High-density lipoprotein subfractions measured in stored serum

1994 ◽  
Vol 40 (9) ◽  
pp. 1713-1716 ◽  
Author(s):  
L L Bausserman ◽  
A L Saritelli ◽  
D Milosavljevic
Keyword(s):  
High Density Lipoprotein ◽  
Dextran Sulfate ◽  
Density Lipoprotein ◽  
Hdl Cholesterol ◽  
High Density ◽  
Manganese Chloride ◽  
Lipoprotein Subfractions ◽  
Hdl Subfractions ◽  
High Density Lipoprotein Subfractions

Abstract We compared the effects of freezing serum on the determination of high-density lipoprotein (HDL) subfractions by two dual-precipitation methods, heparin and manganese chloride/dextran sulfate (HM/DS) (Gidez et al., J Lipid Res 1982;23:1206-23) and DS/DS (Warnick et al., Clin Chem 1982;28:1574), and by ultracentrifugation. Storing serum for 1 month at -70 degrees C resulted in reduced HDL3-cholesterol by ultracentrifugation and reduced total and HDL3-cholesterol by the DS/DS method. There was no change in either total HDL-cholesterol or HDL3-cholesterol with the HM/DS method. Additional studies involving only HM/DS indicated that total HDL-cholesterol in serum stored at 4 degrees C begins to decline after 3 days (-3.1 +/- 3.5%, P < 0.1). HDL was stable at -20 degrees C for 2 weeks but both total and HDL3-cholesterol decreased significantly after 1 month. Storage of serum at -70 degrees C resulted in no changes for 1 year; however, at 18 months, HDL3-cholesterol was reduced 13% (P = 0.002). We conclude that HDL subfractions can be determined accurately in serum as well as in plasma after storage at -70 degrees C for up to 1 year.

Cholesterol distribution between high-density-lipoprotein subfractions HDL2 and HDL3 determined in serum by discontinuous gradient gel electrophoresis

1991 ◽  
Vol 37 (7) ◽  
pp. 1149-1152 ◽  
Author(s):  
Véronique Atger ◽  
Denise Malon ◽  
Marie Claude Bertiere ◽  
Françoise N'Diaye ◽  
Anik Girard-Globa
Keyword(s):  
High Density Lipoprotein ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Hdl Cholesterol ◽  
High Density ◽  
Lipoprotein Subfractions ◽  
Cylindrical Tubes ◽  
High Density Lipoprotein Subfractions ◽  
Gradient Gel Electrophoresis ◽  
Hdl2 Cholesterol

Abstract We used discontinuous gradients of polyacrylamide gel to determine the high-density-lipoprotein (HDL) subfractions HDL2 and HDL3 of serum lipoproteins. Serum (40 microL) prestained with Sudan Black was electrophoresed in cylindrical tubes over successive layers of 3.5%, 6%, 13%, and 17.5% acrylamide gels in a Tris.glycine buffer (3-4 h, 300 V). Very-low- (VLDL) and low-density lipoprotein (LDL) were retained by the 3.5% and 6% gels. HDL2 was concentrated at the interface between the 13% and 17.5% gels, and HDL3 migrated into the 17.5% gel. The distribution between HDL2 and HDL3 was obtained by densitometric scanning. Application of the respective percentages to HDL cholesterol assayed after phosphotung-state-Mg2+ precipitation of VLDL and LDL gave calculated concentrations of HDL2 and HDL3 cholesterol. The calculated values for HDL2 cholesterol were in excellent agreement with those for HDL2 isolated by ultracentrifugation (r = 0.920 for n = 120 sera; differences nonsignificant by Student's paired t-test). Besides being highly discriminating, the method is rapid, easily performed, and economical.

Alterations in high-density lipoprotein subfractions during postprandial lipidaemia induced by fat with and without ethanol

1988 ◽  
Vol 75 (2) ◽  
pp. 135-142 ◽  
Author(s):  
G. Franceschini ◽  
Y. Moreno ◽  
P. Apebe ◽  
L. Calabresi ◽  
E. Gatti ◽  
...  
Keyword(s):  
High Density Lipoprotein ◽  
Density Lipoprotein ◽  
Hdl Cholesterol ◽  
High Density ◽  
Acute Administration ◽  
Cholesteryl Esters ◽  
Lipoprotein Subfractions ◽  
Apo B ◽  
Postprandial Phase ◽  
High Density Lipoprotein Subfractions

1. Serum lipid and apolipoprotein levels, distribution and composition of high-density lipoprotein (HDL) sub-fractions and lecithin:cholesterol acryltransferase activity were analysed in nine normolipidaemic subjects, in whom a hypertriglyceridaemic state was induced by the acute administration of ethanol (40 g) plus fat (70 g) or of fat only. 2. Triglyceride (TG) levels increased by 180% 4–6 h after fat plus ethanol intake, the hypertriglyceridaemic response being inversely correlated with the basal HDL2 mass (r = −0.82). Serum apolipoprotein (apo) B levels rose by 8%, HDL–cholesterol decreased by 10% and HDL–TG increased by 57% at 6–8 h. 3. When ethanol was omitted, serum cholesterol and TG rose by 6% and 70%, respectively; both apo AI and apo B levels went up by 8%, whereas HDL-cholesterol rose progressively (15%) at 12 h. 4. The flotation rates of both HDL2 and HDL, increased, reaching a maximum 6–8 h after ethanol plus fat intake. These changes were due to an increase in TG and phospholipid contents, whereas cholesteryl esters and proteins decreased. 5. The alterations in HDL are attributable to the increase in TG-rich lipoproteins, to the stimulated cholesterol esterification (+ 15%) and to an enhanced transfer of newly formed cholesteryl esters to apo-B-containing lipoproteins in exchange for TG. 6. Changes in HDL properties were evident only when ethanol was given concomitantly with fat. 7. These findings suggest that in the postprandial phase lipoprotein changes may occur, which facilitate an improved removal of cholesterol from tissues.

