中华老年医学杂志
中華老年醫學雜誌
중화노년의학잡지
Chinese Journal of Geriatrics
2015年
4期
382-386
,共5页
陆鹏%郭志睿%张娟%刘静宁%张梦瑶%鲁翔
陸鵬%郭誌睿%張娟%劉靜寧%張夢瑤%魯翔
륙붕%곽지예%장연%류정저%장몽요%로상
金%纳米粒子%粒子大小%免疫测定%肌钙蛋白Ⅰ
金%納米粒子%粒子大小%免疫測定%肌鈣蛋白Ⅰ
금%납미입자%입자대소%면역측정%기개단백Ⅰ
Gold%Nanoparticles%Particle size%Immunoassay%Troponin Ⅰ
目的 可控制备不同直径的单分散胶体级金纳米颗粒(AuNPs),并将其应用于免疫层析法定性检测心肌肌钙蛋白Ⅰ(cTnⅠ),评估其检测效果.方法 以小尺寸AuNPs(15 nm)作为生长中心(种子),采用种子生长-热熟化法,成功地制备了四种单分散、大尺寸、柠檬酸钠稳定的胶体级AuNPs.作为对照,采用传统柠檬酸钠还原法,制备两种常规尺寸的AuNPs(20 nm、40 nm).六种尺寸AuNPs均标记鼠源cTnⅠ单克隆抗体,滴于聚酯结合垫上制成AuNPs结合物垫(金标垫),将与金标垫抗体配对的鼠源抗人cTnⅠ单克隆抗体和羊抗鼠多克隆抗体分别包被检测线和质控线,组装成免疫层析试剂盒检测cTnⅠ,从试剂盒信号强度、灵敏度、特异性、稳定性方面对其进行评估.并将四种大尺寸AuNPs制作的试剂盒与常规尺寸AuNPs制作的试剂盒进行检测效果对比.选取四种大尺寸AuNPs制作的cTnⅠ免疫层析试剂盒中检测效果最佳者进行临床样本检测及与商品试剂盒做检测效果比对.结果 采用种子生长-热熟化法制备的大尺寸AuNPs直径分别为50 nm、65 nm、79 nm、102 nm,尺寸分布呈单分散性.四种大尺寸AuNPs制作的免疫层析试剂盒检测cTnⅠ,其信号强度优于采用20 nm AuNPs制作的试剂盒(均P＜0.01),65 nm的AuNPs制作的试剂盒其信号强度优于各尺寸试剂盒(均P＜0.01).其最低检测限为0.50 μg/L.应用65 nm的AuNPs制作的cTnⅠ试剂盒检测100份血清,与化学发光免疫分析法结果进行对比分析,阳性符合率为97.3％,阴性符合率为100.0％;其灵敏度及同一cTnⅠ标准品浓度下检测信号强度优于艾博公司和博天科智公司同类产品(均P＜0.01).结论 种子生长-热熟化法可控制备直径为50～100 nm单分散、大尺寸、柠檬酸钠稳定的AuNPs,使得各类大尺寸AuNPs免疫层析试剂盒的研发成为可能.直径为65 nm的AuNPs可作为cTnⅠ免疫层析试剂盒的新选择.
目的 可控製備不同直徑的單分散膠體級金納米顆粒(AuNPs),併將其應用于免疫層析法定性檢測心肌肌鈣蛋白Ⅰ(cTnⅠ),評估其檢測效果.方法 以小呎吋AuNPs(15 nm)作為生長中心(種子),採用種子生長-熱熟化法,成功地製備瞭四種單分散、大呎吋、檸檬痠鈉穩定的膠體級AuNPs.作為對照,採用傳統檸檬痠鈉還原法,製備兩種常規呎吋的AuNPs(20 nm、40 nm).六種呎吋AuNPs均標記鼠源cTnⅠ單剋隆抗體,滴于聚酯結閤墊上製成AuNPs結閤物墊(金標墊),將與金標墊抗體配對的鼠源抗人cTnⅠ單剋隆抗體和羊抗鼠多剋隆抗體分彆包被檢測線和質控線,組裝成免疫層析試劑盒檢測cTnⅠ,從試劑盒信號彊度、靈敏度、特異性、穩定性方麵對其進行評估.併將四種大呎吋AuNPs製作的試劑盒與常規呎吋AuNPs製作的試劑盒進行檢測效果對比.選取四種大呎吋AuNPs製作的cTnⅠ免疫層析試劑盒中檢測效果最佳者進行臨床樣本檢測及與商品試劑盒做檢測效果比對.結果 採用種子生長-熱熟化法製備的大呎吋AuNPs直徑分彆為50 nm、65 nm、79 nm、102 nm,呎吋分佈呈單分散性.四種大呎吋AuNPs製作的免疫層析試劑盒檢測cTnⅠ,其信號彊度優于採用20 nm AuNPs製作的試劑盒(均P＜0.01),65 nm的AuNPs製作的試劑盒其信號彊度優于各呎吋試劑盒(均P＜0.01).其最低檢測限為0.50 μg/L.應用65 nm的AuNPs製作的cTnⅠ試劑盒檢測100份血清,與化學髮光免疫分析法結果進行對比分析,暘性符閤率為97.3％,陰性符閤率為100.0％;其靈敏度及同一cTnⅠ標準品濃度下檢測信號彊度優于艾博公司和博天科智公司同類產品(均P＜0.01).結論 種子生長-熱熟化法可控製備直徑為50～100 nm單分散、大呎吋、檸檬痠鈉穩定的AuNPs,使得各類大呎吋AuNPs免疫層析試劑盒的研髮成為可能.直徑為65 nm的AuNPs可作為cTnⅠ免疫層析試劑盒的新選擇.
