Diagnostics
Table 1: Test Validation Data with the ReaScan Strep A Test Sample
GAS control swab (Bioclin, Finland) GAS control swab (Bioclin, Finland) GAS control swab (Bioclin, Finland)
GAS positive control included in one kit lot*
1 mm x 1 mm GAS colony on blood agar dispensed in 1 ml PBS Extraction reagent blank
n
20 20 20 30 10 10
1 mm x 1 mm Streptococcus group B** colony on blood agar dispensed in 1 ml PBS 10 Negative patient samples***
176 CFU/Swab
1.2 x 105 1.2 x 104 1.2 x 103 N/A N/A 0
N/A 0
ReaScan
Value Range 100–112 40–52 10–19
276–347 220–260 0–3 0–5
0–19 Mean of
ReaScan Value 109 46 13
315 239 1 2 1
* The positive control is prepared to a level corresponding to 1.2 x 105 CFU/swab or more (most commercial RADT tests show positive GAS at this level). ** Similarly, Group C, F and G streptococci, (S. milleri, S. anginosus, S. constellatus, S. intermedius), S. salivarius, Enterococcus faecalis, E. casseliflavus, Staphylococcus aureus and Arcanobacterium haemolyticum gave ReaScan values ≤5.0 (= negative).
*** The samples were confirmed as negative with the standard selective sheep blood agar for streptococci cultivation method. Theoretical sensitivity: 1 x 104 CFU/sample. Specificity: only GAS has been found to be reactive in the test. CFU = colony-forming units; GAS = Group A Streptococcus; N/A = not available; PBS = phosphate-buffered saline.
Figure 1A: Outline of the Test Principle Figure 1B: Outline of the Test Procedure
Gold-conjugated GAS antibody
Dispense three drops of extraction reagents 1 and 2 to the assay vial
GAS antigen in the sample
Immerse the swab into the assay vial and rotate it for one minute
Dispense three drops of extraction reagent 3 into the assay vial
Test membrane GAS = Group A Streptococcus.
by the instrument and therefore the observer’s impaired vision or expectations do not interfere with the results.
ReaScan Strep A
The ReaScan Strep A test (Reagena International Ltd, Toivala, Finland) is a modified immunochromatographic lateral-flow test for the detection of GAS from a regular flocked nylon throat swab (microRheologics, Brescia, Italy). This rapid test detects streptococcal Lancefield group A antigen, which is extracted from the specimen by the extraction reagents included in the test kit. The extraction follows a previously well-described procedure.7
The extracted GAS antigen is allowed to
react with gold particles coated with GAS-specific antibodies. Then other membrane-bound GAS antibodies capture the gold particles with the GAS antigen causing the appearance of a red test line (see Figure 1A). An outline of the test protocol is shown in Figure 1B. The intensity of the test line is measured by a ReaScan reader, which gives a numerical value proportional to the amount of antigen in the sample. Since the test result is based on a numerical value, human errors are minimised, thus increasing the diagnostic value of the ReaScan Strep A test.
The discrepant analysis of the ReaScan Strep A test was performed comparing the rapid test with the standard culture method and
144
Pipette 80 µl from the conjugate vial to the test cassette
Wait 15 minutes and read the result with ReaScan©
reader
with the TestPack® Strep A test (Inverness Medical, Cranfield, UK), according to instructions for use. Three parallel swab samples were taken for each of the study methods from 231 volunteers visiting the outpatient clinic of Oulu University Hospital. The standard culture method8
Pipette 100 µl from the assay vial to the conjugate vial and mix well
results were classified as negative or positive (+, ++ or +++). In this method, sheep blood agar for streptococci is used (5 % sheep blood agar, Müller–Hinton + tryptone–soya agar), supplemented with colistin and nalidixic acid (IS-ST-A, Abtek Biological Ltd, UK) and with supplemental carbon dioxide to enhance the growth of streptococci. Streptococcal Grouping Reagent A (DR0586G, Oxoid Ltd, UK) was used.
Results and Discussion
In the present study, we introduced a novel immunochromatographic bacterial Strep A antigen test for the ReaScan rapid test reader. We evaluated the sensitivity, specificity and cross-reactivity of the novel rapid test. Table 1 summarises the test validation data with the ReaScan Strep A test. No cross-reactions with cultured Group B,
EUROPEAN INFECTIOUS DISEASE
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