Enkosi ngokundwendwela www.chinacdoe.com.Inguqulelo yesikhangeli oyisebenzisayo inenkxaso enyiniweyo yeCSS.Ngowona mava angcono, sicebisa ukuba usebenzise isikhangeli esihlaziyiweyo (okanye uvale iMowudi yokuThelela kwi-Internet Explorer).Okwangoku, ukuqinisekisa inkxaso eqhubekayo, siya kunika isayithi ngaphandle kwezitayela kunye neJavaScript.
Iinkqubo zeLab-on-a-chip ezinesakhono sendawo zinika amandla okuxilongwa ngokukhawuleza kunye nokuchanekileyo kwaye ziluncedo kwiindawo ezinobuncwane apho izixhobo ze-biomedical kunye neengcali eziqeqeshiweyo zingekho.Nangona kunjalo, ukudala inkqubo yokuvavanya i-point-care-care kunye nayo yonke into efunekayo kwi-multi-functional dispensing, ukukhutshwa kwemfuno, ukusebenza okuthembekileyo, kunye nokugcinwa kwexesha elide le-reagents kuhlala kungumngeni omkhulu.Apha sichaza iteknoloji yokutshintsha i-lever-actuated micro ehambayo ekwazi ukulawula i-fluids nakweyiphi na indlela, inike impendulo echanekileyo kunye neyomlinganiselo kuxinzelelo lomoya olusetyenzisiweyo, kwaye uhlale uzinzile ngokuchasene nokunyakaza ngokukhawuleza kunye nokunyakaza.Ngokusekwe kwitekhnoloji, sikwachaza uphuhliso lwenkqubo ye-polymerase chain reaction edibanisa intshayelelo ye-reagent, ukuxuba kunye nemisebenzi yokuphendula yonke into kwinkqubo enye, eyenza "isampuli-empendulo-ngaphandle" ukusebenza kuzo zonke iisampuli zempumlo ezivela kwizigulane ze-18 Umkhuhlane kunye nolawulo lwe-18 lomntu ngamnye, kwi-concordance efanelekileyo ye-fluorescence intensity kunye ne-standard polymerase chain reaction (i-Pearson coefficients> 0.9).Ngokusekwe kwitekhnoloji, sikwachaza uphuhliso lwenkqubo ye-polymerase chain reaction edibanisa intshayelelo ye-reagent, ukuxuba kunye nemisebenzi yokuphendula yonke into kwinkqubo enye, eyenza "isampuli-empendulo-ngaphandle" ukusebenza kuzo zonke iisampulu zempumlo ezivela kwizigulane ze-18. ngoMkhuhlane kunye ne-18 yolawulo lomntu ngamnye, kwi-concordance elungileyo ye-fluorescence intensity kunye ne-standard polymerase chain reaction (i-Pearson coefficients> 0.9).Основываясь на этой технологии, мы также описываем разработку системы полимеразной цепной реакции, которая объединяет фунцияет функции, которая объединяет фунциваиваиваивтова реакции в одном процессе, что обеспечивает выполнение «образец-в-ответ-выход» kunye ne-18 отдельных контролей, в хорошем соответствии интенсивности флуоресценции стандартной полимеразной цепной реакцией (Пиэф,9)Ngokusekelwe kule teknoloji, sichaza ukuphuhliswa kwenkqubo ye-polymerase chain reaction system edibanisa imisebenzi yokutofa, ukuxuba, kunye nokusabela kwinkqubo enye, eyenza ukuba isampuli-impendulo iphume kuzo zonke iisampuli zeklinikhi zeempumlo ezivela kwizigulane zomkhuhlane we-18.kunye ne-18 yolawulo lomntu ngamnye, ngokuvumelana kakuhle kunye ne-standard polymerase chain reaction fluorescence intensity (i-Pearson's coefficients> 0.9).Ngokusekwe kobu buchwepheshe, sikwachaza uphuhliso lwenkqubo ye-polymerase chain reaction edibanisa inaliti ye-reagent, ukuxuba, kunye nemisebenzi yokuphendula ukuhlalutya yonke imizekelo yeempumlo zeklinikhi ukusuka kwi-18 kwisampulu ye-sample yezigulane zempumlo. kakuhle ne-standard polymerase chain reaction (i-coefficient kaPearson> 0.9).Iqonga elicetywayo liqinisekisa i-automation ethembekileyo yohlalutyo lwe-biomedical kwaye ngoko inokukhawulezisa ukuthengiswa koluhlu lwezixhobo zokuvavanya i-point-of-care-care.
Izifo zabantu ezivelayo, ezinje ngobhubhani we-COVID-19 ka-2020 osube ubomi bezigidi zabantu, zibeka isoyikiso esikhulu kwimpilo yehlabathi nakwimpucuko yabantu1.Ukufumanisa kwangaphambili, ngokukhawuleza kunye ngokuchanekileyo kwezifo kubalulekile ukulawula ukusasazeka kwentsholongwane kunye nokuphucula iziphumo zonyango.Inkqubo engundoqo yoxilongo lwendalo esekwe kwiilebhu ezikwindawo esembindini apho iisampulu zovavanyo zithunyelwa ezibhedlele okanye kwiiklinikhi zoxilongo kwaye ziqhutywa ziingcali ngoku ithintela ukufikelela kubantu abaphantse babe zibhiliyoni ezi-5.8 kwihlabathi liphela, ngakumbi abo bahlala kwiindawo ezinobuncwane.apho kukho ukunqongophala kwezixhobo ze-biomedical ezibizayo kunye neengcali eziqeqeshiweyo.Iiklinikhi 2. Ngoko ke, kukho imfuneko engxamisekileyo yokuphuhlisa inkqubo ye-lab-on-a-chip engabizi kwaye isebenziseka lula kunye novavanyo lwe-point-of-care (POCT) enokubonelela oogqirha ngolwazi lokuxilonga kwangexesha ukwenza izigqibo zoxilongo olunolwazi. .kunye nonyango 3.
Izikhokelo zoMbutho wezeMpilo weHlabathi (i-WHO) zichaza ukuba i-POCT efanelekileyo kufuneka ifikeleleke, isebenziseke lula (kulula ukuyisebenzisa ngoqeqesho oluncinci), ichaneke (ukuphepha ukunganyaniseki okungeyonyani okanye ukuchaneka okungeyonyani), ikhawuleze kwaye ithembekile (ibonelele ngeempawu ezintle zokuphindaphinda), kwaye iyahanjiswa (ekwaziyo ukugcina ixesha elide kwaye ifumaneke lula kubasebenzisi bokuphela)4.Ukuhlangabezana nezi mfuno, iinkqubo ze-POCT kufuneka zibonelele ngezi mpawu zilandelayo: idosi eguquguqukayo yokunciphisa ungenelelo olwenziwa ngesandla, ukukhutshwa kwemfuno yokukala uthutho lwe-reagent ukuze ufumane iziphumo ezichanekileyo zovavanyo, kunye nokusebenza okuthembekileyo ukumelana nokungcangcazela kokusingqongileyo.Okwangoku, esona sixhobo sisetyenziswa kakhulu se-POCT yi-lateral flow strip5,6 equlathe iileya ezininzi ze-porous nitrocellulose membranes ezityhala isampulu encinci kakhulu phambili, isabela nge-pre-immobilized reagents ngamandla e-capillary.Nangona banenzuzo yeendleko eziphantsi, ukusetyenziswa lula, kunye neziphumo ezikhawulezayo, izixhobo ze-POCT ezisekelwe kwi-flow strip zingasetyenziselwa kuphela iimvavanyo zebhayoloji (umzekelo, iimvavanyo ze-glucose7,8 kunye nokukhulelwa kwe-9,10) ngaphandle kokufuna uhlalutyo lwezigaba ezininzi.ukusabela (umzekelo ukulayishwa kwee-reagents ezininzi, ukuxuba, ukuphindaphinda).Ukongezelela, amandla okuqhuba alawula ukunyakaza kwamanzi (oko kukuthi, amandla e-capillary) awaboneleli ngokuhambelana kakuhle, ngokukodwa phakathi kweebhetshi, okubangelwa ukuveliswa kakubi kwe-11 kunye nokwenza iibhendi zokuhamba kwe-lateral zibe luncedo kakhulu ekufumaneni okulungileyo12,13.
Ukwandiswa kwamandla okuvelisa kwi-micro- kunye ne-nanoscale yenze amathuba okuphuhliswa kwezixhobo ze-POCT ze-microfluidic kwimilinganiselo yobuninzi14,15,16,17.Ngokulungelelanisa iimpawu ze-interface ye-18, i-19 kunye nejometri yeendlela ze-20, i-21, i-22, i-capillary force kunye nesantya sokuhamba kwezi zixhobo zingalawulwa.Nangona kunjalo, ukuthembeka kwazo, ngakumbi kulwelo olumanzi kakhulu, kuhlala kungamkelekanga ngenxa yokungachaneki kwemveliso, iziphene zezinto, kunye nobuntununtunu bokungcangcazela kokusingqongileyo.Ukongezelela, ekubeni ukuhamba kwe-capillary kudalwe kwi-interface ye-liquid-gas, akukho kuhamba okongeziweyo kunokungeniswa, ngakumbi emva kokuzalisa i-microfluidic channel nge-liquid.Ke ngoko, ukufunyaniswa okuntsonkothileyo, amanyathelo amaninzi esitofu sesampulu kufuneka enziwe24,25.
