SARS-CoV-2 Neutralizing Antibody Detection Kit (ELISA)
【MOLAOMOTHEO】
SARS-CoV-2 Neutralizing Antibody Detection Kit e ipapisitse le mokhoa oa tlholisano oa ELISA.
Ho sebelisa purified receptor binding domain (RBD), protheine e tsoang ho viral spike (S) protheine le sele e amohelang.
receptor ACE2, tlhahlobo ena e etselitsoe ho etsisa tšebelisano ea kokoana-hloko ea ho se nke lehlakore.
Li-Calibrator, Litaolo tsa Boleng, le lisampole tsa serum kapa plasma li tsoakoa ka bonngoe hantle ka ho hlapolloa
buffer e nang le haACE2-HRP conjugate e qotsitsoeng ka li-tubes tse nyane. Ebe metsoako e fetisetsoa ka hare
liliba tsa microplate tse nang le sekhechana sa SARS-CoV-2 RBD (RBD) sa immobilized recombinant.
ho qhomisa. Nakong ea ho futhumala ha metsotso e 30, antibody e khethehileng ea RBD ka har'a li-calibrators, QC le
lisampole li tla qothisana lehlokoa le haACE2-HRP bakeng sa ho tlama RBD e sa tsamaeeng lilibeng. Kamora
ho qhoqhoa, liliba li hlatsuoa ka makhetlo a 4 ho tlosa conjugate e sa tlamisoang ea haACE2-HRP. Tharollo ea
Joale TMB e ea eketsoa le ho kenngoa metsotso e 20 mocheso oa kamore, e leng se etsang hore ho be le tsoelo-pele ea
mmala o moputsoa. Tsoelo-pele ea 'mala e emisoa ka ho eketsoa ha 1N HCl,' me monyetla o
e lekantsoe spectrophotometrically ho 450 nm. Matla a 'mala o entsoeng a lekana le
palo ea enzyme e teng, 'me e amana ka tsela e fapaneng le palo ea litekanyetso tse lekantsoeng ka mokhoa o ts'oanang.
Ka ho bapisa le lekhalo la calibration le entsoeng ke li-calibrator tse fanoeng, mahloriso a
Li-antibodies ho sampole tse sa tsejoeng lia baloa.
【LITLHAKISO TSE HLOKAHANG EMPA HA LI ABONE】
1. Metsi a hloekisitsoeng kapa a hloekisitsoeng
2. Lipeipi tse nepahetseng: 10μL, 100μL, 200μL le 1 mL
3. Litlhahiso tsa pipette tse lahliloeng
4. Sebali sa Microplate se khonang ho bala monyetla ho 450nm.
5. Pampiri e monyang
6. Pampiri ea kerafo
7. Vortex mixer kapa e lekanang
【POKETSO LE POLOKO】
1. Lisampole tsa Serum le Plasma tse bokeletsoeng ka har'a li-tubes tse nang le K2-EDTA li ka sebelisoa bakeng sa khithi ena.
2. Mehlala e lokela ho koaheloa 'me e ka bolokoa ho fihlela lihora tse 48 ho 2 °C - 8 °C pele ho tekolo.
Mehlala e tšoaretsoeng nako e telele (ho fihlela likhoeling tse 6) e lokela ho hoamisoa hanngoe feela ho -20 °C pele ho tlhahlobo.
Qoba makhetlo a mangata a ho qhibiliha a leqhoa.
TS'ELISO
【Reagent Tokiso】
1. Li-reagents tsohle li tlameha ho ntšoa ka sehatsetsing ebe li lumelloa ho khutlela mocheso oa kamore pele li sebelisoa
(20° ho isa ho 25°C). Boloka li-reagents tsohle ka sehatsetsing hang ka mor'a hore u li sebelise.
2. Lisampole tsohle le litsamaiso li lokela ho ba vortexed pele li sebelisoa.
3. hACE2-HRP Tokiso ea Tharollo: Hlakola hACE2-HRP concentrate ho 1: 51 dilution ratio with Dilution
Buffer. Mohlala, hlapolla 100 μL ea haACE2-HRP e tsepamisitsoeng maikutlo ka 5.0mL ea HRP Dilution Buffer ho
etsa tharollo ea haACE2-HRP.
4. 1× Tokiso ea Tharollo ea Hlatsoa: Hlatsoa Tharollo ea 20× Hlatsoa ka metsi a hloekisitsoeng kapa a qhibilihisitsoeng ka metsi.
karo-karolelano ea bophahamo ba 1:19. Mohlala, hlapolla 20 mL ea 20× Hlatsoa Tharollo ka 380 mL ea deionized kapa
metsi a halikiloeng ho etsa 400 mL ea 1× Hlatsoa Tharollo.
【Mokhoa oa Teko】
1. Ka li-tubes tse arohaneng, aliquot 120μL ea Tharollo e lokiselitsoeng ea hACE2-HRP.
2. Eketsa 6 μL ea li-calibrator, lisampole tse sa tsejoeng, lisebelisoa tsa boleng ka har'a tube e 'ngoe le e' ngoe 'me u kopanye hantle.
3. Fetisetsa 100μL ea motsoako o mong le o mong o lokisitsoeng mohatong oa 2 ho liliba tse tsamaellanang tsa microplate ho latela
ho tlhophiso e reriloeng esale pele ea tlhahlobo.
3. Koahela poleiti ka Plate Sealer 'me u e kene ka 37°C metsotso e 30.
4. Tlosa Plate Sealer 'me u hlatsoe poleiti ka hoo e ka bang 300 μL ea 1× Hlatsoa Tharollo selibeng ka makhetlo a mane.
5. Tlanya poleiti holim'a thaole ea pampiri ho tlosa mokelikeli o setseng ka lilibeng ka mor'a ho hlatsoa mehato.
6. Kenya 100 μL ea Tharollo ea TMB selibeng se seng le se seng 'me u etsetse poleiti lefifing ka 20 - 25°C metsotso e 20.
7. Kenya 50 μL ea Stop Solution selibeng ka seng ho emisa karabelo.
8. Bala ho monyela ha sebali sa microplate ho 450 nm nakong ea metsotso e 10 (630nm joalo ka sehlomathiso).
kgothaletswa bakeng sa tshebetso e nepahetseng e phahameng).