PRAXISDIENST 243001N-10 NADAL COVID-19 IgG-IgM Test Instruction Manual

PRAXISDIENST 243001N-10 NADAL COVID-19 IgG-IgM Test

Intended Use

The NADAL® COVID-19 IgG/IgM Test is a lateral flow chromatographic immunoassay for the qualitative detection of anti-SARS-CoV-2 IgG and IgM in human whole blood, serum or plasma specimens (see section 12 ‘Limitations’). Note that in the early stages of infection anti-SARS-CoV-2 IgG and IgM may be below the detection limit of the test. This test is intended for use as an aid in the diagnosis of infections with SARS-CoV-2 and the detection of the immune response to COVID-19 infections. The test procedure is not automated and requires no special training or qualification. The NADAL® COVID-19 IgG/IgM Test is designed for professional use only.

Introduction and Clinical Significance

COVID-19 (Corona Virus Disease) is the infectious disease caused by the recently discovered coronavirus SARS-CoV-2. The most common symptoms of COVID-19 are fever, dry cough, fatigue, sputum production, shortness of breath, sore throat and headache. Some patients may have myalgia, chills, nausea, nasal congestion and diarrhoea. These symptoms begin gradually and are mild in most of the cases. Some people become infected but do not develop any symptoms and do not feel unwell. Most people (about 80%) recover from the disease without special treatment. Approximately one in six people who get infected with COVID-19 becomes seriously ill and develops difficulty breathing. Elderly people, and those with pre-existing conditions, such as high blood pressure, heart problems or diabetes, are more likely to develop serious illness. So far, about 2% of infected people have died.
COVID-19 is transmitted via respiratory droplets that are exhaled by infected people via coughing, sneezing or talking. These droplets can be inhaled or ingested directly by other people or can contaminate surfaces, which can then be infectious for several days. Most estimates of the incubation period for COVID-19 range from 1 to 14 days, during which people might already be infectious without showing disease symptoms.

Test Principle

The NADAL® COVID-19 IgG/IgM Test is a lateral flow chromatographic immunoassay for the qualitative detection of anti-SARS-CoV-2 IgG and IgM in human whole blood, serum or plasma specimens.
Anti-human IgM are pre-coated onto the test line region ‘IgM’ and anti-human IgG are pre-coated onto the test line region ‘IgG’ of the membrane. During testing, the specimen reacts with SARS-CoV-2 antigens which are conjugated to coloured particles. The mixture then migrates along the membrane chromatographically by capillary action and reacts with the anti-human IgM and anti-human IgG in the test line region ‘IgM’ and ‘IgG’ of the membrane. The presence of a coloured line in the test line region ‘IgM’ and/or ‘IgG’ indicates a positive result. The absence of a coloured line in the test line region ‘IgM’ and/or ‘IgG’indicates a negative result.
The formation of a coloured line in the control line region ‘C’ serves as a procedural control, indicating that the proper volume of specimen has been added and membrane wicking has occurred.

Reagents and Materials Supplied

• 10 NADAL® COVID-19 IgG/IgM Test cassettes*
• 10 capillary tubes
• 1 buffer (3 mL)**
• 1 package insert
• containing the preservative sodium azide: <0.02%(7.5 ng/test)
  **Phosphate buffer containing the following preservatives: sodium azide: 0.2 mg/mL and kanamycin sulfate: 0.25 g/L

No hazard labelling is required according to Regulation (EC) Nº 1272/2008 CLP. Concentrations are below exemption threshold.

Additional Materials Required

• Specimen collection containers (appropriate for specimen material to be tested)
• Centrifuge (for serum or plasma specimens only)
• Alcohol pads
•  Lancets (for fingerstick whole blood specimens only)
• Timer

Storage & Stability

Test kits should be stored at 2-30°C until the indicated expiry date. Test cassettes are stable until the expiry date printed on the foil pouches. Test cassettes must remain in the sealed foil pouches until use. Do not freeze the test kit. Do not use tests beyond the expiry date indicated on the packaging. Care should be taken to protect test kit components from contamination. Do not use test kit components if there is evidence of microbial contamination or precipitation. Biological contamination of dispensing equipment, containers or reagents can lead to inaccurate results.

Warnings and Precautions

• For professional in-vitro diagnostic use only.
• Carefully read through the test procedure prior to testing.
• Do not use the test beyond the expiration date indicated on the packaging.
• Do not use test kit components if the primary packaging is damaged.
• Tests are for single use only.
•  Do not add specimens to the reaction area (result area).
• In order to avoid contamination, do not touch the reaction area (result area).
• Avoid cross-contamination of specimens by using a new specimen collection container for each specimen obtained.
•  Do not substitute or mix components from different test kits.
•  Do not eat, drink or smoke in the area where specimens and test kits are handled.
•  Wear protective clothing such as laboratory coats, disposable gloves and eye protection when specimens are being assayed.
• Handle all specimens as if they contain infectious agents. Observe established precautions for microbiological risks throughout all procedures and standard guidelines for the appropriate disposal of specimens. does not completely guarantee the absence of transmissible pathogenic agents. It is therefore recommended that these products be treated as potentially infectious and handled in accordance with usual safety precautions (e.g., do not ingest or inhale).
• Temperature can adversely affect test results.
• Used testing materials should be disposed of according to local regulations.