Phakathi kwezixhobo ze-microfluidic, izixhobo ze-centrifugal microfluidic okwangoku sesinye sezisombululo ezilungileyo ze-POCT26,27.Isixhobo sayo sokuqhuba sinenzuzo kuba amandla okuqhuba anokulawulwa ngokulungelelanisa isantya sokujikeleza.Nangona kunjalo, into engalunganga kukuba amandla e-centrifugal ahlala ejoliswe kumda wangaphandle wesixhobo, okwenza kube nzima ukuphumeza ukusabela kwamanyathelo amaninzi afunekayo kuhlalutyo olunzima ngakumbi.Nangona amandla okuqhubela phambili ongezelelweyo (umz. ii-capillaries 28, 29 kunye nabanye abaninzi 30, 31, 32, 33, 34, 35) ukongeza kumandla we-centrifugal aziswa kwidosi yemisebenzi emininzi, ukuhanjiswa kolwelo okungalindelekanga kusengenzeka ngenxa yokuba la mandla ongeziweyo ayimiyalelo ngokubanzi. yobukhulu obungaphantsi kumandla we-centrifugal, ibenza ukuba basebenze kuphela kuluhlu oluncinci lokusebenza okanye ukungafumaneki kwimfuno kunye nokukhululwa kolwelo.Ukubandakanya ukuguqulwa kwe-pneumatic kwi-microfluidics ye-centrifugal njengeendlela ze-centrifugal kinetic 36, i-37, i-38, iindlela ze-thermopneumatic 39 kunye neendlela ezisebenzayo ze-pneumatic 40 ibonakalise ukuba yinto ekhangayo.Ngendlela ye-counterfugodynamic, i-cavity eyongezelelweyo kunye ne-microchannels edibeneyo idibaniswe kwisixhobo sokusebenza kwangaphandle nangaphakathi, nangona ukusebenza kwayo kokupompa (kuluhlu olusuka kwi-75% ukuya kwi-90%) kuxhomekeke kakhulu kwinani lemijikelezo yokupompa kunye ne-viscosity. yolwelo.Kwindlela ye-thermopneumatic, i-membrane ye-latex kunye negumbi lokuhambisa ulwelo lwenzelwe ngokukodwa ukutywina okanye ukuphinda uvule i-inlet xa umthamo womoya ovalelweyo ushushu okanye upholile.Nangona kunjalo, ukuseta ukufudumeza / ukupholisa kwazisa iingxaki zokuphendula ngokukhawuleza kwaye kunqande ukusetyenziswa kwayo kwiimvavanyo ze-thermosensitive (umzekelo, i-polymerase chain reaction (PCR) ukukhulisa).Ngendlela esebenzayo ye-pneumatic, ukukhutshwa kwemfuno kunye nokunyakaza kwangaphakathi kufezekiswa ngokusetyenziswa kwangaxeshanye koxinzelelo olulungileyo kunye nokuchaneka ngokuchanekileyo kwezantya zokujikeleza ngeenjini zesantya esiphezulu.Kukho ezinye iindlela eziphumelelayo zisebenzisa ii-activators ze-pneumatic kuphela (uxinzelelo olulungileyo 41, 42 okanye uxinzelelo olubi 43) kunye noyilo lwevalvu evalekileyo.Ngokufaka uxinzelelo ngokulandelelana kwigumbi le-pneumatic, ulwelo lumpontshelwa phambili ngeperistaltically, kwaye ivalve eqhelekileyo ivaliwe ikhusela umva kolwelo ngenxa ye-peristalsis, ngaloo ndlela iqonda imisebenzi yolwelo entsonkothileyo.Nangona kunjalo, okwangoku kukho kuphela inani elilinganiselweyo letekhnoloji ye-microfluidic enokwenza imisebenzi enzima yolwelo kwisixhobo esinye se-POCT, kubandakanya ukuhanjiswa kwezinto ezininzi, ukukhutshwa kwemfuno, ukusebenza okuthembekileyo, ukugcinwa kwexesha elide, ukuphatha ulwelo oluphezulu lwe-viscosity, kunye nemveliso engabizi kakhulu.Konke ngaxeshanye.Ukungabikho komsebenzi owenziwe ngamanyathelo amaninzi kunokuba sesinye sezizathu zokuba kutheni kuphela iimveliso ze-POCT ezimbalwa zorhwebo ezifana ne-Cepheid, i-Binx, i-Visby, i-Cobas Liat, kunye ne-Rhonda ziye zaziswa ngempumelelo kwimarike evulekileyo ukuza kuthi ga ngoku.
Kweli phepha, siphakamisa i-actuator ye-pneumatic microfluidic esekelwe kwi-green ring micro switch technology (FAST).I-FAST idibanisa zonke iipropati eziyimfuneko ngexesha elifanayo kwi-reagents ebanzi ukusuka kwi-microliters ukuya kwi-milliliters.I-FAST iqulethe ii-elastic membranes, iilevers kunye neebhloko.Ngaphandle kokusetyenziswa koxinzelelo lomoya, i-membrane, i-levers kunye neebhloko zingavalwa ngokuqinileyo kwaye i-liquid ngaphakathi ingagcinwa ixesha elide.Xa uxinzelelo olufanelekileyo lusetyenziswa kwaye lulungelelaniswa nobude be-lever, i-diaphragm iyanda kwaye iqhube i-lever kwindawo evulekile, ivumela ulwelo ukuba ludlule.Oku kuvumela ukubalwa kolwelo olunemisebenzi emininzi kwi-cascade, ngaxeshanye, ngolandelelwano okanye ngendlela ekhethiweyo.
Siye saqulunqa inkqubo ye-PCR usebenzisa i-FAST ukuvelisa iziphumo ze-supply-in-sample ukuze kubonwe i-virus ye-influenza A kunye ne-B (IAV kunye ne-IBV).Sifumene umda ophantsi wokufumanisa (LOD) yeekopi ze-102 / mL, uvavanyo lwethu oluphindaphindiweyo lubonise ukuchaneka kwe-IAV kunye ne-IBV kwaye yavumela i-virus ye-influenza pathotyping.Iziphumo zovavanyo lweklinikhi kusetyenziswa isampulu ye-swab yeempumlo ezivela kwizigulane ezili-18 kunye nabantu abasempilweni abali-18 babonisa i-concordance elungileyo kwi-fluorescence intensity kunye ne-RT-PCR eqhelekileyo (i-Pearson coefficients> 0.9).Iziphumo zovavanyo lweklinikhi kusetyenziswa isampulu ye-swab yeempumlo ezivela kwizigulane ezili-18 kunye nabantu abasempilweni abali-18 babonisa i-concordance elungileyo kwi-fluorescence intensity kunye ne-RT-PCR eqhelekileyo (i-Pearson coefficients> 0.9).Результаты клинических испытаний с использованием образца мазка из носа от 18 пациентов и 18 здоровых лиц показывает хостовыт собошеные сположение сценции стандартной ОТ-ПЦР (коэфффициенты Пирсона > 0,9).Iziphumo zolingo lwezonyango kusetyenziswa isampulu yempumlo yokuswabha kwizigulane ezili-18 kunye nabantu abasempilweni abali-18 babonisa isivumelwano esihle phakathi kokuqina kwe-fluorescence ye-RT-PCR eqhelekileyo (i-Pearson's coefficients> 0.9).0.9 ............ Результаты клинических испытаний с использованием образцов назальных мазков от 18 пациентов и 18 здоровых лиц показать достовиный сположение ю флуоресценции и стандартной ОТ-ПЦР (коэфффициент Пирсона > 0,9).Iziphumo zolingo lwezonyango kusetyenziswa iisampulu ze-swab yeempumlo ezivela kwizigulane ezili-18 kunye nabantu abasempilweni abali-18 babonise isivumelwano esihle phakathi kokuqina kwe-fluorescence kunye ne-RT-PCR eqhelekileyo (i-Coefficient kaPearson> 0.9).Iindleko eziqikelelwayo zesixhobo se-FAST-POCT zimalunga ne-US$1 (iTheyibhile eyoNgezelelweyo 1) kwaye inokwehliswa ngakumbi ngokusebenzisa iindlela zokwenziwa komgangatho omkhulu (umz. ukubumba inaliti).Ngapha koko, izixhobo ze-POCT ezisekelwe kwi-FAST zinazo zonke iimpawu eziyimfuneko ezigunyaziswe yi-WHO kwaye ziyahambelana neendlela ezintsha zovavanyo lwe-biochemical ezifana ne-plasma thermal cycling44, i-amplification-free immunoassays45 kunye ne-nanobody functionalization tests46 eziyintsika yeenkqubo ze-POCT.kunokwenzeka.
Kwikhiwane.I-1a ibonisa isakhiwo se-platform ye-FAST-POCT, equkethe amagumbi amane olwelo: i-pre-storage room, i-mixing room, i-reaction room, kunye ne-waste room.Isitshixo sokulawula ukuhamba kwamanzi yi-FAST yoyilo (ebandakanya i-elastic membranes, i-levers kunye neebhloko) ezibekwe kwigumbi langaphambili lokugcina kunye negumbi lokuxuba.Njengendlela ye-pneumatically actuated, uyilo lwe-FAST lubonelela ngolawulo oluchanekileyo lokuhamba kwamanzi, kubandakanywa ukutshintshwa okuvaliweyo / okuvulekileyo, i-dosing eguquguqukayo, ukukhutshwa kolwelo olufunwayo, ukusebenza okuthembekileyo (umzekelo, ukungabonakali kwi-vibration yendalo), kunye nokugcinwa kwexesha elide.I-platform ye-FAST-POCT iqukethe iileyile ezine: i-backing layer, i-elastic film layer, i-plastic film layer, kunye ne-cover cover, njengoko kuboniswe kumbono owandisiweyo kwi-Fig. ).Zonke iziteshi kunye namagumbi okuthuthwa kwamanzi (ezifana nokugcinwa kwangaphambili kunye namagumbi okuphendula) zifakwe kwi-PLA (i-polylactic acid) substrates ukusuka kwi-0.2 mm (inxalenye encinci) ukuya kwi-5 mm ubukhulu.Ifilim eelastiki imathiriyeli yi-PDMS eyi-300 µm etyebileyo eyanda lula xa uxinzelelo lomoya lusetyenziswa ngenxa “yobungqingqwa bayo obuncinane” kunye nemodyuli esezantsi yokunwebeka (malunga ne-2.25 MPa47).Ifilimu yefilimu ye-polyethylene yenziwe nge-polyethylene terephthalate (PET) kunye nobukhulu be-100 µm ukukhusela ifilimu e-elastic kwi-deformation egqithisileyo ngenxa yoxinzelelo lomoya.Ngokuhambelana namagumbi, i-substrate layer inama-levers axhunyiwe kwi-cover layer (eyenziwe nge-PLA) ngeehenjisi zokulawula ukuhamba kwamanzi.Ifilimu e-elastic ifakwe kwi-backing layer isebenzisa i-tape yokubambelela emacala amabini (i-ARseal 90880) kwaye igqunywe ngefilimu yeplastiki.Iingqimba ezintathu zahlanganiswa kwi-substrate kusetyenziswa i-T-clip design kwi-cover layer.I-T-clamp inesithuba phakathi kwemilenze emibini.Xa ikliphu ifakwe kwi-groove, imilenze yomibini yagoba kancinane, emva koko yabuyela kwimeko yayo yangaphambili kwaye ibophe ngokuqinileyo isiciko kunye nokuxhaswa njengoko bedlula kwi-groove (Fig. S1).Amaleko amane ke adityaniswa kusetyenziswa izihlanganisi.
