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[China]
Address: 101 and 5th Floor, Building 1, No. 68,18th Road, Guangming Hi-Tech Park, Tangjia Community, Fenghuang Street, Guangming District, Shenzhen 518107 ,China
Contact name:Aimee li
Labnovation Technologies, Inc. |
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Monkeypox Virus Real Time PCR Rapid Test Kit
Intended Use
This kit is used for in vitro qualitative nucleic acid detection of
Monkeypox Virus in Human serum, lesion exudate samples and swab
specimens. The test results of this kit are for clinical reference
only and should not be used as the only standard for clinical
diagnosis.
Monkeypox is a rare zoonotic disease characterized by skin
eruptions caused by the Monkeypox virus (MPV) and whose clinical
symptoms are similar to smallpox. Transmitted mainly through
animals, humans can contract monkeypox from bites from infected
animals or from direct contact with the blood, fluids and rashes of
infected animals, and the virus can also be transmitted from person
to person.
Specification
| Product Name | Monkeypox Virus Real Time PCR Kit |
| Detection Channels | FAM |
| IC | VIC |
| Ct Value | 35 |
| Limit of Detection | 200 Copies/mL |
| Storage | -20℃±5℃ |
| Transportation | Under frozen condition |
| Shelf Life | 12 Months |
| Sample Type | Nasal swabs, oropharyngeal swab, saliva, urine, lesion tissue, exudate, and blood, etc. |
| Package | 24 Tests/Kit,48 Tests/Kit,96 Tests/Kit |
| Components | PCR Reaction Buffer, PCR Enzyme Mixture, Positive control, Negative control |
TEST PRINCIPLE
This kit adopts real time fluorescence PCR technology, which is
suitable for nucleic acid detection of monkeypox virus extracted
from clinical samples such as Human serum, lesion exudate samples
and swab specimens Each reaction system contains specific primers
and fluorescent probes to detect the virus, and the monkeypox virus
nucleic acid can be qualitatively detected by collecting the
fluorescent signal generated by PCR amplification.
In addition, specific primers and fluorescent probes for the
internal standard control for human clinical specimen detection are
added to each reaction system, and the collection, transportation
and extraction process of the sample to be measured is monitored,
indicating the false negative of the test result.
Real-Time PCR System: Molarray MA-6000, ABI 7500, ViiATM 7, QuantStudio 5, QuantStudio 6/7 pro, QuantStudio 6/7 flex, Agilent Mx3000P/3005P, Rotor-GeneTM 6000/Q, Bio-Rad CFX96 TouchTM/iQTM 5,Hongshi SLAN-96S/96P, AGS8830, AGS4800.
COMPONENTS
| Specifications | LQ-000301 24T | LQ-000302 48T | LQ-000303 96T |
| PCR Reaction Buffer | 336μL×1 tube | 672μL×1 tube | 672μL×2 tube |
| PCR Enzyme Mixture | 30μL×1 tube | 50μL×1 tube | 100μL×1 tube |
| Positive control | 50μL×1 tube | 100μL×1 tube | 200μL×1 tube |
| Negative control | 50μL×1 tube | 100μL×1 tube | 200μL×1 tube |
| Instruction for use (pcs) | 1 | 1 | 1 |
PROCEDURES
1. Sample preparations
DNA extraction from clinical samples is conducted according to the
corresponding requirements and procedures in the virus DNA
extraction kit. The extracted
DNA can be directly used for detection. If the sample is not detected immediately after extraction, it should be stored at -70°C for standby, avoiding repeated freeze-thaw cycles.
2. Reaction system preparation
(1) System preparation: Take out the reagent from the kit, and
allow it to thaw completely at room temperature. Invert the mixture
and centrifuge immediately. N test reactions (N = number of samples
to be tested + positive control + negative control + 1) are
prepared for reaction systems, respectively, as follows.
| Components | Volume for 1 reaction system | Volume for N reaction system |
| PCR reaction buffer | 14μL | 14μLx N |
| PCR Enzyme mixture | 1μL | 1μLx N |
| Total volume | 15μL | 15μLx N |
(2) Reaction system distribution: Mix well the above reaction buffer, centrifuge, and dispense 15μL of aliquots into PCR tubes suitable for fluorescent PCR instrument. (Centrifuge the PCR tubes at 6,000rpm for 30s and transport to sample treatment zone.)
3. Sample loading (Sample treatment zone)
Add 10μL nucleic acid sample, negative control and positive control
to the above PCR tubes respectively. Cap the tubes tightly and
centrifuge at 6,000rpm for 10s and then transport to PCR
amplification zone.
* Precaution: Adding samples in the following order is
recommended: negative control-> nucleic acid sample ->
positive control.
4. PCR amplification (PCR amplification zone)
4.1 Put the caped PCR tubes into real-time PCR machine for
amplification.
4.2 Fluorescence PCR cycle condition setting
| Program | Temperature | Reaction duration | Number of cycles |
1 | 50°C | 2 min | 1 |
| 2 | 95°C | 2s | 1 |
| 3 | 95°C | 1s | 41 |
| 60°C | 13s/20s/35s |
Collect fluorescent signals at step 3:60℃; 35s for ABI 7500, 20s for SLAN-96S/96P, while 13s for other Real-Time PCR Systems. The total volume:25μL.
4.3 Disposal after detection
Dispose the PCR tubes in a sealed bag after reaction and treat the
used tubes as medical wastes.
5. Settings for result analysis
Set the baseline at a region before the exponential amplification
where the fluorescent signals of all the samples are relatively
stable (no significant fluctuations in all the samples); set the
starting point (cycle number) away from the signal fluctuations at
the starting
phase of fluorescence collection; set the end point (cycle
number)1~3 cycles before the Ct of the first sample to enter
exponential amplification. 4~15 cycles are recommended.
Set the threshold right above the highest point of the negative
control amplification curve (irregular noise curve).
6. Quality control criteria
Prior to evaluating the specimen results, the Positive Control and
Negative Control should be interpreted using the interpretation
table below, and the Positive Control and Negative Control curve
must be performed correctly, otherwise the sample result is
invalid.
| Channels/ Controls | Cycle threshold(Ct)value | |
| FAM | VIC | |
| Positive control | Ct > 40 or UNDET | Ct > 40 or UNDET |
| Negative control | Ct ≤ 35 | Ct ≤ 35 |
7. Interpretation of Test Results
FAM channels for Monkeypox Virus, detection result should be
interpreted as below.
a) Positive: Ct ≤ 38 and amplification curve is S-shaped.
b) Suspected: 38 < Ct ≤ 40 and amplification curve is S-shaped,
a second test is needed. Consider positive if Ct ≤ 40 and
amplification curve is S-shaped for the second test. Considered
negative if Ct > 40 or null Ct and Ct ≤ 35 in VIC channel for
the second test.
c) Negative: Ct > 40 or null Ct and Ct ≤ 35 in VIC channel.
d) Re-test: Ct > 40 or null Ct and Ct > 35 in VIC channel.