Specimen Collection and Preparation

The NADAL® COVID-19 IgG/IgM Test can be performed using whole blood (from venipuncture or fingerstick), serum or plasma.
To collect fingerstick whole blood specimens:

• Wash the patient’s hand with soap and warm water or clean it with an alcohol pad. Allow it to dry.
•  Massage the hand, without touching the puncture site, by rubbing along the hand towards the fingertip of the middle or ring finger.
• Puncture the skin with a sterile lancet. Wipe away the first drop of blood.
• Gently rub the hand from the wrist to the palm, and then to the finger to form a rounded drop of blood over the puncture site.

Fingerstick whole blood should be tested immediately.
Venipuncture whole blood specimens
Containers containing anticoagulants, such as sodium citrate, potassium citrate, sodium heparin, lithium heparin or sodium oxalate should be used for the preparation of venous whole blood or plasma specimens.
Testing should be performed immediately after specimen collection. Do not leave specimens at room temperature for prolonged periods of time.
If the test is to be run within 2 days of specimen collection, whole blood collected by venipuncture should be stored at 2-8°C.
Do not freeze whole blood specimens.
Serum and plasma specimens
Separate serum or plasma from blood as soon as possible to avoid haemolysis. Use only clear, non-haemolysed specimens.
Testing should be performed immediately after specimen collection. Do not leave specimens at room temperature for prolonged periods of time. Serum and plasma specimens can be stored at 2-8°C for up to 3 days. For long-term storage, specimens should be kept at -20°C.
Bring specimens to room temperature prior to testing. Frozen specimens should be completely thawed and mixed well prior to testing. Specimens should not be frozen and thawed repeatedly.
If specimens are to be shipped, they should be packed in compliance with all applicable regulations for the transportation of etiologic agents.
Icteric, lipemic, haemolysed, heat-treated and contaminated specimens may lead to inaccurate test results.

Test Procedure

Bring tests, specimens, buffer and/or controls to room temperature (15-30°C) prior to testing.

1. Remove the test cassette from the foil pouch and use it as soon as possible. The best results will be obtained if the test is performed immediately after opening the foil pouch. Label the test cassette with the patient or control identification.

2. Place the test cassette on a clean and level surface.

3. Holding the capillary tube vertically, draw the specimen (whole blood/serum/plasma) up to the fill line (approximately 10 μL) and add it to the to the specimen well (S) of the test cassette.
   Alternatively, a micropipette (10 µl) may be used.

4. Holding the buffer bottle vertically, add 2 drops (approximately 80 μL) of buffer to the buffer well (B). Avoid air bubbles forming.

5. Start the timer.

6.  Wait for the coloured line(s) to appear. Read the test result after 10 minutes. Do not interpret the result after more than 20 minutes.

Result Interpretation

Positive for IgM
A coloured line develops in the control line region ‘C’ and another coloured line develops in the test line region ‘IgM’.
Positive for IgG
A coloured line develops in the control line region ‘C’ and another coloured line develops in the test line region ‘IgG’.
Positive for IgG and IgM
In addition to the control line ‘C’, a coloured line develops in the test line region ‘IgM’ and another in the test line region ‘IgG’.

Note: The colour intensity in the test line region ‘IgG’ and ‘IgM’ may vary depending on the concentration of anti-SARS-CoV-2 antibodies in the specimen. Therefore, any shade of colour in the test line region ‘IgG’ or ‘IgM’ should be considered positive. Note that this is a qualitative test only and it cannot determine the analyte concentration in the specimen.

Negative
A coloured line develops in the control line region ‘C’. No lines develop in the test line region ‘IgM’ and ‘IgG’.

Invalid
The control line ‘C’ fails to appear. Results from any test which has not produced a control line at the specified reading time must be discarded. Please review the procedure and repeat the test with a new test cassette. If the problem persists, discontinue using the test kit immediately and contact your distributor.
Insufficient specimen volume, incorrect operating procedure or expired tests are the most likely reasons for control line failure.

Limitations

• The NADAL® COVID-19 IgG/IgM Test is for professional in-vitro diagnostic use only. It should be used for the qualitative detection of anti-SARS-CoV-2 antibodies in human whole blood, serum or plasma specimens only. Neither the quantitative value nor the rate of increase in the concentration of anti-SARS-CoV-2 antibodies can be determined with this qualitative test.

• The NADAL® COVID-19 IgG/IgM Test only detects the presence of anti-SARS-CoV-2 antibodies in specimens and should not be used as the sole criterion for a diagnosis of COVID-19.