Umzobo weqonga weqonga obonisa amacandelo ahlukeneyo asebenzayo kunye neempawu ze-FAST.b Umzobo owandisiweyo weqonga le-FAST-POCT.c Ifoto yeqonga elisecaleni kwenkozo yekota yedola yaseMelika.
Indlela yokusebenza yeqonga le-FAST-POCT liboniswe kwi-Figure 2. Amacandelo aphambili ziibhloko kuluhlu lwesiseko kunye neehinges kwi-cover layer, ephumela kuyilo lokuphazamiseka xa iileyile ezine zihlanganiswe kusetyenziswa i-T-shape. .Xa kungekho xinzelelo lomoya lusetyenziswayo (umzobo 2a), ukungenelela okuhambelanayo kubangela ukuba ihenjisi igobe kwaye igqwese, kwaye amandla okutywina asetyenziswa kwi-lever ukucinezela ifilimu e-elastic ngokuchasene nebhloko, kwaye ulwelo olukwindawo yokutywina luchazwe. njengelizwe elitywiniweyo.Kufuneka kuqatshelwe ukuba kulo mbuso, i-lever igobe ngaphandle, njengoko kuboniswe kumbono wecala kwi-Fig. 2a.Xa umoya unikezelwa (umzobo 2b), i-elastic membrane ikhula ngaphandle ukuya kwisigqubuthelo kwaye ityhale i-lever phezulu, ngaloo ndlela ivula i-gap phakathi kwe-lever kunye nebhloko ye-fluid ukuya kwigumbi elilandelayo, elichazwa njengelizwe elivulekileyo. .Emva kokukhululwa koxinzelelo lomoya, i-lever ingabuyela kwindawo yayo yokuqala kwaye ihlale iqinile ngenxa yokuqina kwehinge.Iividiyo zeentshukumo ze-lever ziboniswa kwifilimu eyongezelelweyo i-S1.
A. Idayagram ecwangcisiweyo kunye neefoto xa ivaliwe.Ukungabikho koxinzelelo, i-lever icinezela i-membrane ngokuchasene nebhloko, kwaye i-liquid ivalwe.b Ikwimeko entle.Xa uxinzelelo lusetyenziswa, i-membrane iyanda kwaye ityhale i-lever phezulu, ngoko ke umjelo uvula kwaye umbane unokuhamba.c Qinisekisa ubungakanani beempawu zoxinzelelo olubalulekileyo.Iimpawu zeempawu ziquka ubude be-lever (L), umgama phakathi kwe-slider kunye nehinge (l) kunye nobukhulu be-lever protrusion (t).I-Fs yi-compact force force at throttle point B. q ngumthwalo owabiwa ngokufanayo kwi-lever.I-Tx* imele itorque ephuhliswe sisilevi esinehenjisi.Uxinzelelo olubalulekileyo luxinzelelo olufunekayo ukuphakamisa i-lever kunye nokwenza ukuhamba kwamanzi.d Iziphumo zethiyori kunye novavanyo lobudlelwane phakathi koxinzelelo olubalulekileyo kunye nobukhulu bento.n = Imifuniselo emi-6 ezimeleyo yenziwa kwaye idatha ibonisiwe njengokutenxa okusemgangathweni.Idatha ekrwada inikezelwa njengeefayili zedatha ekrwada.
Imodeli yohlalutyo esekelwe kwi-theory ye-beam iye yaphuhliswa ukuhlalutya ukuxhomekeka koxinzelelo olunzima lwe-Pc apho i-gap ivula i-geometric parameters (umzekelo, uL ubude be-lever, l ngumgama phakathi kwebhloko kunye i-hinge, i-S yi-lever Indawo yoqhagamshelwano kunye ne-liquid t ubukhulu be-lever protrusion, njengoko kuboniswe kwi-Fig. 2c).Njengoko kucacisiwe kwiNqaku lezoNgeniso kunye neFigure S3 eyoNgezelelweyo, isithuba sivula xa \({P}_{c}\ge \ frac{2{F}_{s}l}{SL}\), apho iFs iyitorque. \ ({T}_{x}^{\ast}(={F}_{s}l)\) ukuphelisa imikhosi ehambelana nokuphazamiseka kokuphazamiseka kunye nokubangela ukuba ihenjisi igobe.Impendulo yovavanyo kunye nemodeli yokuhlalutya ibonisa isivumelwano esihle (umzobo 2d), ebonisa ukuba uxinzelelo olubalulekileyo lwe-Pc luyanda ngokunyuka kwe-t / l kunye nokunciphisa i-L, echazwe ngokulula yimodeli ye-beam ye-classical, oko kukuthi i-torque yanda nge-t / Lift. .Ngaloo ndlela, uhlalutyo lwethu lwethiyori lubonisa ngokucacileyo ukuba uxinzelelo olubalulekileyo lunokulawulwa ngokufanelekileyo ngokulungelelanisa ubude be-lever L kunye nomlinganiselo we-t / l, obonelela ngesiseko esibalulekileyo soyilo lweqonga le-FAST-POCT.
Iqonga le-FAST-POCT libonelela ngokusasazwa kwemisebenzi emininzi (eboniswe kwi-Figure 3a kunye ne-inset kunye novavanyo), eyona nto ibaluleke kakhulu kwi-POCT eyimpumelelo, apho ulwelo lunokuhamba kuyo nayiphi na indlela kunye naluphi na umyalelo (i-cascade, ngexesha elinye, ngokulandelelana) okanye i-multichannel ekhethiweyo. ukuhambisa .– umsebenzi wethamo.Kwikhiwane.I-3a(i) ibonisa imowudi ye-cascade dosing apho amagumbi amabini okanye ngaphezulu ahlanjululwayo kusetyenziswa iibhloko ukwahlula ii-reactants ezahlukeneyo kunye ne-lever yokulawula iindawo ezivulekileyo nezivaliweyo.Xa uxinzelelo lusetyenziswa, ulwelo luhamba ukusuka phezulu ukuya kwigumbi elisezantsi ngendlela ye-cascade.Kufuneka kuqatshelwe ukuba amagumbi e-cascade angazaliswa ngamachiza amanzi okanye iikhemikhali ezomileyo ezifana ne-lyophilized powders.Kuvavanyo olukumfanekiso 3a (i), i-inki ebomvu esuka kwigumbi eliphezulu ihamba kunye nomgubo wedayi eluhlaza okwesibhakabhaka (i-sulfate yethusi) ukuya kwigumbi lesibini kwaye ijika luhlaza okwesibhakabhaka xa ifika kwigumbi elisezantsi.Ikwabonisa noxinzelelo lolawulo lokumpompa ulwelo.Ngokufanayo, xa i-lever enye idityaniswe kumagumbi amabini, iba yimowudi yokujova ngaxeshanye, njengoko kubonisiwe kumkhiwane.3a(ii), apho ulwelo lunokusasazwa ngokufanayo phezu kwamagumbi amabini okanye ngaphezulu xa uxinzelelo lusetyenziswa.Ekubeni uxinzelelo olubalulekileyo luxhomekeke kubude be-lever, ubude be-lever bunokutshintshwa ukuze kufezekiswe iphethini yokujova ngokulandelelana njengoko kuboniswe kumkhiwane.3a(iii).I-lever ende (enoxinzelelo olunzima lwe-Pc_long) idibaniswe negumbi B kunye ne-lever emfutshane (enoxinzelelo olunzima i-Pc_short> i-Pc_long) idibaniswe ne-chamber A. Njengoko uxinzelelo lwe-P1 (Pc_long <P1 <Pc_short) lusetyenziswe, kuphela i-liquid ebomvu ebomvu. inokuqukuqela ukuya kwigumbi B kwaye xa uxinzelelo lonyusiwe ukuya kwi-P2 (> Pc_mfutshane), ulwelo oluzuba lunokugeleza ukuya kwigumbi A. Le modi yokutofa ngokulandelelana isebenza kulwelo olwahlukeneyo olutshintshela kumagumbi anxulumeneyo ngokulandelelana, okubaluleke kakhulu kwi-POCT eyimpumelelo. isixhobo.I-lever ende (enoxinzelelo olunzima lwe-Pc_long) idibaniswe negumbi B kunye ne-lever emfutshane (enoxinzelelo olunzima i-Pc_short> i-Pc_long) idibaniswe ne-chamber A. Njengoko uxinzelelo lwe-P1 (Pc_long
a Umzekeliso wezabelo ezinemisebenzi emininzi, okt (i) i-cascading, (ii) ngaxeshanye, (iii) ngokulandelelana, kunye (iv) nesabelo esikhethiweyo.Iigophe zimela ukuhamba komsebenzi kunye neeparitha zezi ndlela zokuhambisa ezine.b Iziphumo zeemvavanyo zokugcinwa kwexesha elide kumanzi adibeneyo kunye ne-ethanol.n = Imifuniselo emi-5 ezimeleyo yenziwa kwaye idatha iboniswa njenge ± sd c.Imiboniso yovavanyo lozinzo xa isixhobo se-FAST kunye ne-capillary valve (CV) isixhobo sasikwi-(i) static kunye (ii) nezimo zokungcangcazela.(iii) Umthamo kunye nexesha le-FAST kunye nezixhobo ze-CV kwiifrikhwensi ezahlukeneyo ze-angular.d Ukupapashwa kweziphumo zovavanyo ngokwemfuno (i) yesixhobo se-FAST kunye (ii) nesixhobo se-CV.(iii) Ubudlelwane phakathi kwevolumu kunye nexesha le-FAST kunye ne-CV izixhobo usebenzisa imodi yoxinzelelo oluphakathi.Zonke iibar zesikali, 1 cm.Idatha ekrwada inikezelwa njengeefayili zedatha ekrwada.
