Ezmedtest RF IgM Test (Serum/Plasma)

INTENDED USE

The IgM rapid Test Cassette (Serum/Plasma) is a lateral flow chromatographic immunoassay for the qualitative detection of IgM antibodies in human serum to Rheumatoid Factor (RF) antigen. It is intended to be used as a screening test and as an aid in the diagnosis of rheumatoid arthritis. Any reactive specimen with the RF IgM Rapid Test Cassette must be confirmed with alternative testing method(s) and clinical findings.

SUMMARY AND EXPLANATION

  • Rheumatoid arthritis (RA) is a chronic inflammatory disease of unknown etiology.
  • 1 Rheumatoid arthritis is a systemic disease characterized by chronic proliferation and inflammation of joint cartilage and supporting structures. RA is mainly defined by clinical criteria, in which systematic pathogenetic studies have been hampered by doubts about the presence of common pathogenetic mechanisms and the relative lack of unique laboratory findings.
  • 2 IgG rheumatoid factor has been reported to be present in sera of patients with rheumatoid arthritis both with and without IgM rheumatoid factor activity.
  • 3,4 RFs are immunoglobulins of any isotype with antibody activity directed against antigenic sites on the Fc portion of IgG molecules. Because of its pentavalent structure and ability to cross-link immunoglobulin G antigen, IgM-RF is the main isotype identified by clinically available diagnostic assays for RF detection.
  • 5 Rheumatoid factors may exist as the mu, gamma, alpha, and epsilon isotypes.
  • 6 Rheumatoid factors are found in 1 to 4% of the general population. RFs are present in 75% of adult patients with the highest incidence of rheumatoid factors occurring in persons over 65 years of age and nearly all patients with Felty and Sjogren syndrome. The clinical correlation of an elevated rheumatoid factor should be interpreted cautiously. Increased titers may accompany a variety of acute immune responses, particularly viral infections and a number of other diseases (e.g., infectious mononucleosis, tuberculosis, leprosy, various parasitic diseases, liver disease, sarcoidosis, and lymphoproliferative syndromes).
  • The earliest tests and those still most widely used rely on the agglutinating properties of the IgM class of rheumatoid factors. Sensitized sheep red blood cell and latex agglutination tests have been developed and routinely employed. These assays are most sensitive for the detection of RF that is of the IgM isotype because of its multivalent structure. These tests provide a dilution which is difficult to standardize and have laborious processing and poor reproducibility.

PRINCIPLE

The RF IgM Rapid Test Cassette (Serum/Plasma) is a lateral flow chromatographic immunoassay based on the principle of the capture technique. The test cassette consists of: 1) a burgundy colored conjugate pad containing human IgG antigens conjugated with colloidal gold (IgG conjugates) and rabbit IgG-gold conjugates, 2) a nitrocellulose membrane strip containing a test band (T band) and a control band (C band). The T band is pre-coated with non-conjugated anti-human IgM antibody, and the C band is pre-coated with goat anti-rabbit IgG.

When an adequate volume of test specimen is dispensed into the sample well of the cassette, the specimen migrates by capillary action across the cassette. The RF IgM if present in the specimen will bind to the IgG conjugates. The immunocomplex is then captured on the membrane by the pre-coated anti-human IgM, forming a burgundy colored T band, indicating a RF IgM positive test result.

Absence of the T band suggests a negative result. The test contains an internal control (C band) which should exhibit a burgundy colored band of the immunocomplex of goat anti-rabbit IgG/rabbit IgG-gold conjugate regardless the presence of any antibodies to H.pylori. Otherwise, the test result is invalid and the specimen must be retested with another device.

MATERIALS PROVIDED

  1. Test Cassette
  2. Disposable Specimen Droppers
  3. Desiccant
  4. Buffer
  5. Package Insert

MATERIALS NOT PROVIDED

  1. Specimen collection container
  2. Centrifuge (for plasma only)
  3. Timer

STORAGE AND STABILITY

The kit can be stored at room temperature or refrigerated (2-30°C). The test device is stable through the expiration date printed on the sealed pouch. The test device must remain in the sealed pouch until use. DO NOT FREEZE. Do not use beyond the expiration date.

WARNINGS AND PRECAUTIONS

  1. For professional in vitro diagnostic use only. Do not use beyond expiration date.
  2. Do not eat, drink or smoke in the area where the specimens or kits are handled.
  3. Handle all specimens as if they contain infectious agents.Observe established precautions against micro-biological hazards throughout all procedures and follow the standard procedures for proper disposal of specimens.
  4. Wear protective clothing such as laboratory coats, disposable gloves and eye protection when specimens are assayed.
  5. Humidity and temperature can adversely affect test results.
  6. Do not perform the test in a room with strong air flow, ie. electric fan or strong airconditioning.

SPECIMEN COLLECTION

  1. RF IgM Rapid Test Cassette (Serum/Plasma) can be performed using either serum or plasma.
  2. Separate the serum or plasma from blood as soon as possible to avoid haemolysis. Only clear, non-haemolysed specimens can be used.
  3. Testing should be performed immediately after the specimens have been collected. Do not leave the specimens at room temperature for prolonged periods.specimens may be stored at 2-8℃ for up to 3 days. For long term storage, specimens should be kept below -20℃.
  4. Bring specimens to room temperature prior to testing. Frozen specimens must be completely thawed and mixed well prior to testing. Specimens should not be frozen and thawed repeatedly.
  5. If specimens are to be shipped ,they should be packed in compliance with usual regulations for transportation of aetiological agents.

