Monkeypox Virus Detection Kit (RT-PCR)

[PACKAGE SPECIFICATION]

24 Tests/Kit；48 Tests/Kit

[INTENDED USE]

The kit is used for qualitative detection of monkeypox virus nucleic acid in vitro and for clinical auxiliarydiagnosis and treatment.

[INSPECTION PRINCIPLE]

The kit can detect monkeypox virus nucleic acid by designing specific primers and TaqMan probes for theconserved sequence of monkeypox virus specific gene. The kit is designed with a two-color fluorescencedetection system. FAM fluorescence is used to detect monkeypox virus nucleic acid and Cy5 fluorescence is usedto detect internal reference genes.

[MAIN COMPONENTS]

Table 1 Main Components
Number Composition Specifications Composition Description1 PCR reaction solution 120 μL/vial Taq DNA polymerase, dNTP, etc2 Primer probe mixture 240 μL/vial Primers and probes
6 Positive quality control 50 μL/vial plasmid DNA7 Negative quality control 50 μL/vial Purified water
Note:

1. Within the validity period of each component, the components in different batch number kits cannot beexchanged.

2. The quality control material is plasmid DNA containing target gene sequence, which does not haveanybiological activity, so it will not cause harm to human. The above ingredients do not include any biological
materials such as animals, pathogens, human tissues and body fluids, so the safety of the product to users andtheenvironment during transportation and use can be guaranteed. Need but Not Provided Material:
Reagent: commercial nucleic acid extraction kit. Consumables: fluorescent quantitative PCR reaction tube.

[STORAGE CONDITIONS AND VALIDITY PERIOD]

1. The kit is stored and transported at - 35 ℃ to 0 ℃ (± 5 ℃) and is valid for 12 months.
2. The kit is valid within 5 times after repeated freeze-thaw verification. See the product label for the production date and service life.

[APPLICABLE INSTRUMENTS]

ABI 7500 PCR Instrument

[SAMPLE REQUIREMENTS]

1. Test specimen type
(1) Plasma: Collect fresh anticoagulant blood (non-heparin);
(2) Rash Exudate: Wipe the rash exudate with a sterile swab, put it into a sterile test tube (containing 1ml sterilenormal saline), plug the test tube tightly with a sterile cotton ball, and send it for sealed examination. Thespecimen can be immediately used for testing. 2.Extraction of nucleic acid from test specimens
Extract the nucleic acid of the specimen to be tested by using the commercialized nucleic acid extractionkit; Theextracted nucleic acid can be stored at 2 ~ 8 ℃ for no more than 2 days and - 20 ± 5 ℃for no more than180days. [TEST PROCEDURE]
1. Take the nucleic acid of the specimen to be tested and the control substance of Negative and Positive properties, shake and mix well, and centrifuge instantaneously for standby. 2. Thaw the kit at room temperature, shake and mix all components, and centrifuge instantaneously for standby. 3. Calculate the volume of PCR reaction solution and primer probe mixture required according to the number oftest specimens n, plus Negative-Positive control (2) and loss (0.5). Take the following table as an example, configure PCR amplification reaction solution in a new EP tube, mix it upside down and centrifugeinstantaneously. Table 2 Preparation of PCR Amplification Reaction Solution
Component PCR Amplification Reaction SolutionPCR reaction solution 5*（n+2.5）µL
Primer probe mixture 10*（n+2.5）µL
Total volume 15*（n+2.5）µL
4. Pack the PCR amplification reaction solution into the PCR reaction tube according to 15 µL/tube, andadd5µLthe samples prepared in step 1, positive control and negative control respectively. The operation shall be carriedout on ice, press the pipe cover, mix upside down, and centrifuge at 2000 rpm for 10 seconds. 5. Carefully place the reaction tube to be detected in the PCR amplification detector, set the PCRamplificationparameters according to the requirements of the following table, and set the target gene fluorescent reporter groupas FAM fluorescence, the internal reference gene fluorescent reporter group as Cy5 fluorescence, andthequenched fluorescent group as NFQ-MGB. 6. Set the reaction procedure of PCR according to the following steps:
Table 3 PCR amplification cycle parameters
Step gathering Number of cycles Temperature Time Collect Fluorescence Signal
1 1 95℃ 10 min no
2 45
95℃ 15 sec no
58℃ 60 sec yes
7. After the sample is put on the machine, save the file and run the program. 8. ABI 7500 software analysis software threshold value setting: the FAM channel threshold value is set to5000.

