Blood Collection

 Proper patient preparation and sampling are essential for an accurate lipid profile evaluation.

Venous sampling for lipid parameters measurement should be performed under the following conditions:

 • Habitual diet, including alcohol consumption, up to 24 h before collection.

 • Whenever possible, discontinue therapies that affect lipemia (diuretics, beta-blockers, etc.) 3 weeks before per forming blood collection.

• Do not test before 2–3 months after childbirth or trauma, such as surgery, or serious illness, such as myocardial infarction (after myocardial infarction, there is an increase in triglycerides and a decrease in cholesterol).

 • Do not test until 2 weeks following an episode of inflammation (inflammation is associated with a reduction in total and LDL cholesterol, which is proportional to basal cholesterol concentration).

• Fasting is not required. Traditionally, fasting for 12–14 h was required for lipid analysis. Recently, it has been shown that analysis performed on samples obtained from fasting and nonfasting patients give overlapping results for TG, C-LDL, and C-HDL. TGs are affected by food, resulting in an increase of only about 27 mg/dL depending on composition and time since the meal.

 • Maintain venous stasis only as long as necessary (maximum 1 min) and remove the tourniquet as soon as the blood begins to flow.

• The sample can be serum or plasma.

• Results should be confirmed on at least two samples collected 7 days apart.

Intraindividual Variability

 There is considerable intraindividual variability in circulating lipid levels. In particular, variability of 5–10% for TC and >20% for TGs has been reported, especially in patients with elevated TGs.

These data are required to calculate significant differences between successive values of the same patient.

 Intraindividual variability is due, in part, to analytical variability but also to environmental factors such as diet, physical activity, and seasonal variations; for example, TG and C-HDL levels increase in winter.

 Measuring Methods

 Enzymatic colorimetric methods are commonly used for lipid assay.

 Total Cholesterol

The CHOD/POD/Trinder enzymatic method is uniformly used in laboratories for the TC assay. According to the National Cholesterol Education Program (NCEP) guide lines, to ensure an adequate quality of the assay, the values obtained by the individual laboratory should not deviate from the value of the reference method by more than 3% and the measurement methods should have an inaccuracy (expressed as a coefficient of variation) of less than 3%. The total error must be <8.9 %.

The desirable value of TC is ≤5.00 mmol/L (≤190 mg/ dL; Table 1).

LDL-Cholesterol

In most laboratories, the C-LDL is calculated by the Friedewald formula:

in mmol/L: C-LDL = TC - C-HDL - (TG/2.2) in mg/dL: C-LDL = TC - C-HDL - (TG/5)

The value of C-LDL calculated according to this formula must consider the following assumptions:

 • Methodological errors that may accumulate during the analysis of each of the three parameters included in the formula (CT, TG, and C-HDL).

 • The formula can be applied assuming that there is a constant cholesterol/triglyceride ratio in VLDL. In the presence of elevated TG values (>4.5 mmol/L or >400 mg/dL), the formula cannot be applied because of the possible presence of circulating chylomicrons and VLDL remnants.

 • The formula may not be applicable when blood is collected under nonfasting conditions. In that case, non-HDL cholesterol should be determined.

 Recently, direct methods (immunological or ultracentrifuge) have been developed for the determination of C-LDL whose values are comparable with those obtained by the formula. The direct methods would seem to have analytical performances superior to the Friedewald formula but, similarly to the formula, cannot be applied in the presence of high TG values.

 According to NCEP guidelines, analytical performance should have an inaccuracy and bias <4 % with a total error <12%.

In general, desirable values should be ≤3.00 mmol/L or ≤115 mg/dL (Table 1). Specifically, based on the subject’s characteristics, the desirable values are:

• ≤3.00 mmol/L or ≤115 mg/dL in healthy subjects without cardiovascular disease and without other risk factors for cardiovascular disease

 • ≤2.60 mmol/L or ≤100 mg/dL in subjects with other risk factors for cardiovascular disease but without current or previous cardiovascular disease

 • ≤1.8 mmol/L or ≤70 mg/dL in subjects with prior or ongoing cardiovascular disease

Table1. Desirable plasma values of lipids, lipoproteins, and apolipoproteins

HDL Cholesterol

 In the past, C-HDL was measured by methods that involved initial precipitation of ApoB-containing lipoproteins (VLDL and LDL) and subsequent determination of the cholesterol present in the supernatant (C-HDL) by enzymatic methods. Currently, these methods have been replaced by “direct” methods that allow measuring C-HDL in a homogeneous phase without the need for prior separation of HDL from other lipoproteins.