Ukugcinwa kwexesha elide le-reagents yenye into ebalulekileyo yesixhobo se-POCT esiphumelelayo esiza kuvumela abasebenzi abangaqeqeshwanga ukuba baphathe ama-reagents amaninzi.Ngelixa iitekhnoloji ezininzi zibonise amandla azo okugcina ixesha elide (umzekelo, ii-microdispensers ezingama-35, iipakethe ezingama-48 ze-blister, kunye neepakethe zentonga ezingama-49), indawo enikezelweyo yokufumana iyadingeka ukulungiselela ipakethe, eyandisa iindleko kunye nobunzima;ngaphezu koko, ezi ndlela zogcino azikuvumeli ukukhutshwa okufunwayo kwaye kukhokelela ekonakaleni kweerejenti ngenxa yokushiyeka kwipakethe.Ikhono lokugcina ixesha elide laqinisekiswa ngokuqhuba uvavanyo olukhawulezayo lobomi usebenzisa i-CNC-machined PMMA material ngenxa yobunzima obuncinci kunye nokuchasana nokungena kwegesi (i-Supplementary Figure S5).Izixhobo zokuvavanya zazaliswa ngamanzi adibeneyo (amanzi adibeneyo) kunye ne-70% ye-ethanol (ukulinganisa i-reagents eguqukayo) kwi-65 ° C kwiintsuku ze-9.Zombini amanzi e-deionized kunye ne-ethanol zigcinwe kusetyenziswa i-aluminium foil ukuvala ukufikelela ukusuka phezulu.I-equation ye-Arrhenius kunye ne-activation yamandla yokungena echazwe kwiincwadi ze-50,51 zisetyenziselwa ukubala ukulingana kwexesha langempela.Kwikhiwane.I-3b ibonisa iziphumo zokulahlekelwa kwesisindo somndilili kwiisampuli ze-5 ezigcinwe kwi-65 ° C kwiintsuku ze-9, ezilingana ne-0.30% yamanzi adibeneyo kunye ne-0.72% ye-70% ye-ethanol kwiminyaka eyi-2 kwi-23 ° C.
Kwikhiwane.I-3c ibonisa uvavanyo lokungcangcazela.Ukusukela ukuba i-capillary valve (CV) iyeyona ndlela idumileyo yokuphatha ulwelo phakathi kwezixhobo ezikhoyo ze-POCT28,29, isixhobo se-CV esiyi-300 µm ububanzi kunye ne-200 µm ubunzulu sasetyenziswa ukuthelekisa.Kuyabonakala ukuba xa zombini izixhobo zihlala zimile, ulwelo olukwiqonga leqonga le-FAST-POCT kunye nolwelo olukwisixhobo se-CV lutshixa ngenxa yokwandiswa ngokukhawuleza komjelo, okunciphisa amandla e-capillary.Nangona kunjalo, njengoko i-orbital vibrator ye-angular frequency inyuka, ulwelo kwiqonga le-FAST-POCT lihlala litywiniwe, kodwa ulwelo olukwisixhobo se-CV luqukuqela ukuya kwigumbi elisezantsi (jonga kwakhona iMovie eyoNgezelelweyo ye-S3).Oku kuphakamisa ukuba i-hinges deformable yeqonga le-FAST-POCT lingasebenzisa amandla anamandla omatshini kwimodyuli ukuvala ngokuqinileyo ulwelo kwigumbi.Nangona kunjalo, kwizixhobo ze-CV, i-liquid igcinwa ngenxa yokulinganisela phakathi kwezigaba eziqinileyo, zomoya, kunye ne-liquid, ukudala ukungazinzi, kunye ne-vibrations kunokuphazamisa ibhalansi kwaye kubangele ukuziphatha okungalindelekanga.Inzuzo yeqonga le-FAST-POCT kukuba libonelela ngokusebenza okuthembekileyo kwaye ligwema ukungaphumeleli phambi kokunyakaza okuqhelekileyo okwenzeka ngexesha lokunikezelwa kunye nokusebenza.
Olunye uphawu olubalulekileyo lweqonga le-FAST-POCT kukukhutshwa kwayo kwimfuno, eyona mfuneko iphambili yohlalutyo lobungakanani.Kwikhiwane.I-3d ithelekisa ukukhutshwa kwemfuno yeqonga le-FAST-POCT kunye nesixhobo seCV.Ukususela kwikhiwane.I-3d (iii) sibona ukuba isixhobo se-FAST siphendula ngokukhawuleza kwisignali yoxinzelelo.Xa uxinzelelo lusetyenziswe kwi-platform ye-FAST-POCT, i-fluid yahamba, xa uxinzelelo lukhutshwe, ukuhamba ngokukhawuleza kuyeka (Umfanekiso 3d (i)).Esi senzo sinokuchazwa ngokubuyela ngokukhawuleza kwe-elastic yehinge, ecinezela i-lever ngasemva kwibhloko, ukuvala igumbi.Nangona kunjalo, ulwelo lwaqhubeka luhamba kwisixhobo se-CV, ekugqibeleni lukhokelele kumthamo wolwelo obungalindelekanga malunga ne-100 µl emva kokukhutshwa koxinzelelo (Figure 3d(ii) kunye neMovie eyoNgezelelweyo S4).Oku kunokucaciswa ngokunyamalala kwe-capillary pinning effect ekumanziseni ngokupheleleyo kwe-CV emva kokutofa okokuqala.
Ukukwazi ukuphatha ulwelo lokumanzi okwahlukeneyo kunye ne-viscosity kwisixhobo esifanayo kuhlala kungumngeni kwizicelo ze-POCT.Ukumanzisa okungalunganga kunokukhokelela ekuvuzeni okanye kwezinye iindlela zokuhamba ezingalindelekanga kwiitshaneli, kunye nezixhobo ezincedisayo ezifana ne-vortex mixers, i-centrifuges kunye nezihlungi zihlala zifunwa ukulungiselela ulwelo oluphezulu lwe-viscous 52.Sivavanye ubudlelwane phakathi koxinzelelo olubalulekileyo kunye neempawu zolwelo (kunye noluhlu olubanzi lokumanzisa kunye ne-viscosity).Iziphumo ziboniswa kwiThebhile 1 kunye neVidiyo ye-S5.Inokubonwa ukuba ulwelo lokumanzisa okwahlukeneyo kunye ne-viscosity lunokutywinwa kwigumbi, kwaye xa uxinzelelo lusetyenziswa, kwanolwelo olune-viscosity ukuya kuthi ga kwi-5500 cP lunokudluliselwa kwigumbi elikufuphi, okwenza kube lula ukufumanisa iisampuli eziphakamileyo. i-viscosity (oko kukuthi isikhohlela, isampulu ebonakala kakhulu esetyenziselwa ukuxilongwa kwezifo zokuphefumla).
Ngokudibanisa ezi zixhobo zingentla apha ngentla, uluhlu olubanzi lwezixhobo ze-POCT ezisekelwe kwi-FAST zinokuphuhliswa.Umzekelo uboniswe kuMzobo 1. Isityalo siqulethe igumbi lokugcinwa kwangaphambili, igumbi lokuxuba, igumbi lokuphendula kunye negumbi lokulahla.Ii-reagents zinokugcinwa kwigumbi lokugcinwa kwangaphambili ixesha elide kwaye emva koko zikhutshwe kwigumbi lokuxuba.Ngoxinzelelo olufanelekileyo, ii-reactants ezixubileyo zinokugqithiselwa ngokukhethiweyo kwigumbi lenkunkuma okanye kwigumbi lokuphendula.
Ngenxa yokuba ukuchongwa kwe-PCR kungumgangatho wegolide wokufumanisa iintsholongwane ezifana ne-H1N1 kunye ne-COVID-19 kwaye kubandakanya amanyathelo okusabela amaninzi, sisebenzise iqonga le-FAST-POCT lokubhaqwa kwe-PCR njengesicelo.Kwikhiwane.I-4 ibonisa inkqubo yokuvavanya i-PCR usebenzisa iqonga le-FAST-POCT.Okokuqala, i-reagent eluting, i-reagent ye-magnetic microbead, isisombululo sokuhlamba A, kunye nesisombululo sokuhlamba u-W safakwa ngombhobho kumagumbi angaphambili okugcinwa kwe-E, M, W1 kunye ne-W2, ngokulandelanayo.Amanqanaba e-RNA adsorption aboniswe kwifig.I-4a kwaye ilandelayo: (1) xa uxinzelelo lwe-P1 (= 0.26 bar) lusetyenziswa, isampuli ihambela kwigumbi M kwaye ikhutshwe kwigumbi lokuxuba.(2) Uxinzelelo lomoya lwe-P2 (= 0.12 bar) lunikezelwa nge-port A exhunywe kumzantsi wegumbi lokuxuba.Nangona inani leendlela zokuxuba zibonise amandla abo ekuxubeni ulwelo kwiiplatifomu ze-POCT (umzekelo, ukuxuba inyoka 53, ukuxuba okungahleliwe kwe-54 kunye ne-batch mixing 55), ukudibanisa kwabo ukudibanisa kunye nokusebenza kakuhle akukwanelisi.Yamkela indlela yokuxuba i-bubble, apho umoya ungeniswa ngaphantsi kwegumbi lokuxuba ukudala amaqamza kulwelo, emva koko i-vortex enamandla inokufikelela ukuxuba okupheleleyo kwimizuzwana.Imifuniselo yokuxuba i-Bubble yenziwa kwaye iziphumo zinikezelwe kuMfanekiso owongezelelweyo S6.Ingabonwa ukuba xa uxinzelelo lwe-0.10 bar lusetyenziswa, ukuxuba okupheleleyo kuthatha malunga nemizuzwana ye-8.Ngokunyusa uxinzelelo kwi-bar ye-0.20, ukuxuba okupheleleyo kufezekiswa malunga nemizuzwana emi-2.Iindlela zokubala ukusebenza kakuhle kokuxuba zinikezelwa kwicandelo leeNdlela.(3) Sebenzisa imagnethi ye-rubidium ukukhupha amaso, emva koko ucinezele i-P3 (= 0.17 bar) nge-port P ukuhambisa ii-reagents kwigumbi lenkunkuma.Kwikhiwane.I-4b, c ibonisa amanyathelo okuhlamba ukususa ukungcola kwisampuli ngolu hlobo lulandelayo: (1) Isisombululo sokuhlamba A ukusuka kwigumbi le-W1 likhutshwe kwi-pressing mixing room P1.(2) Emva koko yenza inkqubo yokuxuba i-bubble.(3) Isisombululo sokuhlamba A sidluliselwa kwigumbi lolwelo lwenkunkuma, kwaye ii-microbeads kwigumbi lokuxuba zitsalwa ngaphandle ngumagnethi.Ukuhlamba i-W (Umfanekiso 4c) yayifana nokuhlamba i-A (Umfanekiso 4b).Kufuneka kuqatshelwe ukuba inyathelo ngalinye lokuhlamba A kunye ne-W lenziwa kabini.Umzobo 4d ubonisa amanyathelo elution elute RNA ukusuka amaso;i-lution kunye nokuxuba amanyathelo okwazisa afana ne-RNA adsorption kunye namanyathelo okuhlamba achazwe ngasentla.Njengoko i-reagents elution idluliselwa kwigumbi lokuphendula le-PCR phantsi koxinzelelo lwe-P3 kunye ne-P4 (= 0.23 bar), uxinzelelo olubalulekileyo lufikelelwa ukutywina ingalo yegumbi lokuphendula le-PCR.Ngokufanayo, uxinzelelo lwe-P4 lukwanceda ukutywina indlela eya kwigumbi lenkunkuma.Ke, zonke ii-reagents ze-elution zasasazwa ngokulinganayo phakathi kwamagumbi amane okusabela e-PCR ukuqalisa ukuphendula kwe-PCR ephindaphindwayo.Le nkqubo ingentla inikezelwe kwi-Supplementary Movie S6.
