中华检验医学杂志
中華檢驗醫學雜誌
중화검험의학잡지
CHINESE JOURNAL OF LABORATORY MEDICINE
2011年
5期
409-414
,共6页
龚岩%王建中%屈晨雪%袁家颖%汪润
龔巖%王建中%屈晨雪%袁傢穎%汪潤
공암%왕건중%굴신설%원가영%왕윤
阿司匹林%血小板活化%血小板功能试验%可重复性%结果
阿司匹林%血小闆活化%血小闆功能試驗%可重複性%結果
아사필림%혈소판활화%혈소판공능시험%가중복성%결과
Aspirin%Platelet activation%Platelet function tests%Reproducibility of results
目的 建立一种新的血小板活化释放试验用于监测阿司匹林的治疗反应.方法 采用ARA活化抗凝全血中血小板,应用血细胞分析仪检测MPC的变化.抽取5名健康志愿者外周血标本,取血后迅速混匀,用于MPC、LTAARA和血小板CD62P表达的检测,以优化检测条件,包括ARA活化浓度(分别为0、0.5、1.0、1.5、5.0、10.0 mmol/L)的选择和ARA活化血小板时间的选择(37℃水浴中5、10、20、30、40、50、60、70、80、90 min);评价该方法的稳定性(取血后0、1、2、3 h分别检测)、重复性(MPC、LTAARA、血小板CD62P表达的检测分别重复测定10次,计算批内及批间CV);并通过体外乙酰水杨酸试验验证该方法监测阿司匹林治疗反应的可行性.ARA活化血小板后用CD61-PerCP和CD62P-PE标记,流式细胞仪测定CD62P表达阳性的血小板百分率,对MPC与CD62P的相关性进行分析.选择口服阿司匹林至少7 d的患者为服药组,共93例;选择未口服或停止口服阿司匹林7 d以上的患者为对照组,共100例.用该方法检测服药组与对照组患者ARA(终浓度0.5 mmol/L)活化前后MPC的变化,并与LTAARA方法进行比较,评价该方法的准确性.结果 ARA终浓度为0.5 mmol/L时,血小板活化充分,MPC与血小板膜表面CD62P呈负相关(r=-0.755,P<0.01).37℃水浴活化30 min后MPC达到稳定,取血后室温3 h内MPC保持稳定,新鲜全血MPC的批内CV为1.35%,固定质控全血批间CV为0.71%和0.74%.随着全血中乙酰水杨酸浓度升高(0～6.95 μmol/L),ARA活化血小板后MPC逐渐升高,CD62P逐渐减低,二者呈负相关(r＝-0.765,P<0.01).服药组MPC差值为(8.2±8.6)g/L,MPC变化率为(3.4±3.6)%,对照组分别为(37.4±10.3)g/L、(15.7±4.0)%,差异均有统计学意义(t=21.522、22.409,P均<0.01).MPC变化率ROC曲线下面积为0.992,当临界值为8.7%时,其监测阿司匹林治疗反应的敏感度为96.8%,特异度为99.0%.MPC变化率与LTAARA具有很好的相关性(r=0.720,P<0.01).结论 本研究建立了一种新的监测阿司匹林治疗反应的血小板活化释放试验,可以替代LTAARA用于临床常规检测.
目的 建立一種新的血小闆活化釋放試驗用于鑑測阿司匹林的治療反應.方法 採用ARA活化抗凝全血中血小闆,應用血細胞分析儀檢測MPC的變化.抽取5名健康誌願者外週血標本,取血後迅速混勻,用于MPC、LTAARA和血小闆CD62P錶達的檢測,以優化檢測條件,包括ARA活化濃度(分彆為0、0.5、1.0、1.5、5.0、10.0 mmol/L)的選擇和ARA活化血小闆時間的選擇(37℃水浴中5、10、20、30、40、50、60、70、80、90 min);評價該方法的穩定性(取血後0、1、2、3 h分彆檢測)、重複性(MPC、LTAARA、血小闆CD62P錶達的檢測分彆重複測定10次,計算批內及批間CV);併通過體外乙酰水楊痠試驗驗證該方法鑑測阿司匹林治療反應的可行性.ARA活化血小闆後用CD61-PerCP和CD62P-PE標記,流式細胞儀測定CD62P錶達暘性的血小闆百分率,對MPC與CD62P的相關性進行分析.選擇口服阿司匹林至少7 d的患者為服藥組,共93例;選擇未口服或停止口服阿司匹林7 d以上的患者為對照組,共100例.用該方法檢測服藥組與對照組患者ARA(終濃度0.5 mmol/L)活化前後MPC的變化,併與LTAARA方法進行比較,評價該方法的準確性.結果 ARA終濃度為0.5 mmol/L時,血小闆活化充分,MPC與血小闆膜錶麵CD62P呈負相關(r=-0.755,P<0.01).37℃水浴活化30 min後MPC達到穩定,取血後室溫3 h內MPC保持穩定,新鮮全血MPC的批內CV為1.35%,固定質控全血批間CV為0.71%和0.74%.隨著全血中乙酰水楊痠濃度升高(0～6.95 μmol/L),ARA活化血小闆後MPC逐漸升高,CD62P逐漸減低,二者呈負相關(r＝-0.765,P<0.01).服藥組MPC差值為(8.2±8.6)g/L,MPC變化率為(3.4±3.6)%,對照組分彆為(37.4±10.3)g/L、(15.7±4.0)%,差異均有統計學意義(t=21.522、22.409,P均<0.01).MPC變化率ROC麯線下麵積為0.992,噹臨界值為8.7%時,其鑑測阿司匹林治療反應的敏感度為96.8%,特異度為99.0%.MPC變化率與LTAARA具有很好的相關性(r=0.720,P<0.01).結論 本研究建立瞭一種新的鑑測阿司匹林治療反應的血小闆活化釋放試驗,可以替代LTAARA用于臨床常規檢測.
