nal 870004N-01/870004N-05 NADAL Borreliosis AG Test Instruction Manual

nal 870004N-01/870004N-05 NADAL Borreliosis AG Test

Intended Use

The NADAL® Borreliosis Ag Test is a membrane-based immunoassay for the rapid, direct detection of Borrelia antigens (B. garinii, B. afzelii and B. burgdorferi sensu stricto and B. spielmanii) in ticks. The test is developed for the examination of small to medium-sized ticks (≤8 mm) only. A positive result with this test indicates the presence of Borrelia in the tick.

Introduction and Clinical Significance

Over 800 species of ticks are known worldwide, which can be divided into two main families: the hard ticks and soft ticks. Soft ticks are usually to be found in the tropics and subtropics. Hard ticks, however, including the castor bean tick, are spread around the world and are carriers of Lyme borreliosis disease and tick-borne encephalitis (TBE). While TBE viruses are in the salivary glands of ticks and can be transmitted directly to the the victim through the tick bite, Borrelia are initially located in the midgut of the tick. Borrelia reach the salivary glands while the tick is sucking blood and are transmitted to the victim only after a few hours. Thus, there is no great risk of a Borrelia infection within the first few hours after being bitten by the tick. The excretion of pathogens begins as early as two hours after being bitten by the tick and reaches its peak only after 72 hours.

Symptoms of Lyme borreliosis disease in humans can be divided into three stages.

• Several days to weeks after infection:
  characteristic skin redness, so-called erythema chronicum migrans, appears around the tick bite and spreads into a ring shape. It occurs in about 70 percent of cases and is followed by general flu-like sysmptoms, such as profuse sweating, fever, headaches, and muscular and joint pain.

• A few weeks to months later:
  Inflammation of the nerves and meninges are possible. A strong, burning pain around the original site of the tick bite and redness, which can spread over the whole body, frequently occur. Swelling of joints, as well as impairments of the locomotor system, the psyche and the cognitive abilities may appear. In some cases, paralysis of, for instance, facial muscles and less often inflammation of the heart and heart rhythm disorder occur.

• Months to years after the infection:
  Cases of chronic or recurring inflammation of joints, mostly in the knees and ankles (lyme arthritis) or skin changes, known as acrodermatitis chronica atrophicans herxheimer occur.

Test Principle

The NADAL® Borreliosis Ag Test is a membrane-based immunoassay for the rapid, direct detection of Borrelia antigens in ticks. The test principle involves polyclonal antibodies, which are used as capture antibodies, along with conjugated Borrelia antibodies on the internal test strip to detect Borrelia antigens in the sample. First, the extracted tick material is added to the sample well of the test cassette. The sample migrates along the internal test strip by capillary action. If Borrelia antigens are present in the sample, an antibody-antigen-antibody complex is formed in the test line region and as a result, a coloured line appears. The appearance of a test line indicates a positive result. If the sample does not contain Borrelia antigens, no antibody-antigen-antibody complex is produced and no coloured line develops in the test line region. The test result should be considered negative. The appearance of a coloured line in the control line region serves as a procedural control, indicating that the proper volume of specimen has been added and membrane wicking has occurred.

Reagents and Materials Supplied

• 1/5 NADAL® Borreliosis Ag test cassette(s) (incl. pipette(s) and desiccant)
• 1/5 tick tube(s) containing buffer (0.2 ml)
• 1/5 stick(s)
• 1 tick removal device
• 1 package insert

Additional Materials Required

Timer

Storage & Stability

The test kit should be stored at 2-30°C until the expiry date printed on the packaging. The test cassette should remain in the sealed foil pouch until use. Do not freeze the test kit.

Warnings and Precautions

• Carefully read through the test procedure prior to testing. Failure to follow the instructions will lead to inaccurate test results.
• Do not use the test beyond the expiration date indicated on the package.
• Do not use the test if the foil pouch is damaged.
• Do not reuse tests.
• Do not add samples 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 tick tube 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.
• The test kit contains products of animal origin. Certified knowledge of the origin and/or sanitary state of the animals 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).
• Humidity and temperature can adversely affect test results.
• Used testing materials should be discarded according to local regulations.
• The tick must be crushed thoroughly, as Borrelia are located in the tick’s colon.
• Keep the sample well of the test cassette clear of any tick particles.
• Only use the stick for the purpose described in the package insert.
• mmediately remove the stick from the reagent mixture after crushing the tick.

