Pneumococcal Urinary Antigen Testing (UAT): Principle, Procedure and Results   5/5 (1)

Streptococcus pneumoniae is one of the most common causes of community-acquired pneumonia (CAP).  Pneumococcal pneumonia has a mortality rate of > 30%, depending on bacteremia, age and underlying diseases.  When not properly diagnosed and treated, S. pneumoniae infection can lead to bacteremia, meningitis, pericarditis, empyema, endocarditis and/or arthritis.Every year S. pneumoniae causes 1 million deaths worldwide in children under 5 years of age and is a leading cause of death in the elderly.

Microbial diagnosis of pneumococcal pneumonia using conventional diagnostic methods remains problematic. Isolation of S. pneumoniae from blood or pleural fluid is regarded as gold standard test, but is positive only in 15-30% of cases. Sputum examination has low specificity due to contamination with oral microflora and invasive samples (such as brochoalveolar lavage, transthoracic needle aspiration) though reliable requires specialist training and may result in complications. Conventional diagnostic techniques includes detection of lanceolate shaped Gram positive cocci in the  sample followed by isolation of alpha-hemolytic, optochin sensitive, bile soluble colonies in the blood agar.   Due to its nature, culture and sensitivity report takes at least 48 hours to arrive so may fail to assist physician for the timely intervention and management of pneumonia cases.

Binax Now Streptococcus Pneumoniae Test
BinaxNOW® Streptococcus Pneumoniae Test

Antigen detection assays are an alternative to standard culture-based methods developed to improve speed and accuracy in the diagnosis of pneumococcal pneumonia. During acute pneumococcal pneumonia, capsular and C-polysaccharide antigens are present in sputum, pleural fluid, blood, and urine. Pneumoccal urinary antigen testing is rapid assay which detects Streptococcus pneumoniae antigen in the urine of patients. BinaxNOW® (Binax) S. pneumoniae test is one of such rapid immunochromatographic membrane test which detects the C-polysaccharide antigen present in all S. pneumoniae. The test may remain positive for several weeks after pneumococcal pneumonia. Pneumoccal urinary antigen test has demonstrated reasonable sensitivity (78% in nonbacteremic cases and 82%–86% in bacteremic cases of community-acquired pneumococcal pneumonia) and good specificity (95%) in different studies. Binax was approved by the US Food and Drug Administration in 1999 for use in diagnosis of pneumonia.

In the suspected cases of pneumococcal meningitis, BinaxNOW can be used to detect c-carbohydrate antigen in the cerebral spinal fluid (CSF) of patients with meningitis.

Materials

  • Provided with the test kits
    • Test Cards: A membrane coated with rabbit antibody specific for S. pneumoniae antigen and with control antibody is combined with rabbit anti-S.pneumoniae antigen and anti-species conjugates in a hinged test card.
    • Reagent A: Citrate/Phosphate buffer with sodium lauryl sulfate, Tween 20 and sodium azide
    • Swabs: Designed for use with Binax Streptococcus pneumoniae (Do not use other swabs).
    • Positive Control Swab: Inactivated S. pneumoniae antigen dried onto swab.
    • Negative Control Swab: S. pneumoniae negative swab.
  • Others:
    • Clock, timer or stopwatch
    • Standard urine collection containers

Principle and Procedure

Binax (Streptococcal antigen testing) procedure
Binax (pneumococcal urinary antigen testing) procedure

Binax pneumoccal urinary antigen test is an immunochromatographic membrane assay test (ICT) used to detect pneumococcal soluble antigen in human urine.  In contrast to other bacterial antigen tests, which detect capsular antigens, the ICT detects the S. pneumoniae C-polysaccharide which is found in the cell wall and is common to all serotypes.

Pneumococcal antigen present in the sample binds to anti-S. pneumoniae conjugated antibody in the ICT strips.  The resulting antigen-conjugate complexes moves by capillary action in the ICT strips and are captured by immobilized anti-S. pneumoniae antibody, forming a Test Line.   Control antibody immobilized in the control region captures anti-species conjugate forming Control Line.  Test results are interpreted by the presence or absence of visually detectable pink-to-purple colored lines.

Sample: Urine

Collect urine specimens in standard containers and transport to the laboratory at room temperature within 24 hours of collection.

Test Procedure:

  1. Allow reagents and cards to equilibrate to room temperature (15-30°C) before testing. Bring patient urine/and or liquid urine control(s) to room temperature.
  2. Remove card from its pouch just before use and lay flat.
  3. Dip an Alere swab into the urine specimen to be tested, completely covering the swab head.
  4. If the swab drips, touch swab to side of the urine container to remove excess liquid.
  5. There are two holes on the inner right panel of the card, insert swab into the BOTTOM hole (swab well). Firmly push upwards to that the swab tip is fully visible in the top hole.
  6. Hold Reagent A vial vertically 1/2 to 1 inch above the card and slowly add three (3) drops of Reagent A to the BOTTOM hole.
  7. Immediately peel off adhesive liner from right edge of the test card. Close and securely seal the card. Gently snap off swab shaft which is purposely scored. Avoid dislodging the swab from the well when doing so. Do not reopen.
  8. Read results in window 15 minutes after closing the card.

Result and Interpretation

  1. Positive result: appearance of two pink-to-purple colored lines (both Test line and Control Line) in 15 minutes or less depending on the amount of antigen present. Specimens with low levels of antigen may give a faint test line. Any visible line is positive.
  2. Negative result:  A negative specimen will give a single pink-to-purple colored Control Line, indicating a presumptive negative result. This Control Line means the detection part of the test was done correctly, but no antigen was detected.
  3. Invalid test: appearance of only Test Line
    (Failure of the Control Line to appear in 15 minutes, whether the Test Line is present or not, indicates as invalid assay).

Advantages of using Pneumococcal Urinary Antigen Testing

Positive pneumococcal urinary antigen test
Positive pneumococcal urinary antigen test
  1. Results are available within 15 minutes so enables timely targeted treatment with pathogen-specific antibiotics
  2. Highly specific for Streptococcus pneumoniae and much more sensitive than blood and sputum cultures
  3. Results are not affected by previous antibiotic therapy

Limitations:

False positive results: Healthy children with nasopharyngeal carriage of S. pneumoniae are likely to have positive urinary antigen test results than children who are not carriers.

Because of this reason, some researchers have claimed that Streptococcus pneumoniae urinary antigen test is not likely to be useful for discriminating between children with and without pneumococcal pneumonia.

References and further readings 

  1. Alere BinaxNOW® Streptococcus pneumoniae Antigen Card

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