RT-PCR HMANGA COVID-19 TEST DAN TLANGPUI

Covid-19 vei leh vei lo endik nana khawl hman lar em em RT-PCR hi a lam pum chu ‘Real Time Polymerase Chain Reaction’ a ni a. Tunlai khawvela technology pawimawh berte zinga mi a ni hial awm e. PCR (Polymerase Chain Reaction) khawl hi DNA tihpunna (amplify) atana hman ṭhin a ni. DNA chu ‘deoxyribonucleic acid’ a ni a, thil nungte inthlah chhawnna atana information pawimawh ber kengtu a ni. 

PCR hmanga DNA tihpun a nih theih nan hian bawlhlo (chemicals) chi hrang hrang bakah temperature hrang hrang, hunbi neia inthlak hman a ni ṭhin. Real Time-PCR (RT-PCR) bikah hian khawl chhunga thil thleng (reaction) awm chu chawp leh chilhin khawl screen aṭangin emaw computer hmangin emaw a en nghal zel theih a (hei hi PCR pangngai aia a changkanna pawh a ni), hei vang hian a hmingah pawh ‘Real Time’ tih telh a ni. 

RT-PCR khawl hmangaCOVID-19 en dan

1. Sample lak: RT-PCR khawl hmanga sample endik tur hi uluk taka lak a ngai a. COVID-19 hi thawkna kawng (respiratory tract) natna a ni a. A hrik SARS-CoV-2 hi hnar, ka leh mit aṭang te a mihring taksaa lut thei a ni. Chuvang chuan sample endik tur chu hnar chhung emaw hrawk emaw aṭanga lak a ni thin. Hnar chhung leh hrawkah swab (plastic hmawlh tawite, a hmawra tuihnang hip bet theiin a tuam) in an han hawlh a, hei hian hnar chhung leh hrawk a tui huh/ hnang chu a han hip a, sample burah dah luh nghal a ni. Sample burah hian virus chhe tur venna tui a awm a (viral transport medium), vur bawmah temperature hniam, 2-8°C velah vawn ṭhat nghal a ni a, a rang thei ang berin endikna hmun (laboratory) panpui tur a ni. Sample hi hun rei zawk chhung vawn ṭhat a ngai a nih chuan temperature hniam takah dah a ngai thung. 

2. RNA lak chhuah: Sample aṭanga RNA (Ribonuclei acid) awm zawng zawng chu lak chhuah hmasak a ni phawt a. DNA ni lova RNA lak chhuah a nih zawk chhan chu SARS-CoV-2 te hian ‘genetic material’ (inthlahchhawn nana an hman) atan RNA zawk an neih vang a ni (DNA an nei ve lo tihna nih chu!). SARS-CoV-2 RNA hi tuamtu kawr thau leh thil dang chhungah a awm a. RNA lak chhuah nan hian hemi kawr hi tihchhiat phawt a ṭul. RNA lak chhuah nan hian bawlhlo (chemicals) leh hmanraw hrang hrang engemawzat a ngai a. A khawih chhung a rei duhin fimkhur a ngai hle. RNA lak chhuahah hian virus-in a va behchhanna mihring timura RNA pawh a tel. Mihring RNA hi test-na step-a felhlel awm thei venpuitu (control) atan hman ṭangkai a ni ṭhin. 

3. RT-PCR: RT-PCR khawl chhungah hian thil thleng (reactions) lian tak hlawm hnih a awm a: 

(i) RNA hmanga DNA siam: 

SARS-CoV-2-in nucleic acid a neih hi RNA zawk a nih avangin he an RNA hmang hian DNA siam chhuah phawt a ngai a. SARS-CoV-2 chauha 'RNA sequence' awm, SARS dang leh virus dangtea awm ve lo (specific segment) chauh te chu  entawnin DNA hi siam a ni. [He DNA insiam hi RNA entawn chunga siam a nih avangin cDNA (complimentary DNA) tih kher a ni ṭhin]. cDNA siam nan hian enzyme pawimawh em em ‘reverse transcriptase’ hman a ni a, chuvang chuan he step pawh hi Reverse Transcription PCR tiin an sawi bawk ṭhin. Tin, SARS-CoV-2 RNA chhunga kan duh lai te thlan chhuah nan hian 'primer' hman a ni a. Primer chu RNA kan entawn tur lai te chauh mil tura siam nucleic acid sequence tawite a ni. 

