GeneFinder™ COVID-19 Plus Real Amp Kit

GeneFinder™ COVID-19 Plus Real Amp Kit

Manufacturer:

OSANG Healthcare Co., Ltd

132, Anyangcheondong-ro, Dongan-gu, Anyang-si, Gyeonggi-do, 14040 Korea

Tel: +82-31-460-9937

Fax: +82-31-460-9933

1. Intended Use

GeneFinder ™ COVID-19 PLUS RealAmp Kit is used for detection of COVID-19 (COIVD-19) virus through reverse Transcription and Real-Time Polymerase Chain Reaction from RNA extracted from Respiratory specimens such as Alveolar lavage fluid, throat swab, sputum. This product can qualitatively detect COVID-19 using Polymerase Chain Reaction.

2. Principle of the Assay

One-Step Reverse Transcription Real-Time polymerase chain reaction is used to confirm the presence of COVID-19 by amplification of RdRp, E and N gene

This product is a In vitro diagnostics (IVD) and is used by professionals in hospitals and laboratories.

3. Kit Contents

Reagents / Materials100 tests/Kit
COVID-19 PLUS Reaction Mixture1,050 ul
COVID-19 PLUS Probe Mixture550 ul
COVID-19 PLUS Positive Control50 ul
COVID-19 PLUS Negative Control50 ul

4. Storage and Handing Requirements

  • All components of the kit should be stored at -20°C or below and kept stable until the expiry date stated on the label.
  • The COVID-19 PLUS Probe Mixture must be stored below -20°C and in the dark.
  • Expires 12 months after date of manufacture. Do not use after expiration date.
  • Expires 6 months after opening the kit. Do not use after use life time.
  • Store the rest of the kit below -20°C.
  • If the kit is defective, do not use.
  • Dispose of unused reagents and waste in accordance with country, federal, state and local regulations.

Note: Inaccurate results can be obtained if the kit is stored at room temperature for a long period of time.

Note: Unnecessary repeated freezing and thawing lead to inaccurate results.

  • Product Description
  1. COVID-19 PLUS Reaction Mixture

COVID-19 PLUS reaction mixture with reagents for reverse transcription and amplification

  • COVID-19 PLUS Probe Mixture

Buffer solution with specific primers and probes which conjugate with nucleic acids of COVID-19 virus and internal control

  • COVID-19 PLUS Positive Control

Positive Control determines for presence of errors/contamination during the test.

Caution. Care should also be taken to avoid cross-contamination of other samples when adding PositiveControl.

  • COVID-19 PLUS Negative Control

To confirm the absence of contamination, negative control reaction should be included at every run as it indicates that reagents have not been contaminated.

Material no. MFK-45

* The product allows the accomplishment of 100 tests, including controls.

6. Required materials

6.1. Provided in the product

LabelCap colorStorageQuantity
COVID-19 PLUS Reaction MixturePurple-20℃1 x 1,050 ul
COVID-19 PLUS Probe MixtureBrown-20℃1 x 550 ul
COVID-19 PLUS Positive ControlRed-20℃1 x 50 ul
COVID-19 PLUS Negative ControlGreen-20℃1 x 50 ul

(100 tests / Kit)

6.2. Required but not provided in the product

  • Applied Biosystems®  7500 / 7500 Fast Real Time PCR Instrument System and CFX96 real time PCR system.
  • Pipettes (1- 20 μl, 20-200 μl, 200-1,000 μl)
  • Pipettes tips with aerosol barrier (RNase, DNase-free)
  • Powder-free gloves (disposable)
  • Vortex mixer or equivalent
  • 1.5 ml tube
  • PCR tube or 96 well plate
  • Bench microcentrifuge
  • RNA isolation kit (Use of QIAamp Viral RNA Mini Kit (Cat. # 52904, Qiagen) is recommended or commercial kits )

7. Warning and Precaution

The GeneFinder™ COVID-19 Plus RealAmp Kit is designed for In vitro diagnostics .

General warnings and precautions

  • Read the instructions in the package carefully before processing samples.
  • Use 0.5% v / v sodium hypochlorite or another disinfectant to clean and disinfect the area around the sample.
  • Decontaminate and dispose of all specimens, reagents and other potentially contaminated materials in accordance with local regulations.
  • Use universal precautions when performing the assay. Handle samples as if capable of transmitting infection.
  • Wear personal protective apparel, including disposable gloves, throughout the assay procedure. Thoroughly wash hands after removing gloves, and dispose of gloves as biohazardous wastes.
  • The material that come into contact with the biological samples must be autoclaved for one hour at 120℃ before disposal.
  • Do not eat, drink, smoke, or apply cosmetics in areas where reagents of samples are handled.
  • Do not pipet by mouth.
  • Do not use a kit after its expiration date.
  • Use aerosol-resistant pipette tips and use a new tip every time a volume is dispensed.
  • Store the reagents recommended temperature.
  • Do not mix reagent from different batches of the kit.
  • Store the kit away from any source of contaminating DNA, especially amplified nucleic acid.
  • Use sterile disposable laboratory materials and do not re-use the tubes and tips.

Material no. MFK-45

  • Alterations in the physical appearance of kit components may indicate instability or deterioration.
  • Use all pipetting devices and instruments with care and follow the manufacturer’s instructions for calibration and quality control.
  • Do not modify the reagent/sample volume used in the test or use in a wrong way which is not recommended.
  • Store COVID-19 PLUS Probe Mixture at -20℃ in a dark place.
  • Procedure

8.1 Preparation of sample

The GeneFinder™ COVID-19 PLUS RealAmp Kit must be used with RNA extracted from Alveolar lavage fluid, throat swab, and sputum samples. RNA extraction is recommended from sample as soon as possible for accurate experiments.

