血脂分析測試的國家指南

1、National Guidelines on Lipid Profile Testing,E-mail:yanshengkai@sina.com,Dr. Sheng-kai Yan , PhDDepartment of Laboratory Medicine, Peking Union Medical College HospitalProfessor zhou xin Department of Laboratory Med

2、icine,Zhongnan hospital of wuhan university.,Preface,Blood lipid analysis has a substantial importance towards the prevention and management of atherosclerosis and coronary heart disease (CHD).It is also being widely

3、applied to the studies of many other related clinical conditions, such as diabetes mellitus, kidney diseases, and metabolic disorders in postmenopausal women.,Recently, the Lipid Expert Panel of the Chinese Society of La

4、boratory Medicine (CSLM) of the Chinese Medical Association (CMA) has come up with the practical guidelines and recommendations on lipid profile testing.lipid profile testing: total cholesterol (TC) triglyce

5、rides (TG)high density lipoprotein–cholesterol (HDL-C)low density lipoprotein-cholesterol (LDL-C)apolipoprotein A1 (ApoA1) apolipoprotein B (ApoB) lipoprotein(a) [Lp(a)],The Guidelines include the content as follows

6、,,PrefacePreanalytical factors affecting lipid test resultsMethods of lipid analysisReagent selection criteria and practical guidelinesClinical significance of cutoff values in the interpretation of lipid profile res

7、ults,See: Chin J Lab Med, 2003, 26(3): 182~184中華檢驗醫(yī)學(xué)雜志,2003,26(3):182-184,Preanalytical Factors Affecting Lipid Test Results,A subject’s lipid profile should be measured when the individual is in a steady metabolic stat

8、e.Subjects should maintain their usual diet and weight for at least 2 weeks prior to the measurement of their lipids or lipoproteins.Subjects should not perform vigorous physical activity during the 24 hours prior to t

9、esting.,Recommendations for minimizing preanalytical variation,Multiple measurements should be performed whthin 2 months, at least one week apart, before making a medical decision about further action. Fasting or non-f

10、asting specimens can be used for TC testing. However, a 12-h fasting specimen is required for TG and recommended for lipoproteins.The subject should be seated for at least 5 minutes before specimen collection.The tourn

11、iquet should not be kept on more than one minute during venipuncture.,Suggestions to reduce the preanalytical variations on blood lipid profile testing,Both serum and plasma are suitable for analysis. Serum is more commo

12、n and convenient. The result obtained from EDTA plasma should be corrected for the dilution effect by a factor of 1.03.The separated serum should be processed as soon as possible. Samples that could not be analyzed

13、within 24 hours, can be stored at 4℃ for one week or at -20℃ for several months and at -70℃ for at least half a year. Repeated freeze and thaw should be avoided.,Suggestions to reduce the preanalytical variations on

14、 blood lipid profile testing,Methods of Lipid Analysis,Analytical Methods For routine lipid analysis,Serum TC: Enzymatic method(CHOD-PAP method)Serum TG: Enzymatic method( GPO-PAP method) Se

15、rum HDL-C: Homogeneous methods Serum LDL-C: Homogeneous methods Serum ApoA1/ApoB and Lp(a): Immunoturbidimetry(ITA) method Immunonephelometry(INA) method

16、 (The first choice would be ITA, followed by INA),Serum HDL-C —Homogeneous methods,Clearance method Synthetic polymer/ detergent HDL-C assay, SPD Daiichi Pure Chemicals Co. Catalase HDL-C assa

17、y, CAT Denka Seiken Randox Co. Reference Diagnostics Polymedco,Serum HDL-C —Homogeneous methods,PEG-modified enzyme HDL-C assay，PEGME Kyowa Medex Co. Roche Diagnostics

18、 Centronic GmbH,,Immunoseparation method, IS International Reagents Corp HDL-C assay，IRC International Reagents Co./Sysmex Antibody immunoseparation HDL-C assay ，AB Wako Pure Chemicals Industry,Serum H

19、DL-C — Homogeneous methods,,Polyanion Polymer/ detergent HDL-C assay，PPD Daiichi Pure Chemicals Co. Genzyme Diagnostics,Serum HDL-C — Homogeneous methods,Clearance method Surfactant LDL-C assay，SUR

20、 Daiichi Pure Chemicals Co. Genzyme Diagnostics Catalase LDL-C assay，CAT Denka Seiken RANDOX Polymedco Kyowa Medex Roche,Serum LDL-C —Homogeneous method,Protecting reagent LDL-C assa

21、y，PRO Wako Chemicals Sigma DiagnosticsCalixarene LDL-C assay，CAL International Reagents Co./Sysmex Solubilization LDL-C assay，SOL,Serum LDL-C —Homogeneous method,Serum Lp(a)—Immunoturbidimetry/immunone

22、phelometry,The reagent should preferably be polyclonal or mixed monoclonal antibodies, that could recognize different epitopes of the Apo(a) molecule.ITA is more preferable than INA.,Spectrophotometers and semi-/automat

23、ic biochemical analyzers would be suitable for analysis once verified for proper functioning. All samplers, dilutors, pipettes and micropipettes must be calibrated.Automatic biochemical analyzers (fully or semi-automati

24、c) are recommended for use in blood lipid testing.,Requirements on Analytical Instruments,Parameters should be set according to the manufacturers’ instructions and assigned calibrator values on the package insert. The p

25、arameters should not be liberally changed.,Setting the Parameters,Laboratories should use accredited quality control (QC) materials with serum matrix as the internal quality control. The QC materials should cover both t