Kwinqanaba le-adsorption ye-RNA, isampuli ingeniswe kwi-inlet M kwaye ifakwe kwigumbi lokuxuba kunye nesisombululo se-bead esigcinwe ngaphambili.Emva kokuxuba kunye nokususa i-granules, i-reagents ihanjiswa kwigumbi lenkunkuma.b kunye c namanyathelo okuhlamba, ngenisa iireyinti ezahlukeneyo zokuhlambela ebezigcinwe ngaphambili kwigumbi lokuxuba, kwaye emva kokuxuba kunye nokususa amaso, udlulisele izinto ezisebenza ngenkunkuma kwigumbi lolwelo lwenkunkuma.d Inyathelo le-Elution: Emva kokuzisa ii-reagents elution, ukuxuba kunye nokutsalwa kweentsimbi, ii-reagents zidluliselwa kwigumbi lokuphendula le-PCR.Iigophe zibonisa ukuhamba komsebenzi kunye neeparamitha ezihambelanayo zezigaba ezahlukeneyo.Uxinzelelo luxinzelelo olwenziwa ngamagumbi ngamanye.Umthamo ngumthamo wolwelo kwigumbi lokuxuba.Zonke iibar zesikali ziyi-1 cm.Idatha ekrwada inikezelwa njengeefayili zedatha ekrwada.
Inkqubo yovavanyo lwe-PCR yenziwa kwaye uMzobo oSongezelelo we-S7 ubonisa iiprofayili ezishushu ezibandakanya imizuzu engama-20 yexesha lokubhalwa kwe-reverse kunye nemizuzu engama-60 yexesha lokuhamba ngebhayisikile eshushu (95 kunye ne-60 °C), kunye nomjikelo omnye we-thermal ube yi-90 s (i-Movie eyoNgezelelweyo ye-S7)..I-FAST-POCT idinga ixesha elincinci lokugqiba umjikelezo we-thermal (imizuzwana ye-90) kune-RT-PCR eqhelekileyo (imizuzwana ye-180 yomjikelezo omnye we-thermal).Oku kunokuchazwa ngummandla ophezulu ukuya kumlinganiselo wevolumu kunye ne-inertia ephantsi ye-thermal yegumbi lokuphendula le-micro-PCR.Umphezulu wegumbi ngu-96.6 mm2 kunye nomthamo wegumbi ngu-25 mm3, okwenza umphezulu we-volume ratio malunga ne-3.86.Njengoko kubonwa kuMzobo owoNgezelelweyo we-S10, indawo yovavanyo lwe-PCR yeqonga lethu inegroove kwiphaneli yangasemva, eyenza umzantsi wegumbi lePCR ube ngama-200 µm ubukhulu.Iphedi ye-elastic conductive thermally ifakwe kwindawo yokufudumala yokulawula ubushushu, iqinisekisa ukudibanisa okuqinileyo kunye nomva webhokisi lovavanyo.Oku kunciphisa inertia ye-thermal yeqonga kunye nokuphucula ukufudumeza / ukupholisa ukusebenza kakuhle.Ngexesha lokuhamba ngebhayisikile ye-thermal, iparafini efakwe kwiqonga iyanyibilika kwaye iqukuqelele kwi-PCR reaction chamber, isebenza njenge-sealant ukuthintela ukuvela kwe-reagent evaporation kunye nokungcoliswa kwendalo (jonga i-Supplementary Movie S8).
Zonke iinkqubo zokufumanisa i-PCR ezichazwe ngasentla zenziwe ngokuzenzekelayo zisebenzisa i-custom eyenziwe ye-FAST-POCT isixhobo, equlethwe yiyunithi yokulawula uxinzelelo olucwangcisiweyo, iyunithi yokukhutshwa kwamagnetic, iyunithi yokulawula ukushisa, kunye neyunithi yesignali ye-fluorescent yokubamba kunye neyunithi yokucubungula.Inqaku, sasebenzisa iqonga le-FAST-POCT lokuzihlukanisa kwe-RNA kwaye emva koko sasebenzisa iisampuli ze-RNA ezikhutshiweyo kwiimpendulo ze-PCR zisebenzisa inkqubo ye-FAST-POCT kunye nenkqubo ye-PCR yedesktop ukuze kuthelekiswe.Iziphumo beziphantse zafana njengoko kubonisiwe kuMzobo oSongezelelo we-S8.Umqhubi wenza umsebenzi olula: ungenisa isampuli kwi-M-chamber kwaye ufaka iqonga kwisixhobo.Iziphumo zovavanyo lobungakanani ziyafumaneka malunga nemizuzu engama-82.Ulwazi oluthe vetshe malunga nezixhobo ze-FAST-POCT zinokufumaneka kwinani elongezelelweyo.C9, C10 kunye neC11.
Umkhuhlane obangelwa yintsholongwane yomkhuhlane A (IAV), B (IBV), C (ICV) kunye ne-D (IDV) yinto eqhelekileyo yehlabathi.Kwezi, i-IAV kunye ne-IBV zinoxanduva lwezona meko zinzima kakhulu kunye nezibetho zonyaka, zosulela i-5-15% yabemi behlabathi, kubangele i-3-5 yezigidi zeemeko ezinzima kwaye kubangele ukufa kwe-290,000-650,000 ngonyaka.Izifo zokuphefumla56,57.Ukuxilongwa kwangaphambili kwe-IAV kunye ne-IB kubalulekile ukunciphisa ukugula kunye nomthwalo wezoqoqosho ohambelanayo.Phakathi kweendlela ezifumanekayo zokuxilonga, i-reverse transcriptase polymerase chain reaction (RT-PCR) ithathwa njengeyona inovakalelo, ithile, kwaye ichanekile (> 99%) 58,59.Phakathi kweendlela ezifumanekayo zokuxilonga, i-reverse transcriptase polymerase chain reaction (RT-PCR) ithathwa njengeyona inovakalelo, ithile, kwaye ichanekile (> 99%) 58,59.Среди доступных диагностических методов полимеразная цепная реакция с обратной транскриптазой (ОТ-ПЦР) считается наиболевичи > 99%)58,59.Phakathi kweendlela ezifumanekayo zokuxilonga, i-reverse transcriptase polymerase chain reaction (RT-PCR) ithathwa njengeyona inovakalelo, ichanekileyo kwaye ichanekile (> 99%) 58,59. Из доступных диагностических методов полимеразная цепная реакция с обратной транскриптазой (ОТ-ПЦР) считается наийстолевие, песни 99%)58,59.Kwiindlela ezifumanekayo zokuxilonga, i-reverse transcriptase polymerase chain reaction (RT-PCR) ithathwa njengeyona ibucayi, ichanekileyo kwaye ichanekile (> 99%) 58,59.Nangona kunjalo, iindlela zendabuko ze-RT-PCR zifuna ukuphindaphinda kwemibhobho, ukuxuba, ukukhupha kunye nokudluliselwa kolwelo, ukunciphisa ukusetyenziswa kwabo ngabasebenzi kwiisetingi ezinobuncwane.Apha, iqonga le-FAST-POCT lalisetyenziselwa ukubonwa kwe-PCR ye-IAV kunye ne-IBV, ngokulandelanayo, ukufumana umda wabo ophantsi wokufumanisa (LOD).Ukongezelela, i-IAV kunye ne-IBV ziye zaphindwa ngokuphindaphindiweyo ukucalula phakathi kweentlobo ezahlukeneyo ze-pathotypes kuzo zonke iintlobo, ukubonelela ngeqonga elithembisayo lokuhlalutya kofuzo kunye nokukwazi ukuphatha ngokuchanekileyo isifo.