According to NCEP guidance, analytical performance should have an inaccuracy <4% and bias <5% with a total error <13%.

Desirable values should be ≥1.00 mmol/L or ≥40 mg/dL for males and ≥1.20 mmol/L or ≥45 mg/dL for females (Table 1).

For the correct evaluation of cardiovascular risk, the risk index, i.e., the CT/C-HDL ratio and the C-LDL/C-HDL ratio, must be considered. The CT/C-HDL ratio in healthy subjects must be <5 for males and <4.5 for females. Regarding the C-LDL/C-HDL ratio, the risk is defined as low when the ratio is 1.47, medium 3.22, moderate 5.0, and high 6.14.

Triglycerides

TGs are commonly assessed by enzymatic methods that measure the free glycerol produced by TGs hydrolysis. Although these methods also measure free glycerol obtained from lipase-mediated hydrolysis of circulating TGs, its con centration is usually negligible. In rare conditions, such as patients with severe multiorgan diseases, a hyperglycerolemic condition may occur, resulting in excessively high false TG values. In these cases, it would be appropriate to use methods for measuring true TG with subtraction of free glycerol.

Triglycerides have a particularly high intraindividual bio logical variability. In particular, smoking, physical exercise, and alcohol can significantly modify TG levels and, therefore, before collecting the sample, it is important to take pre cautions. In addition, the time of sampling is important because, regardless of food intake, TG concentration varies significantly throughout the day. Therefore, collecting the sample in the early morning would be preferable. TGs can be assayed under nonfasting conditions for general screening and cardiovascular risk assessment.

According to NCEP guidelines, analytical performance should have an imprecision and bias <5 % with a total error <15%.

Desirable values should be ≤1.70 mmol/L or ≤150 mg/ dL (Table 1).

Non-HDL Cholesterol

 Non-HDL cholesterol is helpful for quantifying atherogenic lipoproteins. It is obtained by subtracting the C-HDL value the only nonatherogenic cholesterol) from the TC value. It is advisable to evaluate this parameter when TGs are ≥150 mg/dL because TG-rich lipoproteins also carry an atherogenic potential.

Apolipoproteins

The ApoA-I and ApoB apolipoproteins are not evaluated in clinical laboratory practice; their evaluation represents a second level test to be performed in selected cases. In particular, ApoB, being the main apolipoprotein of VLDL, IDL, and LDL, provides information on atherogenic lipoproteins over lapping those provided by non-HDL cholesterol, while ApoA-I provides information on nonatherogenic lipoproteins, overlapping those provided by C-HDL.

There are no standard methods for the apolipoprotein measurement. However, it is recommended that commercial kits be used whose calibrator values are assigned based on internationally recognized preparation.

The reference values for ApoB are ≤100 mg/dL or ≤1.00 g/L and for ApoA-I are ≥125 mg/dL or 1.25 g/L (Table 1).

ApoC-III has recently been identified as a potentially important risk factor. It is a key regulator of TG metabolism and high plasma levels are associated with high plasma levels of TG and VLDL. In addition, mutations that cause loss of function are associated with low TG levels, and reduced cardiovascular risk. However, the role of ApoC-III in clinical practice is not yet known.

Lp(a)

 Several immunometric methods for Lp(a) measurement are available but are not yet standardized. Currently, Lp(a) assay is not recommended for cardiovascular risk assessment in the general population. However, it should be considered in subjects with high risk (familial, high C-LDL, and low C-HDL levels) or with a strong family history of premature atherothrombotic disease. Risk is considered significant when Lp(a) levels are >30 mg/mL.

مواضيع ذات صلة

Diagnosis of Familial Dyslipidemia

5-hydroxyindoleacetic acid (5-HIAA)

17-hydroxycorticosteroids (17-OCHS)

Diagnosis of Dyslipidemia

Familial Hypercholesterolemia

human T-cell lymphotropic virus (HTLV) I/II antibody

human placental lactogen (hPL, Human chorionic somatomammotropin [HCS])

human papillomavirus (HPV test, HPV DNA testing, HPV Genotyping, HPV mRNA testing)

human lymphocyte antigen B27 (HLA-B27 antigen, Human leukocyte A antigen, White blood cell antigens, Histocompatibility leukocyte A antigen)

Nucleic Acid –Based Tests

human chorionic gonadotropin (hCG, Pregnancy tests, hCG beta subunit)

homocysteine (Hcy)