Monkeypox Virus MPV Nucleic Acid Detection Kit Real Time PCR Method

【Intended Use】

This kit is used forthe detection of Monkeypox Virus(MPV) in serum or lesion exudate samples by using real
time PCR systems.

【Test Principle】
The principle of the real-time detection is based on the fluorogenic 5’nuclease assay. During the PCR reaction, the DNA polymerase cleaves the probe at the 5’ end and separates the reporter dye from the quencher dye only when the probe hybridizes to the target DNA. This cleavage results in the fluorescent signal generated by the cleaved reporter dye, which is monitored real-time by the PCR detection system. The PCR cycle at which an increase in the fluorescence signal is detected initially is proportional to the amount of the specific PCR product. Monitoring the fluorescence intensities in real-time allows the detection of the accumulating product without having to re-open the reaction tube after the amplification. The Monkeypox Virus real time PCR Kit contains a specific ready-to-use system for the detection of the Monkeypox Virus through polymerase chain reaction (PCR) in the real-time PCR system. The master contains reagents and enzymes for the specific amplification of the Monkeypox Virus DNA. Fluorescence is emitted and measured by the real time systems´ optical unit during the PCR. The detection of amplified Monkeypox Virus DNA fragment is performed in fluorimeter channel FAM with the fluorescent quencher BHQ1. DNA extraction buffer is available in the kit and serum or lesion exudate samples are used for the extraction of the DNA. In addition, the kit contains a system to identify possible PCR inhibition by measuring the Cy5 fluorescence of the internal control (IC). An external positive control defined as 1×10 7copies/ml is supplied which allow the determination of the gene load.

【Components of the Diagnostic Kit】

【Applicable Instrument】
The diagnostic kit is applicable to MA-6000, ABI series, Bio-Rad series, Roche LightCycler R480, Cepheid
SmartCycler, Rotor-Gene series and other multi-channel real-time quantitative PCR instruments.

【Specimen Requirements】
1.Sample types: serum, or lesion exudates samples

2.Specimens can be extracted immediately or frozen at -20°C to -80°C.

3.Transportation of clinical specimens must comply with local regulations for the transport of etiologic agents.

【The Sample Result Judgment 】
1.Ifthe test sample detects a typical S-type amplification curve in the FAM≤38, and Cy5 has a typical amplification curve, theCtis≤38.the samplecanbe judgedtobe MPVpositive.

2.If the test sample has no Ct or Ct > 38 in the FAM channels, and there is a typical S-type amplification curve in the Internal Control channel(Cy5), the Ctis ≤38.the sample canbe judgedtobeMPVnegative.

3.If the test sample have no typical S-type amplification curve (No Ct value) or Ct value> 38 is detected in the FAM, andCy5 channel does not have a typical amplification curve (No Ct value) or Ct value> 38, it means that there is a problem with the quality of the sample or a problem with the operation. If the result is invalid, you should find and eliminate the cause,collectthe sampleagain, andrepeatthe test(ifthe test resultis stillinvalid,please contact the company).

【Limitations of Detection Method】
1.Test results of the diagnos tickit can be used only for clinic reference. The clinical diagnosis and treat ment of patients should be considered in conjunction with their symptoms, signs,medicalhistory and other related conditions.

2.False negative results may occur when the concentration of the detecte dnucleicacid in the test sampleis below the minimumde tection limit of this kit.

3.Improperhandlingofthetestedsampleduringcollection,transportation, storage, andprocessingcaneasily resultin DNA degradation and falsenegative results.

4.When samples are cross-contaminated during collection,transportation, storage, and rocessing,it is easy togetfalse positive results.

【Product Performance Index】
1.LOD:The limitofdetectionis200copies/ml. 2. Precision:Coefficientof variation(CV%)ofprecisionCtvaluewithinbatch≤3%. 3.Specificity:Thereisnocross reactionbetweenthekit andpositive samples, suchasSmallpoxVirus,CowpoxVirus, VacciniaVirus etc.

【Precations】
1.The entire detection process should be performed strictly in accordance with there quirements of this manual in the reagent preparation area, sample processing area, and PCR amplification area, and the experimental clothes,instruments, and consumables ineacharea should be used in dependently and cannotbe mixed.

2.Negative andpositive controls shouldbe setfor each experiment.

3.All reagents in the kit should be fully thawed and mixed at room temperature and centrifuged immediately before use.

4.All negative and positive controls in the kit should be transferred to the sample preparation area and stored separately before the firstuse.

5. To prevent fluorescence interference, avoid touching the PCR reaction tube directly with bare hands, and avoid any marking on the PCR reaction tube.

6. The instrument amplification related parameters should be set in accordance with the relevant requirements of this manual, and differentbatchesof reagents cannotbemixed.

7.The product waste during the experiment should be detoxified before being discarded.

【Index of Symbol】

【FACTORY WORKSHOP】

【EXPORTER】

Magnus Internationa Limited

F12, New City Internationa Mansion A, 234 Huapao Ave.

Liuyang, Hunan Province 410300 China

Contact: Goodwellmedical@gmail.com

【AUTHORIZED REPRESENTATIVE】

Lotus NL B.V.

Koningin Julianaplein 10, 1e Verd, 2595AA, The Hague, Netherlands.

【PRODUCT AND STANDARDS】