Kwikhiwane.I-5a ibonisa iziphumo zovavanyo lwe-HAV PCR usebenzisa i-150 µl ye-RNA yentsholongwane ecocekileyo njengesampulu.Kwikhiwane.I-5a (i) ibonisa ukuba kwi-HAV yoxinaniso lweekopi ze-106 / ml, i-fluorescence intensity (ΔRn) inokufikelela kwi-0.830, kwaye xa i-concentration iyancipha ukuya kwiikopi ze-102 / ml, i-ΔRn isenokufikelela kwi-0.365, ehambelana nephezulu kunoko. yeqela elingenanto elingalunganga lokulawula (0.002), malunga namaxesha angama-100 aphezulu.Ukulinganisa ubungakanani obusekelwe kwiimvavanyo ezithandathu ezizimeleyo, i-curve yokulinganisa i-linear yaveliswa phakathi kwe-log concentration kunye ne-cycle threshold (Ct) ye-IAV (Fig. 5a (ii)), i-R2 = 0.993, ukusuka kwi-102-106 iikopi / mL.iziphumo zivumelana kakuhle neendlela eziqhelekileyo ze-RT-PCR.Kwikhiwane.I-5a(iii) ibonisa imifanekiso ye-fluorescent yeziphumo zovavanyo emva kwemijikelo ye-40 yeqonga le-FAST-POCT.Sifumene ukuba iqonga le-FAST-POCT liyakwazi ukubona i-HAV ephantsi njengeekopi ze-102 / mL.Nangona kunjalo, indlela yendabuko ayinayo ixabiso le-Ct kwiikopi ze-102 / mL, okwenza kube yi-LOD malunga neekopi ze-103 / mL.Siye saqikelela ukuba oku kungenxa yobuchule obuphezulu bokuxuba amaqamza.Uvavanyo lovavanyo lwe-PCR lwenziwa kwi-IAV RNA esulungekileyo yokuvavanya iindlela ezahlukeneyo zokuxuba, kubandakanya ukuxubana kwe-shake (indlela yokuxuba efanayo njengakwindlela eqhelekileyo yokusebenza kwe-RT-PCR), ukuxuba i-vial (le ndlela, i-3 s kwi-0.12 bar) kwaye akukho kuxubana njengeqela lolawulo. ..Iziphumo zinokufumaneka kwi-Supplementary Figure S12.Ingabonwa ukuba kugxininiso lwe-RNA ephezulu (iikopi ezili-106/mL), amaxabiso e-Ct eendlela ezahlukeneyo zokuxuba aphantse afane nokuxutywa kweqamza.Xa i-RNA i-concentration yehla kwiikopi ze-102 / mL, ukuxuba ukuxuba kunye nokulawula kwakungekho ixabiso le-Ct, ngelixa indlela yokuxuba i-bubble isanika ixabiso le-Ct ye-36.9, eyayingaphantsi kwe-Ct threshold ye-38. Iziphumo zibonisa uphawu oluxubileyo oluxubileyo iivesicles, eziye zaboniswa nakwezinye iincwadi, ezinokuchaza ukuba kutheni uvakalelo lweqonga le-FAST-POCT liphezulu kancinane kune-RT-PCR eqhelekileyo.Kwikhiwane.I-5b ibonisa iziphumo zohlalutyo lwe-PCR yeesampuli ze-IBV RNA ezihlambulukileyo ukusuka kwi-101 ukuya kwi-106 iikopi / ml.Iziphumo zazifana novavanyo lwe-IAV, ukufumana i-R2 = 0.994 kunye ne-LOD yeekopi ezili-102/mL.
uhlalutyo lwe-PCR ye-influenza A virus (IAV) kunye nogxininiso lwe-IAV ukusuka kwi-106 ukuya kwi-101 iikopi / mL usebenzisa i-TE buffer njengolawulo olubi (NC).(i) Ixesha lokwenyani ijiko le-fluorescence.(ii) I-curve calibration curve phakathi kwe-logarithmic IAV RNA yoxinaniso kunye nomjikelezo womjikelo (Ct) we-FAST kunye neendlela zokuvavanya eziqhelekileyo.(iii) Umfanekiso wefluorescent we-IAV FAST-POCT emva kwemijikelo engama-40.b, ukufunyaniswa kwe-PCR yentsholongwane ye-influenza B (IBV) kunye (i) nexesha langempela le-fluorescence spectrum.(ii) Igophe lokulinganisa umgca kunye (iii) FAST-POCT IBV umfanekiso we-fluorescence emva kwemijikelo engama-40.Umda ophantsi wokufumanisa (LOD) we-IAV kunye ne-IBV usebenzisa iqonga le-FAST-POCT laliyi-102 iikopi / mL, ephantsi kuneendlela eziqhelekileyo (iikopi ze-103 / mL).c Iziphumo zovavanyo ezininzi ze-IAV kunye ne-IBV.I-GAPDH yasetyenziswa njengolawulo olulungileyo kwaye i-TE buffer yasetyenziswa njengolawulo olubi ukuthintela ukungcoliseka okunokwenzeka kunye nokwandiswa kwemvelaphi.Iintlobo ezine zeesampulu ezahlukeneyo zinokuhlukaniswa: (1) i-GAPDH-kuphela iisampulu ezingalunganga ("IAV-/IBV-");(2) usulelo lwe-IAV ("IAV +/IBV-") kunye ne-IAV kunye ne-GAPDH;(3) Usulelo lwe-IBV (“IAV-/IBV +”) nge-IBV kunye ne-GAPDH;(4) Usulelo lwe-IAV/IBV (“IAV +/IBV +”) nge-IAV, IBV kunye ne-GAPDH.Umgca onamachaphaza umele umda.n = Imifuniselo emi-6 ezimeleyo ngokwebhayoloji yenziwe, idatha iboniswa ± ukutenxa okusemgangathweni.Idatha ekrwada inikezelwa njengeefayili zedatha ekrwada.
Kwikhiwane.I-5c ibonisa iziphumo zovavanyo lwe-multiplexing lwe-IAV/IBV.Apha, i-virus lysate isetyenziswe njengesisombululo sesampuli endaweni ye-RNA ecocekileyo, kunye ne-primers ezine ze-IAV, i-IBV, i-GAPDH (ulawulo oluhle) kunye ne-TE buffer (ulawulo olubi) zongezwa kumagumbi amane okusabela ahlukeneyo kwi-platform ye-FAST-POCT.Ulawulo olulungileyo nolungalunganga lusetyenziswa apha ukuthintela ukosuleleka okunokwenzeka kunye nokwandiswa kwemvelaphi.Iimvavanyo zahlulahlulwe ngamaqela amane: (1) iisampuli ze-GAPDH-negative ("IAV-/IBV-");(2) I-IAV-isulelwe ("IAV +/IBV-") ngokuchasene ne-IAV kunye ne-GAPDH;(3) IBV-.abosulelekileyo (“IAV-”) -/IBV+”) IBV kunye neGAPDH;(4) IAV / IBV ("IAV +/IBV +") ukusuleleka nge-IAV, IBV kunye ne-GAPDH.Kwikhiwane.I-5c ibonisa ukuba xa iisampulu ezingalunganga zisetyenzisiwe, ukuqina kwe-fluorescence ΔRn yegumbi lokulawula elilungileyo laliyi-0.860, kwaye i-ΔRn ye-IAV kunye ne-IBV yayifana nolawulo olubi (0.002).Kumaqela e-IAV+/IBV-, IAV-/IBV+ kunye ne-IAV+/IBV+, iikhamera ze-IAV/GAPDH, IBV/GAPDH kunye ne-IAV/IBV/GAPDH zibonise ubunzulu be-fluorescence, ngokulandelelanayo, ngelixa ezinye iikhamera zibonise ubunzulu be-fluorescence ngasemva. inqanaba le-40 emva kokuhamba ngebhayisikili eshushu.Ukusuka kwiimvavanyo ezingentla, iqonga le-FAST-POCT libonise ukuchaneka okubalaseleyo kwaye lisivumele ukuba ngaxeshanye sisebenzise iintsholongwane zomkhuhlane ezahlukeneyo.
Ukuqinisekisa ukusetyenziswa kweklinikhi ye-FAST-POCT, siye savavanya iisampuli zekliniki ze-36 (imizekelo yeempumlo ze-swab) kwizigulane ze-IB (n = 18) kunye nokulawulwa kwe-non-IB (n = 18) (Umfanekiso 6a).Ulwazi lwesigulane lunikezelwa kwiTheyibhile eyoNgezelelweyo 3. Isimo sosulelo lwe-IB saqinisekiswa ngokuzimeleyo kwaye iprotocol yokufunda yavunywa yiZhejiang University First Affiliated Hospital (Hangzhou, Zhejiang).Isampulu nganye yezigulane yahlulahlulwe yangamacandelo amabini.Enye yacutshungulwa kusetyenziswa i-FAST-POCT kwaye enye yacutshungulwa kusetyenziswa inkqubo yePCR yedesktop (SLAN-96P, China).Zombini iimvavanyo zisebenzisa izixhobo ezifanayo zokucoca kunye nokufumanisa.Kwikhiwane.I-6b ibonisa iziphumo ze-FAST-POCT kunye ne-PCR ye-reverse transcription (RT-PCR).Siqhathanise i-fluorescence intensity (FAST-POCT) kunye -log2 (Ct), apho i-Ct ingumda womjikelo we-RT-PCR eqhelekileyo.Kwakukho ukuvumelana okuhle phakathi kwezi ndlela zimbini.I-FAST-POCT kunye ne-RT-PCR ibonise ukulungelelaniswa okuqinisekileyo okuqinileyo kunye ne-Pearson's ratio (r) ixabiso le-0.90 (Umfanekiso 6b).Emva koko siye savavanya ukuchaneka kokuxilongwa kwe-FAST-POCT.Ukuhanjiswa kwe-Fluorescence intensity (FL) kwiisampuli ezilungileyo kunye nezibi zinikezelwe njengomlinganiselo ozimeleyo wokuhlalutya (Umfanekiso 6c).Amaxabiso e-FL ayephezulu kakhulu kwizigulane ze-IB kunezolawulo (****P = 3.31 × 10-19; uvavanyo lwe-t-tailed ezimbini) (umzobo 6d).Emva koko, iimpawu ze-IBV zokusebenzisa i-receiver (ROC) i-curves yacwangciswa.Sifumene ukuba ukuchaneka kokuxilonga kwakukuhle kakhulu, kunye nommandla ongaphantsi kwe-curve ye-1 (Umfanekiso 6e).Nceda uqaphele ukuba ngenxa yomyalelo wemaski osisinyanzelo e-China ngenxa ye-COVID-19 ukusukela ngo-2020, asikhange sichonge abaguli abane-IBD, ngoko ke yonke imizekelo yekliniki eqinisekileyo (okt, iisampulu ze-swab yeempumlo) yayize-IBV kuphela.