Dual-precipitation method evaluated for determination of high-density lipoprotein (HDL), HDL2, and HDL3 cholesterol concentrations.

1988 ◽  
Vol 34 (11) ◽  
pp. 2322-2327 ◽  
Author(s):  
V C Dias ◽  
H G Parsons ◽  
N D Boyd ◽  
P Keane
Keyword(s):  
High Density Lipoprotein ◽  
Dextran Sulfate ◽  
Density Lipoprotein ◽  
Hdl Cholesterol ◽  
Comparison Method ◽  
High Density ◽  
Precipitation Method ◽  
Low Density ◽  
Present Technique

Abstract We evaluated a dual-precipitation method for determining cholesterol in high-density lipoprotein (HDL) and its subfractions HDL2 and HDL3. After total HDL was isolated by precipitation of very-low-density (VLDL) and low-density (LDL) lipoproteins with polyethylene glycol (Mr 8000), HDL2 was isolated from total HDL by precipitation with dextran sulfate (Mr 15,000), leaving HDL3 in the supernate. Concentration of total HDL cholesterol after precipitation of VLDL and LDL with PEG showed significant proportional and constant biases of -3.8% and 0.04 mmol/L, respectively, when compared with a phosphotungstic acid-based comparison method, although results by the two methods were correlated highly (r = 0.99, P less than 0.001). HDL2 and HDL3 cholesterol concentrations measured with the present technique were not different from those obtained by density-gradient ultracentrifugation or by combined precipitation-ultracentrifugation.

High-density lipoprotein subfractions as measured by differential polyanionic precipitation and rate zonal ultracentrifugation.

1982 ◽  
Vol 28 (10) ◽  
pp. 2040-2043 ◽  
Author(s):  
H S Simpson ◽  
F C Ballantyne ◽  
C J Packard ◽  
H G Morgan ◽  
J Shepherd
Keyword(s):  
High Density Lipoprotein ◽  
Density Lipoprotein ◽  
Hdl Cholesterol ◽  
Protective Role ◽  
High Density ◽  
Lipoprotein Subfractions ◽  
Functional Roles ◽  
High Density Lipoprotein Subfractions ◽  
Hdl2 Cholesterol

Abstract Recent interest in the putative protective role of high-density lipoprotein (HDL) and its subfractions against atherosclerosis has highlighted the need for a rapid, simple subfractionation procedure. Here we compared HDL subfractionation by two recently developed polyanionic-precipitation methods with the values obtained by rate zonal ultracentrifugation. A similar result for total HDL cholesterol was obtained by all three methods. However, HDL2 cholesterol as measured by the precipitation procedures was significantly higher than the zonal value, and HDL3 was lower. This reflects the different underlying principles involved in the separations and highlights the need for a clearer understanding of the functional roles of the HDL fractions.

Ultracentrifugation in swinging-bucket and fixed-angle rotors evaluated for isolation and determination of high-density lipoprotein subfractions HDL2 and HDL3.

1983 ◽  
Vol 29 (4) ◽  
pp. 656-663 ◽  
Author(s):  
P N Demacker ◽  
D F van Sommeren-Zondag ◽  
A F Stalenhoef ◽  
P M Stuyt ◽  
A van't Laar
Keyword(s):  
High Density Lipoprotein ◽  
Density Lipoprotein ◽  
High Density ◽  
Lipoprotein Subfractions ◽  
Fixed Angle ◽  
Serum Pool ◽  
Solvent Density ◽  
High Density Lipoprotein Subfractions ◽  
Coomassie Brilliant

Abstract We evaluated the density-gradient ultracentrifugation method in a swinging-bucket rotor (Anal Biochem 111, 149-157, 1981) in a slightly modified version for isolation and determination of high-density lipoprotein (HDL) subfractions. We prestained the serum with Coomassie Brilliant Blue R, which did not change the hydrated densities of the lipoproteins, and after only 2.2 X 10(8) gav . min obtained an equilibrium distribution of the lipoproteins along the gradient. The density distribution of the HDL of 120 sera obtained from apparently healthy persons and from patients with different types of hyperlipoproteinemia was bimodal. The HDL2 could be isolated in the density range 1.072-1.098 kg/L and the HDL3 at 1.100-1.176 kg/L, the latter fraction being more heterogeneous. At a solvent density of 1.100 we obtained similar results for HDL2-and HDL3-cholesterol by ultracentrifugation in two different fixed-angle rotors with tube angles of 15 degrees or 35 degrees. Independent of the rotor and the ultracentrifugation technique, subfractionation at d = 1.100 resulted in more distinct stained entities than ultracentrifugation at d = 1.125. In the swinging-bucket rotor procedure, interference by sinking pre-beta-lipoproteins was minimized because, having hydrated densities between 1.058 and 1.075, they could be removed without aspirating the HDL2. The method is both accurate and precise. For HDL2- and HDL3-cholesterol determined in a thawed frozen serum pool, CVs were 8.8 and 6.3%, respectively (n = 18).

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