목적 건립일충신적혈소판활화석방시험용우감측아사필림적치료반응.방법 채용ARA활화항응전혈중혈소판,응용혈세포분석의검측MPC적변화.추취5명건강지원자외주혈표본,취혈후신속혼균,용우MPC、LTAARA화혈소판CD62P표체적검측,이우화검측조건,포괄ARA활화농도(분별위0、0.5、1.0、1.5、5.0、10.0 mmol/L)적선택화ARA활화혈소판시간적선택(37℃수욕중5、10、20、30、40、50、60、70、80、90 min);평개해방법적은정성(취혈후0、1、2、3 h분별검측)、중복성(MPC、LTAARA、혈소판CD62P표체적검측분별중복측정10차,계산비내급비간CV);병통과체외을선수양산시험험증해방법감측아사필림치료반응적가행성.ARA활화혈소판후용CD61-PerCP화CD62P-PE표기,류식세포의측정CD62P표체양성적혈소판백분솔,대MPC여CD62P적상관성진행분석.선택구복아사필림지소7 d적환자위복약조,공93례;선택미구복혹정지구복아사필림7 d이상적환자위대조조,공100례.용해방법검측복약조여대조조환자ARA(종농도0.5 mmol/L)활화전후MPC적변화,병여LTAARA방법진행비교,평개해방법적준학성.결과 ARA종농도위0.5 mmol/L시,혈소판활화충분,MPC여혈소판막표면CD62P정부상관(r=-0.755,P<0.01).37℃수욕활화30 min후MPC체도은정,취혈후실온3 h내MPC보지은정,신선전혈MPC적비내CV위1.35%,고정질공전혈비간CV위0.71%화0.74%.수착전혈중을선수양산농도승고(0～6.95 μmol/L),ARA활화혈소판후MPC축점승고,CD62P축점감저,이자정부상관(r＝-0.765,P<0.01).복약조MPC차치위(8.2±8.6)g/L,MPC변화솔위(3.4±3.6)%,대조조분별위(37.4±10.3)g/L、(15.7±4.0)%,차이균유통계학의의(t=21.522、22.409,P균<0.01).MPC변화솔ROC곡선하면적위0.992,당림계치위8.7%시,기감측아사필림치료반응적민감도위96.8%,특이도위99.0%.MPC변화솔여LTAARA구유흔호적상관성(r=0.720,P<0.01).결론 본연구건립료일충신적감측아사필림치료반응적혈소판활화석방시험,가이체대LTAARA용우림상상규검측.
Objective To establish a new method for monitoring aspirin response by platelet activated-release experiment.Methods The platelets in whole blood were activated by ARA,and the MPC was measured by hematology analyzer.Blood samples were drawn from five healthy volunteers for measuring MPC,LTAARA and platelet membrane CD62P expression.Blood samples were mixed thoroughly right after venipuncture. The concentration of ARA (0,0. 5,1.0,1.5,5.0 and 10. 0 mmol/L) and the time for platelet activation (5,10,20,30,40,50,60,70,80 and 90 min in 37℃ water bathe) were optimized to evaluate the stability (0,1,2 and 3 h after venipuncture) and reproducibility (MPC, LTAARA and platelet membrane CD62p were measured ten times and the CV was calculated). Platelets were mixed with acetylsalicylic acid at different concentrations in vitro in order to verify the validity for monitoring aspirin response. The percentage of CD62p positive platelets after activated by ARA was measured using flow cytometer with CD61-PerCP and CD62p-PE antibodies. The correlation between MPC and CD62P was determined. 100 patients without taking or stop taking aspirin more than 7 days and 93 patients who took aspirin at least 7 days were enrolled. Duplicate measurements of platelet function were performed using the change of MPC (ARA 0. 5 mmol/L) and LTA (ARA 0. 5 mmol/L) among two patient groups to evaluate the accuracy of the new method. Results Platelcts were completely activated by ARA at final concentration of 0. 5 mmol/L. MPC negatively correlated with platelet membrane CD62p(r=-0. 755,P<0. O1 ). MPC was stable for 30 minutes in 37 ℃ water bathe after ARA activation. The result of MPC was consistent at room temperature within 3 hours after blood collection. This method had good reproducibility. CV in batch using fresh whole blood was 1.35% and CV between batches using commercial control whole blood were 0. 71% and 0. 74%. With the concentration of acetylsalicylic acid increased (0-6. 95 μmol/L), MPC increased as CD62P decreased, which showed negative correlation (r=-0. 765 ,P <0. 01 ). The difference of MPC before and after ARA activation (ΔMPC) and MPC variance ratio of the group taking aspirin were ( 8. 2±8. 6) g/L and ( 3.4±3.6) %, and they were (37.4±10. 3 ) g/L and ( 15.7±4.0) % in control group, respectively.ΔMPC and MPC variance ratio showed significant difference between the two groups ( t=21. 522, 22. 409, all P < 0. 01 ). Area under the ROC curve for MPC variance ratio was 0. 992 with sensitivity of 96. 8% and specificity of 99.0% for monitoring the aspirin response using the cut-off of 8. 7%. MPC variance ratio had good correlation with LTAARA (r = 0. 720, P < 0. O1 ). Conclusions A new method for monitoring aspirin response by platelet activated-release experiment is established. It may replace LTAARA for routine clinical examination.