Uyilo lwesifundo seklinikhi.Iisampulu ze-36 ezipheleleyo, ezibandakanya i-18 iisampulu zesigulane kunye ne-18 yokulawula umkhuhlane, zahlaziywa kusetyenziswa iqonga le-FAST-POCT kunye ne-RT-PCR eqhelekileyo.b Ukuvavanya ukuhambelana kohlalutyo phakathi kwe-FAST-POCT PCR kunye ne-RT-PCR eqhelekileyo.Iziphumo zihambelana kakuhle (Pearson r = 0.90).c Amanqanaba e-Fluorescence intensity kwizigulane ezili-18 ze-IB kunye nolawulo lwe-18.d Kwizigulane ze-IB (+), amaxabiso e-FL ayephezulu kakhulu kuneqela lokulawula (-) (****P = 3.31 × 10-19; uvavanyo lwe-t-tailed; n = 36).Kwindawo nganye yesikwere, umakishi omnyama embindini umele i-median, kwaye imigca engezantsi kunye nephezulu yebhokisi imele i-25th kunye ne-75th percentiles, ngokulandelanayo.I-whiskers yandisa kwiindawo ezincinci kunye nezona ziphezulu zedatha, ezingaqwalaselwanga ngaphandle.e ROC ijika.Umgca onamachaphaza u-d ubonisa ixabiso lomqobo eliqikelelwa kuhlalutyo lwe-ROC.I-AUC ye-IBV yi-1. Idatha ekrwada ibonelelwa njengeefayili zedatha ekrwada.
Kweli nqaku, sibonisa i-FAST, eneempawu ezifunekayo kwi-POCT efanelekileyo.Iinzuzo zeteknoloji yethu ziquka: (1) I-dosing eguquguqukayo (i-cascade, ngaxeshanye, ilandelelana kwaye ikhethiweyo), ukukhululwa kwimfuno (ukukhululwa ngokukhawuleza kunye nokulinganisa koxinzelelo olusetyenzisiweyo) kunye nokusebenza okuthembekileyo (i-vibration kwii-150 degrees) (2) ukugcinwa kwexesha elide. (iminyaka eyi-2 yokuvavanya ngokukhawuleza, ukulahleka kwesisindo malunga ne-0.3%);(3) ukukwazi ukusebenza kunye nolwelo kunye noluhlu olubanzi lokumanzisa kunye ne-viscosity (i-viscosity ukuya kwi-5500 cP);(4) Uqoqosho (Ixabiso lezinto eziqikelelweyo zesixhobo se-FAST-POCT PCR malunga ne-US $ 1).Ngokudibanisa i-multifunctional dispensers, i-platform ye-FAST-POCT edibeneyo yokufumanisa i-PCR ye-influenza A kunye ne-B viruses yaboniswa kwaye yasetyenziswa.I-FAST-POCT ithatha kuphela imizuzu engama-82.Iimvavanyo zeklinikhi kunye neesampuli ze-36 ze-swab zempumlo zibonise i-concordance efanelekileyo kwi-fluorescence intensity kunye ne-RT-PCR eqhelekileyo (i-Pearson coefficients> 0.9).Iimvavanyo zeklinikhi kunye neesampuli ze-36 ze-swab zempumlo zibonise i-concordance efanelekileyo kwi-fluorescence intensity kunye ne-RT-PCR eqhelekileyo (i-Pearson coefficients> 0.9).Клинические тесты с 36 образцами мазков из носа показали хорошее соответствие интенсивности флуоресценции стандартной ОТ-ПЦенция ОТ-П,фенции стандартной ОТ-П,фенцийIimvavanyo zeklinikhi ezineisampulu ezingama-36 zeempumlo zempumlo zibonise isivumelwano esihle kunye nokuqina kwe-fluorescence ye-RT-PCR eqhelekileyo (i-Pearson's coefficients> 0.9) .RT-PCR Клинические испытания 36 образцов мазков из носа показали хорошее совпадение интенсивности флуоресценции со стандартной ОТ-эфинценции стандартной ОТ-ПефенцийUvavanyo lwezonyango lweesampulu ze-36 zempumlo ze-swab zibonise ukuvumelana okulungileyo kokuqina kwe-fluorescence kunye ne-RT-PCR eqhelekileyo (i-coefficient ka-Pearson> 0.9).Ngokuhambelana nalo msebenzi, iindlela ezahlukeneyo ezivelayo ze-biochemical (umzekelo, i-plasma thermal cycling, i-amplification-free immunoassays, kunye ne-nanobody functionalization assays) ibonise amandla abo kwi-POCT.Nangona kunjalo, ngenxa yokunqongophala kweqonga le-POCT elidityanisiweyo ngokupheleleyo kunye nelomeleleyo, ezi ndlela zifuna ngokungaphephekiyo iinkqubo ezahlukeneyo zangaphambili (umzekelo, i-RNA isolation44, incubation45 kunye ne-washing46), ehambelana ngakumbi nomsebenzi wangoku kunye nezi ndlela zokuphumeza imisebenzi ye-POCT ephezulu iiparamitha ezifunekayo.landa-kwimpendulo-imveliso yokusebenza.Kulo msebenzi, nangona ipompo yomoya esetyenziselwa ukuvula i-valve ye-FAST incinci ngokwaneleyo ukuba ihlanganiswe kwisixhobo se-benchtop (Umfanekiso we-S9, i-S10), isasebenzisa amandla amakhulu kwaye ivelise ingxolo.Ngokomgaqo, iimpompo zepneumatic zefom encinci zinokutshintshwa ngezinye iindlela, ezinjengokusetyenziswa kwamandla ombane wombane okanye ukusebenza komnwe.Uphuculo olongezelelweyo lunokubandakanya, umzekelo, ukulungelelanisa iikiti kwiimvavanyo ezahlukeneyo kunye nezichanekileyo ze-biochemical, usebenzisa iindlela ezintsha zokufumanisa ezingadingi iinkqubo zokufudumeza / zokupholisa, ngaloo ndlela zibonelela ngeqonga le-POCT elingenaso isixhobo kwizicelo ze-PCR.Sikholelwa ukuba iqonga le-FAST libonelela ngendlela yokulawula ulwelo, sikholelwa ukuba itekhnoloji ye-FAST ecetywayo ibonisa amandla okudala iqonga eliqhelekileyo kungekuphela nje uvavanyo lwe-biomedical, kodwa kunye nokubeka iliso kokusingqongileyo, ukuvavanywa komgangatho wokutya, izixhobo kunye nokuhlanganiswa kweziyobisi. ..
Ukuqokelela kunye nokusetyenziswa kwemizekelo ye-swab yeempumlo zomntu kuye kwavunywa yiKomiti yeeNqobo zokuziphatha yeSibhedlele esiManyano neYunivesithi yaseZhejiang (IIT20220330B).Iisampuli ze-36 ze-swab zempumlo zaqokelelwa, ezibandakanya abantu abadala abayi-16 abangaphantsi kweminyaka engama-30 ubudala, abantu abadala abayi-7> iminyaka engama-40 ubudala, kunye ne-19 yamadoda, i-17 yabasetyhini.Iisampuli ze-36 ze-swab zempumlo zaqokelelwa, ezibandakanya abantu abadala abayi-16 abangaphantsi kweminyaka engama-30 ubudala, abantu abadala abayi-7> iminyaka engama-40 ubudala, kunye ne-19 yamadoda, i-17 yabasetyhini.Было собрано 36 образцов мазков из носа, в которых приняли участие 16 взрослых < 30 лет, 7 взрослых старше 40 лемун17 инге 19Kwaqokelelwa imizekelo yamashumi amathathu anesithandathu kwimpumlo yabantu abadala abali-16 abangaphantsi kweminyaka engama-30 ubudala, abasi-7 abangaphezulu kweminyaka engama-40 ubudala, amadoda ali-19 kunye nabasetyhini abali-17..Idatha yedemografi inikwe kwiTheyibhile eyoNgezelelweyo yesi-3. Imvume enolwazi ifunyenwe kubo bonke abathathi-nxaxheba.Bonke abathathi-nxaxheba barhanelwa ukuba banomkhuhlane kwaye bavavanywa ngokuzithandela ngaphandle kwembuyekezo.
Isiseko se-FAST kunye nesivalo senziwe nge-polylactic acid (PLA) kwaye iprintwe ngu-Ender 3 Pro 3D umshicileli (i-Shenzhen Transcend 3D Technology Co., Ltd.).Iteyiphu enamacala amabini yathengwa kwi-Adhesives Research, Inc. Model 90880. Ifilimu ye-PET eyi-100 µm ubukhulu yathengwa kwaMcMaster-Carr.Zombini i-adhesive kunye nefilimu ye-PET yasikwa kusetyenziswa i-Silhouette Cameo 2 cutter esuka kwi-Silhouette America, Inc. Ifilimu e-elastic yenziwe ngezinto ze-PDMS ngokubunjwa kwe-injection.Okokuqala, i-200 µm engqingqwa ye-PET isakhelo yasikwa kusetyenziswa inkqubo ye-laser kwaye yancamathelwa kwiphepha le-PMMA elishinyeneyo le-3 mm kusetyenziswa i-100 µm yeteyiphu yokuncamathelisa kabini.I-precursor ye-PDMS (i-Sylgard 184; iCandelo A: Ingxenye B = 10: 1, i-Dow Corning) yaza yagalelwa kwi-mold kunye nentonga yeglasi yasetyenziselwa ukususa i-PDMS engaphezulu.Emva kokunyanga kwi-70 ° C. kwiiyure ezi-3, ifilimu ye-PDMS ye-300 μm eshinyeneyo inokuxolwa kubumba.