Virus RNA chu DNA a siam a ni ang tho hian sample-a mihring RNA awm chu primer bik hmanga DNA a siam a ni ve a, control atana hman ṭhin a ni. Sample positive tur chuan virus RNA aṭangin DNA hi a insiam ngei ngei a ngai a. SARS-CoV-2 hrik kai lo a nih chuan sample-ah chuan virus RNA a tel lova, chuvang chuan virus RNA aṭanga DNA insiam tur a awm lovang. 

(ii) DNA tihpun: RT-PCR hian SARS-CoV-2 DNA (RNA hmanga siam) chu a tipung a. DNA a tihpun rual hian RT-PCR khawl screen emaw computer emaw aṭanga hmuh theih nghal turin signal a lo lang ṭhin. DNA insiam pung vak a awm chuan sample chu SARS-CoV-2 kai (positive) a ni tih a lantir. Hemi signal a rawn lan theihna tur hian bawlhlo pawimawh tak 'flourescent probe' (thim zinga eng thei rawng) hman a ni a. Hemi probe hian DNA a insiam pun apiangin signal a pe chhuak ta ṭhin a ni. DNA insiam a tam poh leh signal a sang ṭhin. 

Test kalpui dan

RT-PCR khawla sample dah luh nan hian 96-well plate hman a ni a. He plate hi kil li nei, kutphah tiat vela hlai (rectangle), khuar (well) te reuh te te 96 nei a ni. A khuarah hian tui 0.2 ml (200µl) vel lek a leng a. Khuar hi 96 awm mah se, khuar zawng zawng hi sample-in hnawh khah a ni lo. RT-PCR in hna a thawh chhunga felhlel leh dik lo awm thei hrang hrang venpui turin khuar ṭhenkhat hi sample ni lo thildang hmanga hnawh khah a ni. 

Step1: Screening

Screening atan hian pooled testing hman a ni. Khuar pakhat zelah hian sample panga, mihring panga ami chhunkhawm a ni a, chung sample chhunkhawm te result chu a hlawmin ṭum khatah RT-PCR khawlin a rawn pe chhuak ṭhin. Result hi a negative a nih chuan sample pangate chu an negative vek tihna a ni a. A positive erawh chuan sample panga zingah hian positive an awm tihna a ni thung. 

Step 2: Validation (ennawn lehna)

Screening-a positive awmna khuar hi a bika ennawn leh ṭhin a ni a. Sample panga inpawlh aṭang khan tu ber nge positive tih finfiah a ni. Hemi atan hian sample hi inpawlh (pool) tawh lovin khuar pakhat zelah a mal malin RT-PCR khawlah ennawn leh a ni ṭhin. 

Test-naa harsatna awm theite

1. RNA lak chhuah hi sample tam thamah chuan hun leh tha sen a hautak duh hle. (Automated RNA extractor thar hian thawktute hun leh tha senral nasa taka a chhawk beisei a ni)

2. Screening-ah positive an tam chuan validation tur a tam zel a, a mal mala tihnawn leh vek a ngaih ṭhin avangin sample khawih chhung a rei phah. 

3. Sample lak hi uluk viau a ngai a, sample quality a ṭhat loh chuan RNA lak chhuahah harsatna a thlen thei. 

Note: A tlangpuiin plate pakhata khuar 92 hi sample chhunkhawm (pooled sample) dah nan hman ṭhin a ni. Sample en tur hi viral target (SARS-CoV-2 RNA hmun hnih a mi, ‘primer’) pahnih zela kalpui a nih chuan pooled sample 46 a dah theih tihna a ni a, a bak khuar 4 te hi chu venpuitu atan hman a ni. 

Pooled sample pakhatah sample 5 chhun khawm ni ta se, sample 46 dah luh a nih chuan vawi khat test-ah mimal sample 230 a test theih dawn tihna a ni. Sample chhunkhawmte zingah hian negative an tam chuan pooled testing hi a hlawk hle a, positive an tam erawh chuan a mal te te a ennawn leh vek a ngaih avangin test chhung a rei a, khuar pakhata sample pakhat test nen hun hman rei chhung a inthlau em em chuang lo thei tho a ni.