TYDAL – S
(STAINED TYDAL ANTIGENS FOR SLIDE AND TUBE TEST)
SUMMARY:
Salmonella typhi & Salmonella paratyphi are the causative agents of "Enteric Fever". The antigens of typhoid and paratyphoid consist
of two distinct fractions - the stable somatic 'O' Antigen and the labile flagellar 'H' antigen. The paratyphoid antigens are further classified into
A & B species. In typhoid and paratyphoid, the 'H' antigen is type specific whereas the 'O' antigen is group specific.
Tydal-S antigens are the standardised smooth suspension of killed bacterial antigens for qualitative and semi-quantitative
detection of S. typhi and S. paratyphi antibodies. The different color to each antigen facilitates the differentiation of antigens and clear
reading of agglutination. It also avoids the possible error of mixing and/or misinterpretation. As undiluted serum is used in Slide Test, it is a
simple, rapid and convenient screening test. The slide test antigens are standardised in such a way that they can be used for either slide or
tube technique. In doubtful cases, it is recommended to perform the tube technique for obtaining conclusive results. A marked rise in the
titre to one sero type (above 1:80) suggests infection. Diagnostically a rising antibody titre of at least four fold (two tube difference) is
considered more significant than a single test. It is observed that individuals immunised with TAB vaccine may show a moderately high titre
for all the antigens.
REAGENTS:
Reagent A1 S. typhi 'O' Antigen
Reagent A2 S. typhi 'H' Antigen
Reagent A3 S. paratyphi 'AH' Antigen
Reagent A4 S. Paratyphi 'BH' Antigen
Reagent A5 Positive Control
STORAGE AND STABILITY:
All reagents are stable at 2-8°C till the expiry date mentioned on the labels.
SAMPLE COLLECTION & STORAGE OR SPECIMEN AND STORAGE:
1. No special preparation of the patients is required prior
to sample collection by approved technique. Do not use haemolysed samples.
2. Clean and dry glassware free from detergents must be used for sample collection.
3. Though freshly collected serum is preferable, store samples at 2-8°C in case of delay in testing for up to 72 hrs.
PRECAUTION:
1. Bring All the reagents and samples to room temperature before use.
2. Shake all the antigens thoroughly before use
3. Avoid using turbid, contaminated or inactivated serum.
ACCESSORIES:
1. Slide- 1No.
PROCEDURE:
I. Rapid Screening Slide Test:
1. On a slide with six circles, place 50 µl of test serum in each of the first four circles and 50 µl ml each of Positive Control and Normal
Saline in each of the last two circles respectively.
2. Add one drop each of 'O', 'H', 'AH' and 'BH' antigens in the first four circles respectively and one drop of any one antigen in the
remaining two circles.
3. Mix the contents of each circle separately and spread it in the entire circle.
4. Rock the slide gently for One minute and observe for agglutination.
INTERPRETATION OF RESULTS:
Agglutination with Positive Control and no agglutination with Normal Saline validate test results. No agglutination up to one minute is a
negative test, and indicates the absence of corresponding antibodies.
Agglutination within one minute is a positive test, and indicates presence of corresponding antibodies. Then proceed for semi quantitative
slide or tube technique for determination of antibody tire.
Do not observe result after one minute.
I. Semi-Quantitative Slide Test:
1. Put one drop of normal saline in the first circle and 5 µl, 10 µl, 20 µl, 40 µl & 80 µl of test serum in the remaining five circles
respectively. The corresponding titres obtained will be 1:320, 1:160, 1:80, 1:40, 1:20 respectively.
2. To each of the above circles, add one drop of the appropriate antigen, which gives an agglutination in the Screening Slide Test.
3. Mix the contents of each circle separately and spread it in the entire circle.
4. Rock the Slide gently for one minute & observe for agglutination.
NOTE: This method is recommended for obtaining quick approximately titers only
INTERPRETATION OF RESULTS:
The lowest volume of serum, which shows clear agglutination, indicates the cut off level of the positive test and the corresponding antibody
titre as per the tube technique is given below:
SerumVolume Antibody Titre Serum Volume Antibody Titre
80 µl 1:20 10 µl 1:160
40 µl 1:40 5 µl 1:320
20 µl 1:80
QUANTITATIVE METHOD:
II. Tube technique using slide antigens:
1. Perform the assay for all four antigens or for that which has given a positive result in the Screening Slide Test.
2. Take a set of six test tubes (10 x 75mm) for each antigen. Dilute the serum sample and set up the test as indicated in the table.
TUBE NO. 1 2 3 4 5 6
Dilution Saline Control 1:20 1:40 1:80 1:160 1:320
Normal Saline 1.0 ml 1.9 ml 1.0 ml 1.0 ml 1.0 ml 1.0 ml
Test Serum - 0.1 ml - - - -
Diluted Serum from - - 1.0 ml 1.0 ml 1.0 ml 1.0 ml 1.0 ml
tube 2 Discard
Appropriate One drop One drop One drop One drop One drop One drop
Antigen
3. Mix well after each addition and incubated at 37°C for 18-20 hours.
4. Observe for agglutination. The highest dilution for Serum which shows clear-cut agglutination indicates the antibody titre.
NOTE:
1. Sera from normal individuals may show agglutination up to 1: 40 dilution.
2. Agglutination titre greater than 1:80 is considered significant and usually suggestive of infection.
3. Tydal is only screening test.
4. The correlation of test results with typical clinical signs, symptoms and patient's history should be taken into account before arriving at
the final diagnosis.
5. As with all diagnostic procedures, the Physician should evaluate data obtained by use of this kit in light of other clinical information.
6. For accuracy of results, the procedure has to be followed meticulously.
REFERENCES
1. Cruickshank, R. (1982) Medical Microbiology, 12th Edition, P.403.
2. Felix, A. (1942) Brit. Med. J., 11, 597.
