up considerably lesser quantities of P12. This was utilized for setting up a model system, in which mixtures of these two respective cell types could be identified and separated from each other on the basis of their relative rates of uptake of the fluorescent fatty acid.
Nahas
N
N
Fibach
E
E
Giloh
H
H
Gatt
S
S
D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D007700:Kinetics; D007850:Lauric Acids / Q000378:metabolism*; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism; D007942:Leukemia, Experimental / Q000378:metabolism; D015470:Leukemia, Myeloid, Acute; D051379:Mice
D007850:Lauric Acids; C040596:12-(1-pyrene)dodecanoic acid