ACL与抗β2GP1测定推荐指南发布

摘要：1.样本要求 : 血清：建议用无溶血和无脂样本，避免样本过热（56℃以上30分钟）。 血浆：厂商需详细说明样本类型，包括使用的抗凝剂。样本中血小板含量应小于10000/ul。使用血浆时要考虑抗凝剂所致的稀释作用。

2.aCL及抗β2GP1亚型测定: IgM型及IgG型aCL及抗β2GP1抗体均需测定。当其它测定均阴性但仍怀疑APS时，需测定IgA型aCL及β2GP1。

3.抗 原: aCL：应使用心磷脂+（牛或人）-β2GP1。 抗β2GP1：应使用人源性β2GP1，测定时使用负电荷（高结合力或γ-照射）板。

4.结果定量: aCL：需测定GPL/GML单位，建立范围（低度、中度和高度阳性）。抗β2GP1：暂无通用的测定单位，建议建立国际或通用测量单位。

5. 标准: 厂商及实验者需选择可靠的标准来准备再次校准（多克隆或单克隆）。采用已发表的操作步骤，参照首次标准来对比和验证所建议的再次校准。如有可能，应使用患者的血清进一步确定测定系统的一致性程度。更重要的是，产品及质控标准应遵从FDA的最佳良好操作规范（GMP, Good Manufacturing Practices）指南或等效的质量保证程序。需有从推荐标准到任何再次校准的描绘记录。

6. 校准曲线: 需有多点校准、正确的统计拟合及计算方法。每检测一次，均应有校准曲线。当测量读数与期望值之间的相关系数小于0.90，不能使用该校准曲线。

7. 精确度: ELISA中，CV（变异系数）应小于0.10。需多层面（包括低~中滴度阳性界值）评估精确度。 对于商业试剂盒，产品操作手册上应标明预期精度。

8.阳性/阴性对照: 每做一次测定时，至少需一个阳性对照，以监测两次测定间的差异。理论上，阳性对照值应在试剂盒的低~中滴度界值范围内，以获得推荐的精度值。阳性对照值还应在测定的线性范围内。同样，每做一次测定时，需一个阴性对照，阴性对照值应低于界值。当阴性/阳性对照值落在所设立的范围外时，本次测定应作废。

9. 单次/重复测定: 若单次测定能获得满意的精确度，无需做重复测定。

10.界值计算: 采用非参数百分比法确定界值。鼓励厂商根据第95和99百分位来确定和报告界值。临床实验室使用商业试剂盒时，需核对厂商所提供的界值。

11.类风湿因子的干扰: 类风湿因子可影响测试结果，这应在评价时加以强调.

12.结果汇报: 结果应以单位或半定量范围报告。包括阴性、“可疑”、中度阳性和高度阳性。

13.注释：强烈建议在对检测结果进行解释时，应列入有助于临床医生的评价。厂家应公开来自于临床研究的信息，这也许有助于结果的解释。

附原文：1.Specimen requirements：Serum: Heat inactivation at 56°C for 30 minutes should be avoided. Use of nonhemolyzed, nonlipemic samples is recommended. Plasma: Manufacturers must specify the specimen type, including the anticoagulant used. Platelet-poor plasma (<10 ,000/ul) is required. use of plasma should take into consideration the dilution factor that may be produced because of the anticoagulant.2.isotype of acl and anti-β2gpi tested：the igg and igm isotypes are recommended for both acl and anti-β2gpi. the iga isotype is recommended for both acl and anti-β2gpi when results of all other tests are negative and aps is still suspected. 3.antigen：acl: cardiolipin + (bovine or human) -β2gpi should be used. anti-β2gpi: β2gpi of human origin should be used，on a negatively charged (“high” binding or gamma-irradiated) plate. 4.quantitation of results：acl: gpl/mpl units should be measured and ranges (low-positive, medium/moderate-positive, highpositive) established. anti-β2gpi: universal units of measurement are not available. development/establishment of international/universal units of measurement is recommended. 5.standards：manufacturers and test users are strongly encouraged to select a reliable standard to prepare secondary calibrators (polyclonal or monoclonal). the proposed secondary calibrators should be compared and validated against the primary standard, using published and accepted procedures. selected groups of actual patient sera should be used if possible, to further establish the extent of agreement in the assay/test system. most importantly, the production and the quality control of the standards should be subjected to fda good manufacturing practices guidelines or an equivalent quality assurance program. a record of traceability from the recommended standards to any secondary calibrators is required. 6.calibration curves :multipoint calibration and use of statistically correct fitting and calculation methods are required. a calibration curve should be included in each run. the calibration curve should be rejected if the correlation coefficient between assay readings and expected values of the calibrators is <0.90. 7.precision : cv of <10% is recommended for elisas. precision should be evaluated at multiple levels of positivity, including low-positive to medium/moderatepositive cutoff level. for commercial kits, expected precision should be published in manufacturers’ instruction manuals. 8.positive/negative control: incorporation of at least 1 “external” positive control in every run to monitor interassay variation is recommended. ideally, use of a sample with a value approximately at the low-positive to medium/moderate-positive cutoff of the assay/kit and aiming to achieve the recommended precision (cv) is strongly encouraged. the value of this control should be also within the linear range of the assay/kit. similarly a “negative” control with values below the cutoff of the assay should be used in each run. a run should be rejected if the result with either the positive or the negative control falls out of the established range. 9.singlet/duplicate measurements : if precision is satisfactory based on single testing, then duplicate measurements may not be required. 10.cutoff calculation :cutoffs should be established using a nonparametric percentile method. manufacturers are encouraged to establish and to report cutoffs based on the 95th and 99th percentiles. clinical/diagnostic laboratories using commercial kits should validate/verify cutoffs provided by the manufacturer. 11.rheumatoid factor interference: rheumatoid factor can affect the results of the tests, and this should be addressed in the interpretative comments. 12. reporting of results: results should be reported in units and in semiquantitative ranges. interpretations should include negative, “indeterminate” or grey zone, medium/moderate-positive, and high-positive. 13.interpretative comments: inclusion of comments to assist clinicians in the interpretation of test results is strongly recommended. manufacturers should disclose information based on evidence derived from clinical studies that may assist with the interpretation of results.

引自：Gabriella Lakos,1 Emmanuel J. Favaloro,2 E. Nigel Harris,3 Pier Luigi Meroni,4 Angela Tincani,5 Richard C. Wong,6 and Silvia S. Pierangeli7 International Consensus Guidelines on Anticardiolipin and Anti-β2-Glycoprotein I Testing. Arthritis Rheumatism，2012, 1–10 DOI 10.1002/art.33349