[POSITIVE JUDGMENT VALUE AND INTERPRETATION OF TEST RESULTS]

• 
  1. The negative quality control FAM/Cy5 channel has no signal, indicating that there is no pollution inthetest,
  and the result analysis can be continued.
  2. The Ct value of positive quality control FAM/Cy5 channel ≤ 35 indicates that the experimental system is
  normal and the result analysis can be continued.
  3. All test sample reaction holes shall have Cy5 signal, and the CT value of Cy5 signal shall be ≤ 35. If there is no
  Cy5 signal or Cy5 Ct> 35, it is necessary to re-extract the sample nucleic acid and repeat the test.
  4. Detect the FAM channel of the sample. If there is an S-shaped amplification curve and the Ct value is ≤ 35, the
  sample is positive for monkeypox virus nucleic acid detection; If there is no amplification curve or Ct value > 35,
  the sample is negative for monkeypox virus nucleic acid detection.
  [LIMITATIONS OF TEST METHODS]
  1. The test results of this kit are only for clinical reference and should not be used as the basis for the diagnosis of
  patients. Clinicians should comprehensively judge the test results in combination with other diagnostic results.
  2. Negative results can not completely exclude the existence of Monkeypox Virus nucleic acid. Excessive
  degradation of nucleic acid or the concentration of target gene in the amplification reaction system is lower than
  the detection limit can also lead to negative results.
  3. Unreasonable specimen collection, transportation and treatment, as well as improper test operation and
  experimental environment may lead to false negative or false positive results.
  4. This kit is limited to the specified sample type and detection system.

•  
• [PERFORMANCE CHARACTERISTICS]

1. Positive coincidence rate: three enterprises' positive reference materials P1-P3 were tested, and the test results
should be positive, with a positive coincidence rate of 100%.
2. Negative coincidence rate: three negative reference materials N1-N3 of enterprises were tested, and the test
results should be negative, with a negative coincidence rate of 100%.
3. Precision: test the precision quality control products J1-J2 of the enterprise, repeat the test for 10 times, and the
coefficient of variation (CV%) calculated by Ct value CV ≤ 5.0%.
4. Limit of Detection: when the monkeypox virus content is 500 copies/ml, the target gene can be accurately
detected.

[PRECAUTIONS]

1. This kit is only used for in vitro diagnosis. Please read the instructions carefully before use and operate instrict
accordance with the instructions. 2. The operation shall be carried out by personnel with professional experience or qualified training. 3. The laboratory shall be used separately according to reagent preparation area, sample processing area, reactionsolution preparation area and amplification detection and analysis area. During operation, work clothes, hats, shoes, gloves, etc. Should be fully worn. Items in each area shall be used exclusively and shall not be cross usedto avoid pollution. 4. The test results will be affected by the source of the specimen itself, the collection process, specimenquality, transportation conditions, pretreatment and other factors, as well as the quality of nucleic acid extraction, themodel of fluorescent quantitative PCR instrument, the operating environment and the limitations of current
molecular biology technology. As a result, very few specimens may get false positive or false negative test results. Users should understand the potential errors that may exist in the detection process and the limitationsofaccuracy. 5. Before use, the kit shall be fully melted, mixed and centrifuged to make the liquid concentrate at the bottomofthe reagent tube. 6. All reagents in this kit have been specially prepared for the above tests. Replacing any reagent in the kit at will
may affect the use effect. The components of kits of different batch numbers cannot be mixed. 7. Please use this kit within the validity period. Do not use expired kit components. 8. Strictly prevent pollution, and pay attention to prevent leakage when preparing the reaction solution, soastoprevent fluorescent substances from polluting the instrument. 9. The test table shall be wiped with 75% alcohol before and after use, and the workbench andvariousexperimental supplies shall be disinfected regularly.

[INDEX OF SYMBOLS]

• Lot number For in vitro diagnostic use onlyTemperature Limitation Consult instruction for use
• Use by Contains sufficient for <n>tests
• Fragile, Handle With Care Keep dry
• Manufacturer This Way Up
• Authorized Representative in the
• European Community
• Catalogue number
• Date of manufacture CE Marking