Uhlahlelo lwamanyathelo kunye neMiba yokongezwa kweeSampulu ze-ELISA

Ukunyamekela kufuneka kuthathwe ukuphepha i-hemolysis enzima kwindawo yokuqala. IHemoglobin ineqela le-heme, elinomsebenzi ofana neeperoxide. Ngoko ke, ekuzimiseleni kwe-ELISA kunye ne-HRP njenge-enzyme ebhaliweyo, ukuba i-hemoglobin concentration kwi-serum specimen iphezulu, kulula ukubhengezwa kwisigaba esiqinileyo kwinkqubo yokuzala, ukuze uphendule kunye ne-HRP substrate eyongezwe kamva kuphuhliso lombala. Iisampuli zeSerum ezahlulwe ngokusebenza kwe-aseptic zingagcinwa kwi-2 ~ 8℃kwiveki, kunye nokugcinwa komkhenkce kuyacetyiswa ukuba ibhaktheriya ibandakanyeka. Iisampulu kufuneka zigcinwe ngaphantsi -70 isidanga ixesha elide. Ukuqokelela isampuli kunye nokwahlula serum kufuneka ingqalelo ukuphepha ukungcoliseka ibhaktheriya kangangoko kunokwenzeka, ukukhula kwebhaktheriya, secretion yayo ezinye enzymes inokuba antigens, amajoni omzimba kunye nezinye iiproteni isiphumo ukubola; Kwelinye icala, ezinye ii-endogenous enzymes zebhaktiriya, ezinje nge- -igalactosidase ye-Escherichia coli, ngokwazo ziya kubangela uphazamiseko olukhethekileyo kuvavanyo oluphawulwe nge-enzyme ehambelana nayo. Kwimeko ye-turbidities okanye i-flocculent ebangelwa ukungcoliseka kwebhaktheriya ngexesha lokugcinwa kwesampuli, i-supernatant kufuneka ithathwe ukuze ibonwe emva kwemvula ye-centrifugation. Umkhenkce{13}}ukunyibilikiswa kwesampulu zeserum kufuneka kuphetshwe ngenxa yokusilela kwamandla. Amandla okucheba ngoomatshini eveliswa kukunyibilika okuphindaphindiweyo{15}} komfanekiso kuya kutshabalalisa iiproteni kunye nezinye iimolekyuli kumzekelo, okukhokelela kwiziphumo ezingezizo ezingalunganga. Ukongezelela, ingqalelo kufuneka ihlawulwe kwakhona ekuxutyweni kweesampuli ze-freeze-thaw, kwaye i-oscillations enobudlova ayifanele yenziwe. Ukuxuba kufuneka kuguqulwe ngokuphindaphindiweyo.

Kwi-ELISA, kukho amanyathelo amathathu ngokubanzi okongeza iisampulu, oko kukuthi, ukongeza i-enzyme conjugate, yongeza i-substrate. Emva kokunyibilika kwe-serum efriziweyo, iprotheni igxininiswe kwindawo kwaye isasazwe ngokungalingani. Kufuneka ixutywe ngokupheleleyo, ngobunono kwaye igweme amaqamza. Inokuxutywa ijonge phantsi, kwaye musa ukuxubha ngamandla kumxube. Ukulungiswa kweesampuli ze-plasma kufuna uncedo lwe-anticoagulants, ngelixa iisampuli ze-serum zingafumaneka kuphela emva kokudibanisa kwendalo ye-serum kunye nokuchithwa kwegazi. I-Diluent nayo inokongezwa kwi-plate hole, kwaye emva koko i-serum specimen yongezwa kuyo, kwaye emva koko ishukunyiswe kwi-oscillator encinci ye-1 ngomzuzu ukuqinisekisa ukuxuba. Ngokubanzi, iisampuli ze-serum ezilinganiswe kwiintsuku ze-5 zingagcinwa kwi-4℃C, kwaye ezo zilinganiswe ngaphezu kweveki kufuneka zigcinwe kwiqhwa elibandayo. Uluhlu olubanzi lweesampulu lungasetyenziselwa ukuzimisela kwe-ELISA. Ulwelo lomzimba (olufana neserum), incindi (amathe) kunye nelindle (elifana nomchamo, ilindle) zonke zinokusetyenziswa njengeisampulu ukumisela i-antibody okanye i-antigen components. Inxalenye yekhithi ye-ELISA engafunekiyo kolu vavanyo kufuneka igcinwe efrijini kwangethuba.