簡介：中文中文2830字出處出處WANGJ,GUODONGLIUAULTRASENSITIVEELECTRICALBIOSENSINGOFPROTEINSANDDNA鈥CARBONNANOTUBEDERIVEDAMPLIFICATIONOFTHERECOGNITIONANDTRANSDUCTIONEVENTSJJOURNALOFTHEAMERICANCHEMICALSOCIETY,2004,1261030101超靈敏免疫和超靈敏免疫和DNADNA電化學生物分析電化學生物分析應用碳納米管放大分子識別和傳導過程應用碳納米管放大分子識別和傳導過程蛋白質(zhì)和DNA檢測技術在基因疾病的診斷和治療，傳染性疾病的檢測，藥物的開發(fā)，生物戰(zhàn)爭預警中起著非常重要的作用。這些生物檢測通常依賴DNA雜交或抗原抗體相互作用，可達到超靈敏度的檢測。由于電化學傳感器具有靈敏度高、簡單、可微型化，低成本和高需求的特點，非常適用于生物檢測。酶標記在蛋白質(zhì)和DNA超靈敏電化學生物親和性檢測中起著很大的作用。HLELLER，小組5，6通過DNA上連接HRP酶標記物和采用一種可加快電子傳遞的氧化聚合物可實現(xiàn)DNA的高靈敏度的電化學檢測（低至5ZMOL）。WILLNER,S小組7～8通過生物催化沉積酶反應產(chǎn)物可獲得信號的多重放大，從而實現(xiàn)極低的檢測限（25AMOL），酶聯(lián)電化學蛋白質(zhì)檢測信號可通過雙酶底物體系底物循環(huán)或者酶產(chǎn)物的離子交換富集產(chǎn)物來進行放大。然而，在電化學生物檢測中，放大生物識別傳導信號仍是一個重大的挑戰(zhàn)。為了滿足蛋白質(zhì)和核酸電化學檢測高靈敏度的要求，我們需要新的方法，通過酶生物催化反應來放大信號。在本文中，我們利用碳納米管（CNTS）顯著放大蛋白質(zhì)和DNA的識別和電化學傳導信號。CNTS所特有的電學性能、化學性能和機械性質(zhì)使其非常適用于電化學傳感器1，2。大多數(shù)CNT傳感器主要是利用CNTS特有的表面性質(zhì)來促進在生物催化裝置中電子轉移反應，在我們新的生物親和性檢測中（圖1），CNTS起著放大識別和傳導信號的雙重作用，也就是CNTS上載有大量酶及積累了大量酶反應的產(chǎn)物。這些新奇的方法和CNTS預富集的功能反映了CNTS具有很大的比表面積，并可用ALP酶標來證實。通過采用CNTS的放大處理從而降低檢測限的一些方法已被報道過，因此很適用于電化學DNA檢測。圖，這些微觀圖片是HITACHIH7000儀器在工作電壓為75KV下拍下的。從圖3DNA分子雜交（A）和抗原抗體生物檢測中可以看到由于CNT的雙重放大作用引起了傳感信號的明顯增強。基于單酶標記物和一個玻碳電極的傳統(tǒng)檢測方法既不能對10PGML1的目標DNA（A，A）也不能對80PGML1的IGG（B，B）產(chǎn)生響應?；谳d有ALP酶的CNTS（B）的第一放大步驟為這些分析物的低濃度檢測提供了方便。單ALP酶檢測即使在分析物濃度較高（1000倍）的條件下仍顯示一個較低的信號（圖中未標出）。改進方法后的檢測靈敏度達到將近104，正好與每個CNT上載有的ALP酶估計量相一致。用涂覆上鏈霉親合素的聚苯乙烯代替CNT來裝載粒子獲得的靈敏度增強約僅為單酶檢測的50倍。在信號第二放大途徑，即用CNT修飾傳感器可獲得更強的DNA和蛋白質(zhì)檢測信號（約是用鏈霉親合素修飾聚苯乙烯檢測的30倍）（C）。后者反映了在CNT層強烈地吸附著大量游離的Α奈酚。在CNT上富集產(chǎn)物的現(xiàn)象可用由沉積時間的不同而引起的Α奈酚信號的突增來描述（與裸電極上產(chǎn)生的時間信號關系相比較；見圖2的支持信息）。圖310PGML1目標寡核苷酸（A）和80PGML1IGG（B）分別在玻碳電極（A）單ALP酶標（B）和載有大量ALP酶標的CNT上用計時電勢分析法進行檢測產(chǎn)生的信號。在檢測中（C）除了使用CNT修飾的玻碳電極外與（B）均相同。磁性粒子量，50UG；分別進行20和30分鐘的DNA雜交和抗原/抗體免疫反應；樣品體積，50UL。檢測，往樣品中加入50ULΑ奈基磷酸鹽（50MM）溶液進行酶反應20分鐘。產(chǎn)物Α奈酚的測量在裸或者是已用

簡介：MICROSATELLITEINSTABILITYAND/ORLOSSOFHETEROZYGOSITYINYOUNGGASTRICCANCERPATIENTSINITALYYIHHORNGSHIAO1,DANIELABOVO2,MARIAGUIDO2,CARLOCAPELLA3,MAUROCASSARO2,GRAZIELLABUSATTO2,VALENTINARUSSO2,4,ANGELOSIDONI5,ANNARPARENTI2ANDMASSIMORUGGE21LABORATORYOFCOMPARATIVECARCINOGENESIS,NCIFCRDC,NATIONALINSTITUTESOFHEALTH,FREDERICK,MD,USA2DEPARTMENTOFONCOLOGICALANDSURGICALSCIENCES,CATTEDRADIISTOCHIMICAEIMMUNOHISTOCHIMICAPATOLOGICA,IIICATTEDRADIANATOMIAPATOLOGICA,UNIVERSITA