TEST PROCEDURE

Allow the test cassette, specimen,buffer, and/or controls to equilibrate 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. Best results will be obtained if the assay is performed within one hour.
  2. Place the test cassette on a clean and level surface. Be sure to label the device with specimen’s ID number.
  3. With a 5 μL mini plastic dropper provided, draw serum or plasma specimen to exceed the specimen line as showed in the following image and then transfer drawn specimen into the sample well ( S ). Then add 2 drops (about μL) of buffer to the buffer well ( B ) immediately and start the timer. Avoid trapping air bubbles in the specimen well (S). Please see the illustration below.

Note: Practice a few times prior to testing if you are not familiar with the mini dropper. For better precision, transfer specimen by pipette capable to deliver 5 μL of volume.

  1. Wait for the red line(s) to appear. The result should be read within 10 minutes. Do not interpret the result afte

5 μL of serum/plasma
2 drops of buffer 12
Buffer
Specimen Line 9 3
B ID RF B ID RF 6
s IgM s IgM 10 Minutes
C C C C C
T T T T T
Positive Negative Invalid

RF IgM Rapid Test Cassette (Serum/Plasma)

INTERPRETATION OF RESULTS

POSITIVE: Two distinct red lines appear. One line should be in the control region (C) and another line should be in the test region (T).

NEGATIVE: One red line appears in the control region (C). No apparent red or pink line appears in the test region (T).

INVALID: Control line fails to appear. Insufficient specimen volume or incorrect procedural techniques are the most likely reasons for control line failure. Review the procedure and repeat the test with a new test Strip. If the problem persists, discontinue using the test kit immediately and contact your local distributor.

QUALITY CONTROL

  • A procedural control is included in the test. A red line appearing in the control region (C) is the internal procedural
  • control. It confirms sufficient specimen volume and correct procedural technique.
  • Control standards are not supplied with this kit; however it is recommended that positive and negative controls be
  • tested as a good laboratory practice to confirm the test procedure and to verify proper test performance.

LIMITATIONS

  1. Only if test instructions are rigidly followed will optimum results be achieved.
  2. Though the RF IgM Rapid Test Cassette is a reliable screening assay, diagnosis must not be based solely on the result of this test but should be complemented with others (ie RF-Waaler test) along with the clinical examination.
  3. The RF IgM Rapid Test Cassette (Serum/Plasma) is limited to the qualitative detection of IgM antibodies to RF in human serum or plasma. The intensity of the test band does not have linear correlation with RF IgM titer in the specimen.
  4. It is occasionally found in the serum of patients with polyartheritis nodosa systematic lupus erythematosus, and a variety of chronic inflammatory illness such as tuberculosis, leprosy, syphilis, and bacterial endocarditis. Sera tested from these related diseases showed positive reactions in approximately 6% of tested cases.
  5. The values obtained from this assay are intended to be an aid for diagnosis only. Each Physician must interpret the results in conjunction with the patient’s history, physical findings and other diagnostic procedures.

PERFORMANCE CHARACTERISTICS

Sensitivity and Specificity:

A study was performed using 400 patient sera obtained from outside clinical laboratories. These samples were tested using both the RF IgM Rapid Test Cassette (Serum/Plasma) and Roche Tina-quant RF II test following the manufacturers’ package inserts.The following data were obtained:

Method Roche Tina-quant RF II Total Results
RF IgM Rapid Results Positive Negative
Positive 108 3 111
Test Cassette
Negative 5 284 289
Total Results 113 287 400

Relative sensitivity: 95.6%

Relative specificity: 99.0%

Accuracy: 98.0%

REFERENCES

  1. Harris, E. A. 1990. Rheumatoid arthritis: Pathophysiology and Implications for Therapy. N. Engl. J. Med. 322: 1277-1289.
  2. Klareskog, L., J. Ronnelid, S. Gudmundsson, and A. Karlsson-Parra. 1991. Rheumatoid arthritis. Current Opinion in Immunology. 3: 912-916.
  3. Carson, D. A. 1985. Rheumatoid factor. In: Textbook of Rheumatology. W. N. Kelley, E. D. Harris, R. S. Sledge, eds. Philadelphia: W.B. Saunders. pp 664-679.
  4. Mannik, M., F. A. Nardella. 1985. IgG rheumatoid factors and self association of these antibodies. Clin. Rheum. Dis. 11: 551- 572.
  5. Sager, D., R. M. Wernick, M. P. Davey. 1992. Assays for Rheumatoid factor: A review of their utility and limitations in clinical practice. Laboratory Medicine. 23 (1): 15-18.
  6. Linker, J. B. III, and R. C. Williams, Jr. 1986. Tests for detection of rheumatoid factors. In: Man. Clin. Lab. Immunol, 3rd Edition. N. R. Rose, H. Friedman, and J. L. Fahey. eds. ASM, Wash., DC. pp 759-761.