Specimen Collection and Preparation

Remove the tick carefully and ensure that the head is also removed. The use of the tick removal device is recommended. After removing the tick and before starting the test, you can store the tick in the supplied specimen collection tube. Testing of the tick directly after the removal yields the best results. A thorough crushing of the tick is crucial for optimal testing results, as Borrelia to be verified are located in the tick’s colon.

Test Procedure

The optimal test procedure depends on the size of a tick. The test is developed for the examination of small to medium-sized ticks (≤8 mm) only.

1. Place the tick into the tick tube containing buffer.

2. Crush the tick with the supplied stick until the tick’s colon is crushed and the liquid takes on colour. Use the sharp and the dull sides of the stick for this purpose. Then remove the stick.

3. When ready to perform the test, open the foil pouch and remove the test cassette. Place it on a clean, flat surface.

4. Fill the pipette with the tick-buffer mixture. Ensure that you do not take up any tick particles.

5.  Add the tick-buffer mixture to the sample well of the test cassette. Ensure that you do not add any tick particles to the sample well, as they may obstruct it.
   The liquid will start to migrate along the internal test strip.

6. Read the test result after 10 minutes. Do not interpret the result after more than 20 minutes.

Result Interpretation

Positive:
Two coloured lines appear on the membrane. One line appears in the control line region (C) and the other line appears in the test line region (T). The line in the test line region (T) indicates that Borrelia antigens have been detected in the tick.

The less Borrelia antigens are present in the tick, the weaker the test line is (T). Even a weak test line should be interpreted as a positivie test result. The control line is not a reference line and may have a different colour intensity as the test line.

Negative:
Only one coloured line appears in the control line region (C). No apparent coloured line appears in the test line region (T). No Borrelia antigens have been detected in the tick.

Invalid:
The control line 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 he test with a new test cassette. If the problem persists, discontinue using the test kit immediately and contact your local distributor.

Note: Insufficient specimen volume, incorrect operating procedure or expired tests are the most likely reasons for the control line failure.

Quality Control

A coloured line appearing in the control line region (C) is considered an internal procedural control. It confirms sufficient specimen volume, adequate membrane wicking and correct procedural technique.

Limitations

• The test is developed for the examination of small to medium-sized ticks (≤8 mm) only.
• A positive result with this test indicates the presence of Borrelia in the tick.

13. Performance Characteristics

A comparative study has investigated which Borrelia species in spiked ticks can be detected with the help of the NADAL® Borreliosis Ag Test in comparison with the polymerase chain reaction (PCR). The following Borrelia species: B. garinii, B. afzelii, B. burgdorferi sensu stricto and B. spielmanii were detected with both test systems (comparison study 2012/2013).

The NADAL® Borreliosis Ag Test

• Sensitivity: 92.86%
• Specificity: 95.83%
• Positive predictive value: 86.67%
• Negative predictive value: 97.87%
• Total Test Performance: 95.16%

References

1. J. Eckert, K.T. Friedhoff, H. Zahner, P. Deplazes: „Lehrbuch Der Parasitologie für die Tiermedizin“, Enke Verlag, 2.vollst. überarb. Aufl. 2008 W.
2. Luttmann, K. Bratke, M. Küpper, D. Myrtek: „Der Experimentator Immunologie“, Spektrum Akademischer Verlag, S.110 f., 3. Auflage 2009.

Symbol

• Consult instructions for use
• Temperature limitation
• Batch code
• Do not reuse
• Use by
• Catalogue Number
• Manufacturer
• Sufficient for tests

UK & Ireland:

• Tel: +49 941 290 10-18
• Free Tel –UK: 0808 234 1237
• Free Tel – IRE: 1800 555 080
• Fax: +49 290 10-50

nal von minden GmbH
Carl-Zeiss-Strasse 12 • 47445 Moers • Germany www.nal- vonminden.com • info@nal- vonminden.com Fon: +49 2841 99820-0 • Fax: +49 2841 99820-1

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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