′DEGLISTUDI,PADOVA,ITALY3DEPARTMENTOFCLINICALANDBIOLOGICALSCIENCE,UNIVERSITYOFPAVIA,VARESE,ITALY4DEPARTMENTOFPATHOLOGY,UNIVERSITYOFCATANIA,CATANIA,ITALY5DEPARTMENTOFPATHOLOGY,UNIVERSITYOFPERUGIA,PERUGIA,ITALYGASTRICCANCERSARERARELYDIAGNOSEDBEFORETHEAGEOF40YEARSANDTHEINCIDENCEREACHESAPEAKDURINGTHE7THDECADEINTHEGENERALPOPULATIONAMOLECULARMECHANISMOFEARLYTUMORONSETMAYBEDETERMINEDBYCOMPARINGMICROSATELLITEINSTABILITYMSI,INDICATIVEOFERRORPRONEMISMATCHREPAIR,ANDLOSSOFHETEROZYGOSITYLOHBETWEENGASTRICCANCERSINPATIENTSI40YEARSOFAGEANDTHOSEOFOLDERAGESTHREETO5CHROMOSOMALLOCI,WHEREMSIAND/ORLOHARECOMMONLYFOUNDINGASTRICCANCERSINTHEGENERALPOPULATION,WEREEXAMINEDINFORMALINFIXED,PARAFFINEMBEDDEDSAMPLESFROM102PATIENTSI40YEARSOFAGEUSINGAPOLYMERASECHAINREACTIONBASEDNONRADIOACTIVESCREENINGMETHODMSIAND/ORLOHATAMINIMUMOF1LOCUSWEREDETECTEDIN11/102PATIENTSTHEFREQUENCYOFMSIAND/ORLOHATTHED11S904LOCUSWASSIGNIFICANTLYHIGHERTHANTHATATTHED2S119,D2S123,D5S409ANDIFNAREGIONSNOPREFERENTIALGENETICCHANGESATTHED11S904LOCUSWEREOBSERVEDINELDERLYPATIENTSAMONGSEVERALCLINICOPATHOLOGICALVARIABLES,ASTATISTICALLYSIGNIFICANTASSOCIATIONWITHMSIAND/ORLOHWASOBSERVEDONLYFORTUMORSLOCATEDATTHECARDIA,COMPAREDWITHTUMORSATTHEANTRUMANDTHECORPUSOURFINDINGSSUGGESTTHATAUNIQUEMECHANISMMAYBEINVOLVEDININCREASINGTHESUSCEPTIBILITYOFTHED11S904LOCUSFOREITHERMSIORLOH,ESPECIALLYFORCARDIATUMORSINYOUNGPATIENTSEARLYONSETOFGASTRICCANCERSINPATIENTSI40YEARSOFAGEISASSOCIATEDWITHGENETICCHANGESATPREFERENTIALCHROMOSOMALLOCI,INCLUDINGD11S904INTJCANCER8259–62,1999?1999WILEYLISS,INCALTHOUGHGASTRICCANCERINCIDENCEHASBEENDECLINING,ITWASSTILLRANKEDASTHE2NDMOSTCOMMONCANCERANDTHE2NDLEADINGCAUSEOFCANCERDEATHINTHEWORLDINTHE1990SPARKINETAL,1993BECAUSEOFTHEAGGRESSIVENATUREOFGASTRICCANCERS,THEOVERALL5YEARSURVIVALRATEISLESSTHAN20BREAUXETAL,1990GASTRICCANCERSUSUALLYOCCURINPATIENTSOLDERTHAN50YEARSOFAGEONLYABOUT5OFGASTRICCANCERPATIENTSAREYOUNGERTHAN40YEARSNEUGUTETAL,1996GASTRICCANCERSINPATIENTS?40YEARSOFAGEAREMOREAGGRESSIVETHANTHOSEINELDERLYPATIENTSFUJIMOTOETAL,1994COMPARISONOFGENETICALTERATIONSINGASTRICCANCERSBETWEENPATIENTS?40YEARSOFAGEANDOLDERPATIENTSMAYIDENTIFYASPECIFICMOLECULARMECHANISMASSOCIATEDWITHANEARLYONSETOFTUMORMICROSATELLITESARESHORTTANDEMLYREPEATEDDNASEQUENCESANDPRESENTTHROUGHOUTMAMMALIANGENOMESREPETITIVESEQUENCESOFDI,TRIANDTETRANUCLEOTIDEREPEATSAREPRESENTINMORETHAN50OFTHEHUMANGENOMEBECKMANANDWEBER,1992THETANDEMREPEATSOFDCDANAREPARTICULARLYABUNDANTALTERATIONSINTHENUMBEROFREPEATSPERSITE,KNOWNASMICROSATELLITEINSTABILITYMSI,HAVEBEENIMPLICATEDINVARIOUSHUMANDISEASES,INCLUDINGNEOPLASMSSPEICHER,1995MICROSATELLITESEQUENCESAREALSOHIGHLYPOLYMORPHICPOLYMORPHISMDUETODIFFERENTNUMBEROFREPEATSISVERYUSEFULTODETERMINEGENETICALTERATIONS,SUCHASLOSSOFHETEROZYGOSITYLOHINTHISSTUDY,WEEXAMINED5CHROMOSOMALLOCI,WHEREMSIAND/ORLOHARECOMMONLYFOUNDINGASTRICCANCERSINTHEGENERALPOPULATIONRHYUETAL,1994CHONGETAL,1994NAGELETAL,1995TAMURAETAL,1995LINETAL,1995SERUCAETAL,1995BUONSANTIETAL,1997,INGASTRICCANCERPATIENTS?40YEARSOFAGETHEFREQUENCYOFMSIAND/ORLOHANDTHEIRASSOCIATIONWITHOTHERCLINICOPATHOLOGICALDATAAREDISCUSSEDMATERIALANDMETHODSPATIENTSFORMALINFIXED,PARAFFINEMBEDDEDTISSUEBLOCKSFROM102PATIENTS?40YEARSOFAGEMEANAGE35YEARSRANGE16–40YEARSWERERETRIEVEDFROMTHEARCHIVESOFMANYITALIANHOSPITALSWITHCOMPARABLEGASTRICCANCERINCIDENCEDEMOGRAPHICANDPATHOLOGICALDATA,INCLUDINGAGE,GENDER,SITEOFTUMORANDTUMORSTAGEUSINGTHETNMSYSTEMBEAHRSANDMYERS,1983,WEREOBTAINEDFROMACOLLABORATIVEMULTICENTERSTUDYTHESITEOFTUMORWASCATEGORIZEDINTOTHEANTRUM,CORPUSORCARDIAOFSTOMACHNOCANCEREXTENDEDBEYONDTHEGASTRICESOPHAGEALJUNCTIONACCORDINGTOTHELAURE′NSYSTEM1965,GASTRICCANCERSWERECLASSIFIEDASEITHERINTESTINALORDIFFUSETYPEWHENBOTHPHENOTYPESCOEXISTED,CLASSIFICATIONOFTHETUMORTYPEWASBASEDONTHEMOSTREPRESENTATIVEHISTOLOGYGASTRITISNONATROPHICVSATROPHIC/METAPLASTICWASCLASSIFIEDUSINGTHEHOUSTONUPDATEDSYDNEYSYSTEMDIXONETAL,1996ALLCASESWEREJOINTLYASSESSEDBY2AUTHORSMCANDMRDNAEXTRACTIONNEOPLASTICANDADJACENTNONNEOPLASTICAREASWEREMICRODISSECTEDSEPARATELYFROMUNSTAINEDFORMALINFIXED,PARAFFINEMBEDDEDSECTIONSFORDEPARAFFINIZATION,PROTEINASEKDIGESTIONANDDNAPURIFICATION,ASDESCRIBEDPREVIOUSLYSHIAOETAL,1994MICRODISSECTEDNEOPLASTICAREASCONTAINEDATLEAST50TUMORCELLSWHENPOSSIBLE,LYMPHOCYTESWEREUSEDASANONNEOPLASTICCELLCONTROLPOLYMERASECHAINREACTIONPCRFIVECHROMOSOMALLOCIWITHCANDINUCLEOTIDEREPEATSD2S119,D2S123,D5S409,IFNAANDD11S904WERESELECTEDFORPCRPRIMERSD2S123AFM093XH3AANDAFM093XH3MD5S409AFM184YB6AANDAFM184YB6MIFNAIFNAPCR21ANDIFNAPCR22D11S904AFM081ZA5AANDAFM081ZA5MWEREOBTAINEDFROMTHEGENOMEDATABASEWEBSITEWWWGDBORGFORD2S119,UPPER5?CCAGTTTGGAAGCATTT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