목적 가공제비불동직경적단분산효체급금납미과립(AuNPs),병장기응용우면역층석법정성검측심기기개단백Ⅰ(cTnⅠ),평고기검측효과.방법 이소척촌AuNPs(15 nm)작위생장중심(충자),채용충자생장-열숙화법,성공지제비료사충단분산、대척촌、저몽산납은정적효체급AuNPs.작위대조,채용전통저몽산납환원법,제비량충상규척촌적AuNPs(20 nm、40 nm).륙충척촌AuNPs균표기서원cTnⅠ단극륭항체,적우취지결합점상제성AuNPs결합물점(금표점),장여금표점항체배대적서원항인cTnⅠ단극륭항체화양항서다극륭항체분별포피검측선화질공선,조장성면역층석시제합검측cTnⅠ,종시제합신호강도、령민도、특이성、은정성방면대기진행평고.병장사충대척촌AuNPs제작적시제합여상규척촌AuNPs제작적시제합진행검측효과대비.선취사충대척촌AuNPs제작적cTnⅠ면역층석시제합중검측효과최가자진행림상양본검측급여상품시제합주검측효과비대.결과 채용충자생장-열숙화법제비적대척촌AuNPs직경분별위50 nm、65 nm、79 nm、102 nm,척촌분포정단분산성.사충대척촌AuNPs제작적면역층석시제합검측cTnⅠ,기신호강도우우채용20 nm AuNPs제작적시제합(균P＜0.01),65 nm적AuNPs제작적시제합기신호강도우우각척촌시제합(균P＜0.01).기최저검측한위0.50 μg/L.응용65 nm적AuNPs제작적cTnⅠ시제합검측100빈혈청,여화학발광면역분석법결과진행대비분석,양성부합솔위97.3％,음성부합솔위100.0％;기령민도급동일cTnⅠ표준품농도하검측신호강도우우애박공사화박천과지공사동류산품(균P＜0.01).결론 충자생장-열숙화법가공제비직경위50～100 nm단분산、대척촌、저몽산납은정적AuNPs,사득각류대척촌AuNPs면역층석시제합적연발성위가능.직경위65 nm적AuNPs가작위cTnⅠ면역층석시제합적신선택.
Objective To prepare the monodisperse,colloidal gold nanoparticles (AuNPs) with controllable sizes (50 nm,65 nm,79 nm and 102 nm) for the qualitative detection of cardiac troponin Ⅰ (cTnⅠ) by immunochromatography assay,and to evaluate the effectiveness of the detection.Methods Four kinds of monodisperse citrate-stabilized AuNPs were prepared using small AuNPs as "growth centers" (seeds) by a seeded growth thermal aging protocol.As controls,two conventional AuNPs (20 nm,40 nm) were prepared by the traditional citrate-reduction method.The mouse monoclonal antibody against cTnⅠ labeled AuNPs were dropped on polyester mat to make AuNPs conjugate pad.The detection line and quality control line of immunochromatography assay kits for detection of cTnⅠ were coated by mouse anti human cTnⅠ monoclonal antibody paired with antibody in AuNPs and goat anti mouse polyclonal antiboy respectively.The six kinds of AuNPs were employed as color-labels in immunochromatography assay kits for detecting cTnⅠ,and the corresponding detection effects were evaluated in signal intensity,sensitivity,specificity and stability.The assay kit with the best performance was chosen and compared with the commercialized kits for the detection of cTnⅠ in clinical samples.Results Four kinds of monodisperse AuNPs with large sizes of 50 nm,65 nm,79 nm,102 nm respectively were successfully synthesized by the seeded growth thermal aging method.The signal strength of four kinds of kits produced by the four large-sized AuNPs was superior to the kits produced by 20 nm AuNPs in detecting cTnⅠ(all P＜0.01).The signal strength of the kits produced by 65nm AuNPs showed the best performance among the six kinds of AuNPs(all P＜0.01).The lowest detectable limit was 0.50 ng/ml.To compare the agreement of results from chemiluminescent immunoassay versus the results from kits produced by 65nm AuNPs,100 serum samples have been used for detecting cTnⅠ.Their positive coincidence rate was 97.30％ and negative coincidence rate was 100％,the sensitivity and signal strength of the kits produced by 65nm AuNPs was superior to similar products which produced by ABON and Bottests company(all P＜0.01).Conclusions Monodisperse,largesized,citrate-stabilized AuNPs are controllably prepared by a seeded growth-thermal aging method.The development of large-size AuNPs-based immunochromatography assay kits is feasible.65 nm AuNPs can be a suitable candidate for cTnⅠ immunochromatography assay kit.Our findings provides a new idea for the current immunochromatography assay kits which still adopt small-sized AuNPs as color labels.