• As with all diagnostic tests, all results should be interpreted in conjunction with other clinical information available to the physician.

•  At the beginning of the disease, the concentration of anti-SARS-CoV-2 IgM may be below the detection limit of the test.

• The continued presence or absence of antibodies cannot be used to determine the success or failure of therapy.

•  Results from immunosuppressed patients should be interpreted with caution.

•  A positive test result can also occur in case of negative PCR results because antibodies are still present in the blood after the illness and can be detected.

•  If the test result is negative and clinical symptoms persist, additional testing using other clinical methods is recommended. A negative result does not at any time preclude the possibility of a SARS-CoV-2 infection.

Expected Values

A primary SARS-CoV-2 infection is characterised by the presence of detectable IgM antibodies at the beginning of the disease. The subsequent presence of IgG antibodies can indicate a previous SARS-CoV-2 infection for months – even when the pathogen is no longer detectable by PCR. These IgG antibodies can indicate immunity. However, in some cases antibodies can only be detected in sufficient quantities long after the infection. In case of a sufficient innate or a sufficient cellular immune response, the absence of detectable specific antibodies is also conceivable.

Performance Characteristics

Clinical performance
Diagnostic sensitivity and specificity
The NADAL® COVID-19 IgG/IgM Test was evaluated with clinical specimens whose status was confirmed using RT-PCR. The following table demonstrates the combined results of the IgG and IgM test lines (negative: IgG and IgM; positive: IgM and/or IgG).

• Diagnostic sensitivity: 94.1% (86.8% – 98.1%)*
• Diagnostic specificity: 99.2% (97.7% – 99.8%)*
• Overall agreement: 98.2% (96.6% – 99.2%)
• 95% confidence interval

To illustrate the dependence of sensitivity on the time point of specimen collection, specimens of additional clinical studies are grouped according to the time period between the onset of symptoms and specimen collection (1-14 and 15-35 days) in the following section:

Diagnostic sensitivity IgM:

• 0-14 days: 67.4% (59% – 74.8%)*
• 15-68 days: 95.1% (88% – 98.1%)*

Diagnostic sensitivity IgG:

• 0-14 days: 60.6% (52.1% – 68.5%)*
• 15-68 days: 85.2% (75.9% – 91.3%)*

Diagnostic sensitivity IgM/IgG (combined):

• 0-14 days: 70.5% (62.2% – 77.6%)*
• 15-68 days: 96.3% (89.7% – 98.7%)*

Interfering substances
SARS-CoV-2 IgG/IgM negative and positive serum specimens spiked with the following interfering substances showed no interference with the NADAL® COVID-19 IgG/IgM Test.

• Acetaminophen 200 mg/L
• Acetylsalicylic acid 200 mg/L
• Albumin 20 g/L
• Ascorbic acid 20000 mg/L
• Bilirubin 10000 mg/L
• Caffeine 200 mg/L
• Creatine 2000 mg/L
• Ethanol 1%
• Gentisic acid 200 mg/L

Cross-reactivity
Anti-Influenza virus type A, anti-influenza virus type B, anti-RSV, H1N1, H3N2, H7N9, anti-Haemophilus influenzae, anti-adenovirus, anti-HBsAg, anti- HBV, anti-T. pallidum, anti-H. pylori, anti-HIV, anti-HCV, ANA and HAMA positive specimens were tested using the NADAL® COVID-19 IgG/IgM Test. No cross-reactivity with the specimens was observed when tested using the NADAL® COVID-19 IgG/IgM Test. It is not ruled out that SARS-CoV, MERS-CoV and rheumatoid factor positive specimens may show cross-reactivity with the NADAL® COVID-19 IgG/IgM Test.
Precision
Repeatability and reproducibility
Precision was established by testing 10 replicates of negative and anti-SARS- CoV-2 IgG/IgM positive specimens. Repeatabi-lity was established within the reproducibility study. Testing was performed using 3 independent NADAL® COVID-19 IgG/IgM test lots.
The NADAL® COVID-19 IgG/IgM Test demonstrated acceptable repeatability and reproducibility. The negative and positive values were correctly identified

99% of the time.

References

1. Coronavirus disease 2019 (COVID-19) in the EU/EEA and the UK – ninth update, 23 April 2020. Stockholm: ECDC; 2020.
2.  Weiss SR, Leibowitz JL, Coronavirus pathogenesis, Adv Virus Res 2011; 81:85-164.
3.  Cui J, Li F, Shi ZL, Origin and evolution of pathogenic coronaviruses, Nat Rev Microbiol 2019; 17:181-192.
4. Su S, Wong G, Shi W, et al, Epidemiology, genetic recombination, and pathogenesis of coronaviruses, TrendsMicrobiol 2016;24:490-502.

References

• nal von minden GmbH - Spezialist für hochwertige in-vitro Diagnostik
• nal von minden GmbH - Spezialist für hochwertige in-vitro Diagnostik

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