Iifoto zokusasazwa okuguquguqukayo, upapasho olufunwayo kunye nokusebenza okuthembekileyo kuthathwa ngekhamera yesantya esiphezulu (i-Sony AX700 1000 fps).I-orbital shaker esetyenziswe kuvavanyo lokuthembeka yathengwa kwi-SCILOGEX (SCI-O180).Uxinzelelo lomoya lwenziwa yi-compressor yomoya, kwaye abalawuli abaninzi bedijithali bokuchaneka koxinzelelo basetyenziselwa ukulungelelanisa ixabiso loxinzelelo.Inkqubo yokuvavanya ukuziphatha kokuhamba ngolu hlobo lulandelayo.Inani elimiselweyo lolwelo lifakwe kwisixhobo sokuvavanya kwaye ikhamera yesantya esiphezulu yayisetyenziselwa ukurekhoda ukuziphatha kokuhamba.Imifanekiso esaqhubeka yathathwa kwiividiyo zokuziphatha okuhamba ngamaxesha amisiweyo, kwaye indawo eseleyo yabalwa kusetyenziswa isoftware ye-Image-Pro Plus, eyathi yaphindaphindwa ngobunzulu bekhamera ukubala umthamo.Iinkcukacha zenkqubo yokuvavanya ukuziphatha okuhambayo zinokufumaneka kwi-Supplementary Figure S4.
Tofa i-50 µl yee-microbeads kunye ne-100 µl yamanzi adibeneyo kwisixhobo sokuxuba i-vial.Iifoto zokusebenza ezixubileyo zithathwe ngekhamera yesantya esiphezulu rhoqo ngemizuzwana ye-0.1 kwiingcinezelo ze-0.1 bar, i-0.15 bar kunye ne-0.2 bar.Ulwazi lwePixel ngexesha lenkqubo yokudibanisa inokufumaneka kule mifanekiso kusetyenziswa isoftware yokulungisa ifoto (Photoshop CS6).Kwaye ukudibanisa ukusebenza kakuhle kunokufezekiswa ngale Equation 53 ilandelayo.
apho uM kukuxutywa kakuhle, N linani lilonke lesampulu yeepikseli, kunye ne-ci kunye \(\bar{c}\) zizigxina eziqhelekileyo nezilindelekileyo.Ukudibanisa ukusebenza kakuhle ukusuka kwi-0 (0%, engaxutywanga) ukuya kwi-1 (100%, ixutywe ngokupheleleyo).Iziphumo ziboniswe kuMzobo owoNgezelelweyo we-S6.
Ikiti ye-RT-PCR yexesha langempela ye-IAV kunye ne-IBV, kuquka i-IAV kunye ne-IBV RNA iisampulu (cat. No. RR-0051-02/RR-0052-02, Liferiver, China), Tris-EDTA buffer (TE buffer no. B541019 , I-Sangon Biotech, i-China), i-Positive Control ye-RNA Purification Kit (Ingxenye ye-Z-ME-0010, i-Liferiver, i-China) kunye ne-GAPDH Isisombululo (Ingxenye yeNombolo ye-M591101, i-Sangon Biotech, i-China) ifumaneka ngokurhweba.Ikhithi yokucoca i-RNA ibandakanya isithinteli esibophelelayo, ukuhlamba A, ukuhlamba i-W, i-eluent, i-microbeads magnetic, kunye ne-acrylic carrier.I-IAV kunye ne-IBV yexesha langempela leekiti ze-RT-PCR ziquka i-IFVA nucleic acid PCR umxube wokufumanisa kunye ne-enzyme ye-RT-PCR.Yongeza i-6 µl ye-AcrylCarrier kunye ne-20 µl yamaso kazibuthe ukuya kwi-500 µl yesisombululo se-buffer esibophezelayo, hlunguza kakuhle uze ulungise isisombululo se-bead.Yongeza i-21 ml ye-ethanol ukuhlamba i-A kunye ne-W, xubha kakuhle ukufumana izisombululo zokuhlamba i-A kunye ne-W, ngokulandelanayo.Emva koko, i-18 µl yomxube we-fluorescent PCR kunye ne-IFVA nucleic acid kunye ne-1 µl ye-enzyme ye-RT-PCR zongezwa kwi-1 µl yesisombululo se-TE, yashukunyiswa kwaye yafakwa i-centrifuged imizuzwana emininzi, ukufumana i-20 µl ye-IAV kunye ne-IBV primers.
Landela le nkqubo ilandelayo yokucoca i-RNA: (1) I-RNA adsorption.I-Pipette 526 µl yesisombululo se-pellet kwityhubhu eyi-1.5 ml ye-centrifuge kwaye wongeze i-150 µl yesampulu, emva koko ushukumise ityhubhu ngesandla phezulu naphantsi amaxesha ali-10.Dlulisa i-676 µl yomxube kwikholamu ye-affinity kunye ne-centrifuge kwi-1.88 x 104 g imizuzwana engama-60.Iidreyini ezilandelayo ziyalahlwa.(2) Inqanaba lokuqala lokuhlamba.Yongeza i-500 µl yesisombululo sokuhlambela A kwikholamu ye-affinity, i-centrifuge kwi-1.88 x 104 g ye-40 s, kwaye ulahle isisombululo esichithiweyo.Le nkqubo yokuhlamba iphindwe kabini.(3) inqanaba lesibini lokuhlamba.Yongeza i-500 µl yesisombululo sokuhlamba W kwikholamu yokubambisana, i-centrifuge kwi-1.88×104 g ye-15 s kwaye ulahle isisombululo esichithiweyo.Le nkqubo yokuhlamba iphindwe kabini.(4) Uvavanyo.Yongeza i-200 µl ye-eluate kwi-affinity column kunye ne-centrifuge kwi-1.88 x 104 g nge-2 min.(5) I-RT-PCR: I-eluate ifakwe kwi-20 μl yesisombululo se-primer kwi-tube ye-PCR, emva koko ityhubhu ifakwe kwi-PCR ye-real-time test apparatus (SLAN-96P) ukuqhuba inkqubo ye-RT-PCR.Yonke inkqubo yokufumanisa ithatha malunga nemizuzu eyi-140 (imizuzu engama-20 yokuhlanjululwa kwe-RNA kunye nemizuzu eyi-120 yokufunyanwa kwe-PCR).
I-526 µl yesisombululo se-bead, i-1000 µl yesisombululo sokuhlamba A, 1000 µl yesisombululo sokuhlamba W, 200 µl ye-eluate kunye ne-20 µl yesisombululo se-primer yadityaniswa kwangaphambili kwaye yagcinwa kumagumbi okubona uM, W1, W2, E kunye ne-PCR.Indibano yeqonga.Emva koko, i-150 µl yesampulu yafakwa ngombhobho kwigumbi elingu-M kwaye iqonga le-FAST-POCT lafakwa kwisixhobo sovavanyo esiboniswe kuMfanekiso owongezelelweyo S9.Emva kwemizuzu engama-82, iziphumo zovavanyo zazifumaneka.
Ngaphandle kokuba kuphawulwe ngenye indlela, zonke iziphumo zovavanyo zinikwa njengentsingiselo ± SD emva ubuncinane beziphindaphindo ezintandathu kusetyenziswa kuphela iqonga le-FAST-POCT kunye neesampulu ezizimeleyo ngokwebhayoloji.Akukho datha ikhutshiweyo kuhlalutyo.Imifuniselo ayizenzelwanga.Abaphandi abakhange babe yimfama kwimisebenzi yeqela ngexesha lovavanyo.
Ukufumana ulwazi oluthe kratya malunga noyilo lwesifundo, bona iNgxelo yoPhando lweNdalo eqhagamshelwe kweli nqaku.
Idatha exhasa iziphumo zolu phononongo iyafumaneka kwiiNkcukacha ezoNgezelelweyo.Eli nqaku libonelela ngedatha yokuqala.
I-Chagla, Z. & Madhukar, P. COVID-19 booster kumazwe atyebileyo iya kulibazisa amayeza okugonya kubo bonke.I-Chagla, Z. & Madhukar, P. COVID-19 booster kumazwe atyebileyo iya kulibazisa amayeza okugonya kubo bonke.I-Chagla, Z. kunye ne-Madhukar, P. COVID-19 booster kumazwe atyebileyo iya kulibazisa amayeza okugonya kuye wonke umntu.I-Chagla, Z. kunye ne-Madhukar, P. COVID-19 revaccination kumazwe atyebileyo iya kulibazisa ugonyo kuye wonke umntu.Iyeza lesizwe.27, 1659-1665 (2021).
Faust, L. et al.Uvavanyo lwe-SARS-CoV-2 kumazwe anengeniso ephantsi kunye nephakathi: ukufumaneka kunye nokufikeleleka kwicandelo lezempilo labucala.usulelo lwemicrobial.22, 511–514 (2020).
I-World Health Organization.Ukuxhaphaka kwehlabathi kunye neziganeko zezifo ezikhethiweyo ezinokuthi zinyangeke ngesondo: uphononongo kunye noqikelelo.I-Geneva: WHO, WHO/HIV_AIDS/2 https://apps.who.int/iris/bitstream/handle/10665/66818/WHO_HIV_AIDS_2001.02.pdf (2001).
Fenton, EM et al.Imicu yovavanyo lwecala lokuqukuqela lwe-2D emininzi.Isicelo se-ASS.i-alma mater.Inter Milan.1, 124-129 (2009).
Schilling, KM et al.Isixhobo sokuhlalutya esisekwe kwi-microfluidic efakwe ngokupheleleyo.umva.Ikhemikhali.84, 1579-1585 (2012).
Lapenter, N. et al.I-immunochromatography esekelwe kwiphepha elikhuphisanayo kunye ne-enzyme-modified electrode ivumela ukubeka iliso ngaphandle kwamacingo kunye nokuzimisela kwe-electrochemical ye-urinary cotinine.Iinzwa 21, 1659 (2021).
Zhu, X. et al.Ukulinganisa iimpawu zebhayoloji zesifo kunye neqonga le-nanozyme-edibeneyo edibeneyo yolwelo olusecaleni kusetyenziswa iglucometer.isivamvo sebhayoloji.I-Bioelectronics.126, 690-696 (2019).
Boo, S. et al.Umtya wovavanyo lokukhulelwa ukufumanisa iibhaktheriya ze-pathogenic usebenzisa i-concanavalin A-human chorionic gonadotropin-Cu3 (PO4)2 i-nanoflowers edibeneyo, ukuhlukana kwamagnetic kunye nokufunda nge-smart phone.I-Microcomputer.Iphephancwadi.185, 464 (2018).