8.1 RNA Extraction

Commercialized extraction kit should be used for collection of RNA Extracted samples. QIAamp viral RNA Mini Kit

(Qiagen, Germany, Cat. # 52904) is recommended for extraction. Please carry out RNA extraction according to the

manufacturer’s instructions.

Extracted RNA samples are more vulnerable than DNA that it is suggested to avoid repeated freezing and thawing and to keep at -70°C.

When extracting RNA, be sure to extract it according to the manufacturer’s instructions.

8.2 Preparation of reagents

Thaw all components thoroughly at room temperature before using. Mix gently, spin down the contents for 5 seconds, and then test it immediately..

  1. Mix 10 μl of COVID-19 PLUS Reaction Mixture, 5 μl of COVID-19 PLUS Probe Mixture to prepare RT-PCR Master mixture as described in the following table (Table 1). Prepare enough volume of Master mixture for all the reactions plus extra amounts to prevent possible pipetting error.

Note: Total Master mixture number

= n sample + 1 positive control + 1 negative control + 1 extra

Number of Samples Solution1 TEST 3 TEST 5 TESTTotal volume
of Master Mixture
COVID-19 PLUS Reaction Mixture 10 ul30 ul 50 ul10x(n+3)
COVID-19 PLUS Probe Mixture 5 ul15 ul25 ul 5x(n+3)
Total (COVID-19 PLUS Master Mixture)15 ul45 ul 75 ul20x(n+3)

Table 1. Master Mixture preparation

Important: Adequate controls should be used in each run to ensure reliable results.

  • Place 15 μl of RT-PCR Master mixture into each PCR tube or optical 96 well plate Note: To avoid any bubbles, do not vortex the tubes at this step.

Material no. MFK-45

  • Add each of 5 μl of sample RNA into the corresponding PCR tube/well-plate for amplification and mix them with pipetting
  • Place 5 μl of Positive Control and Negative Control into the each PCR tube or well-plate in the same way. Note: Total Reaction volume is 20 μl per sample.

Note: Insufficient mixing of the Master mixture may lead to inaccurate result..

  • Transfer PCR tube or well-plate for testing into the real-time thermal cycler and start the thermal cycle for the amplification.

Figure 1. Schematic workflow for test

8.3 Setting of the RealTime Amplificatin

8.4 Data analysis

  1. Select Amplification Plot at Analysis Mode.
  • Select Analysis Settings.
  • Set Threshold Values, Baseline start and end values.

Material no. MFK-45

  • *When target RNA is detected in a sample amplification reaction, Internal control (IC) may give the result as Ct Not applicable (N/A). In fact, low-efficiency amplification reaction for internal control may be displaced by competition from high-efficiency amplification reaction for Target gene. In such a case, it shall be determined as positive.
  • If test result is not valid as follows, it is recommended to retest.
  • In case of Ct value of Internal control is Not applicable
  • In case of Invalid result.

10. Quality Control

Validating the whole analysis procedure (of each extraction and amplification session by processing a negative tested sample and a positive tested sample or a calibrated reference material) is recommended.

  1. Procedure Limitation
  • The users must be trained and familiar with this technology prior to the use of this device.
  • Any diagnostic results generated must be interpreted in conjunction with other clinical or laboratory findings. It is the user’s responsibility to validate system performance for any procedures used in their laboratory which are not covered by the OSANG Healthcare performance studies.
  • Use this product only with RNA extracted from the following human biological samples: Alveolar lavage fluid, throat swab, sputum
  • A negative result does not exclude the possibility of infection, because results are dependent on appropriate specimen collection and absence of inhibitors. The presence of PCR inhibitors may cause invalid results with this product..
  1. Trouble shooting

13. Performance characteristics

Analytical Sensitivity : LOD

Analytical sensitivity with Target RNA was performed with GeneFinder™ COVID-19 PLUS RealAmp Kit. As a result of experiments at various concentrations, the analytical sensitivity of the product was confirmed as 10 copies

  • reaction for all target genes (RdRp, E, N gene).

Analytical Specificity : Cross Reactivity

The cross reactivity tests were performed using COVID-19 Standard materials and RNA of 14 negative reference materials; None of negative reference materials were detected in any of the test performed.

#Name
1Influenza A (H1N1/09)
2Influenza A (H3N2)
3Influenza A (H5N1)
4Influenza B
5Rhinovirus
6Respiratory syncytial virus (A/B)
7Parainfluenza 1 virus
8Parainfluenza 2 virus
9Parainfluenza 3 virus
10Parainfluenza 4 virus
11Adenovirus
12Human Bocavirus
13Measles virus
14Mycoplasma spp.

Reproducibility (Between Lots, operators, and places)

Reproducibility between lots, inspectors, and test sites with different manufacturing dates for the same concentration of standard material was found to be within the CV% titration range (<5 CV%) and showed reproducibility.

Repeatability

Repeated every 2 days, the test is carried out by replication; all results are taken as the reference value (5% sustained CV basis).

Interference

Mucin, NaCl, Blood, Respiratory syncytial virus A, and PBS were repeated four times for each of high, medium and low concentrations. As a result of testing with RNA extracted from swab containing interfering substances, it was confirmed that some substances above a certain concentration act as interfering agents in reverse transcription and PCR reactions. At the appropriate concentration, all showed positive test results of 100% and confirmed that they were not affected by the interference materials.

14. Symbols used on Labels

Lot or batch numberCaution
Catalogue numberStore below temperature shown
In Vitro Diagnostic Medical DeviceExpiry date
Consult Instruction For UseManufacturer