26、he normal and pathological ranges. There are QC materials specifically designed for lipid analysis that laboratories could consider to purchase.,Quality Control,While choosing individual QC materials, one should conside

27、r carefully the dynamic range and the target values for the corresponding analytical methods. A parallel run should be performed for an overlapping period with both the current and new lot control materials. Enrolment

28、to external quality assessment programs is a MUST.,Quality Control,Reagent Selection Criteria and Practical Guidelines,,Inaccuracy and Imprecision,Inaccuracy(Bias) Imprecision(CV) Total errors * TC ≤

29、±3% ≤3% 8.9% TG ≤±5% ≤5% ≤15% HDL-C ≤±5% ≤4% ≤13% LDL-C ≤&

30、#177;4% ≤4% ≤12% ApoAI ≤±3% ≤±5% ApoB ≤±3% ≤±5% Lp(a) ≤±4% ≤±10% *Tot

31、al errors=Bias%+1.96CV,Analytical Sensitivity,When phenol is employed in enzymatic analysis of serum TC, the absorbance of TC = 5.2 mmol/L at 500nm (A500nm) is about 0.30-0.35. Therefore, A500nm of 0.005 should give 0.08

32、 mmol/L TC.The sensitivity of TG enzymatic analysis should be A500nm ≥0.2 at TG 2 mmol/L.,Analytical Sensitivity,In using homogeneous assays for HDL-C and LDL-C, the minimal measurable level should be 0.01 mmol/L.The l

33、owest detection limits for serum ApoA1 and ApoB by immunoturbidimetry or immunonephelometry should be 0.5 g/L, and that for Lp(a) should be 5 mg/L.,Linearity,The upper limit of linearity is 13 mmol/L when using the dilut

34、ion ratio of 1:100 in enzymatic analysis of TC. It will lower the upper limit if smaller dilution ratios are used.The linearity of the enzymatic TG assay should at least be 11.3 mmol/L (1000 mg/dL). The linearity of h

35、omogeneous assays for HDL-C and LDL-C should at least be 2.59 mmol/L and 7.77 mmol/L respectively.,Linearity,The linearity of serum ApoA1 and ApoB by ITA or INA should not be less than 2.0 g/L and that of Lp(a) not less

36、than 800 mg/L, respectively.,Specificity,In enzymatic analysis of serum TC, the color reaction is subject to certain degree of spectral interferences from various non-cholesterol sterols. Normally, there is only negligib

37、le amount (about 1%) of these non-cholesterol sterols in the blood. In enzymatic analysis of TG, the lipoprotein lipase (LPL) can hydrolyze TG and also mono-glycerides and di-glycerides. The latter two constitute about

38、3% of total TG measured.,Specificity,The recoveries in homogeneous assays of HDL-C and LDL-C, immunoturbidimetry of serum ApoAI, Apo B and Lp(a) should preferable be in the range of 90%-110%. In general, the measurements

39、 should not be affected by other lipoproteins.,There is no significant interference up to hemoglobin concentration of 2g/L, bilirubin concentration of 0.1g/L in enzymatic analysis of serum TC.Interference in enzymatic

40、analysis of TG is similar to that of the TC assay. There will be negative interference when bilirubin >100?mol/L or ascorbic acid>170?mol/L. Hemoglobin will cause spectral interferences. Grossly hemolysed samples a

41、re not suitable for analysis.,Interferences,There is no significant interference of TG < 5.65 mmol/L (500 mg/dl), bilirubin <513 ?mol/L (30 mg/dl), and Hb <5 g/L in most homogeneous assays for HDL-C and LDL-C, a

42、s well as the immunoturbidimetric or immunonephelomentric assays for serum ApoA1, ApoB and Lp(a).,Interferences,Reagent Stability,Lyophilized reagents can usually be stored at least for 1 year if kept unopened at 2℃~8℃.

43、Reconstituted cholesterol and TG enzymatic reagents should be stored at 2℃~8℃for 2 days. Reagents should be discarded if pink color is seen. The absorbance at 500nm of the reagent blank should be ≤0.05. Unopened reagen

44、t solutions should be stable for at least 6 months at 2℃~8℃ and for 1 month after being opened.,Reaction Rates,The reaction should not be longer than 5 min and 8 min at 37℃ for enzymatic analysis of serum cholesterol and

45、 TG respectively. The endpoint of immunoturbidimetric assays for serum ApoA1, ApoB and Lp(a) could be determined according to their reaction curves shown in the automatic analyzer. Generally, reaction time of 8 ~ 10 min

46、utes is acceptable.,Calibration,The user should use the calibration materials provided by the manufacturer. The calibration materials should be traceable to the reference methods. One should avoid using different brands

47、 of calibration materials as to ensure consistence of the performance.,Calibration,The international standard serum preparations of WHO-IFCC should be used in the ITA or INA assays for ApoA1, ApoB and Lp(a). At least fi

48、ve different concentrations should be used to cover the dynamic range of the measurement. Data reduction method should be used for curve-fitting. Quality control preparations should never be used as standard materials f

49、or calibration.,It is essential to have standardization in lipid analysis in order to ensure the accuracy of measurements and most importantly, traceability of the results. A routine lipid profile testing should include

50、 TC,TG, HDL-C and LDL-C. Laboratories could perform ApoA1, ApoB and Lp(a) if they were capable and on the basis of conditioned clinical judgment.,Conclusion,Blood lipid profile testing mainly reflects the status of lipi

51、d metabolism of an individual and could be used for risk assessment for CHD. One should not use a risk indicator as a diagnostic tool. Yet, its application outlined a strategy and therapeutic approach for primary preven