Name: Siti Hawa Bte Azali
Admin No.: 0704087F
During the 1st week of my attachment , i was assigned to work in the Microbiology Department where specimens are tested for presence of any pathogenic organisms and where identification of causative agents were carried out. This section is important as it aids in the final confirmation of clinical diagnosis.
Several tests were introduced to me by the section supervisor,some of which were familiar whereas others were not. Among the microbiological tests that i was assigned to practice on and eventually carry out are Gram and AFB Staining, Syphillis Test- VDRL and TPHA, PYR Enterococci test,Mycoplasma Test, etc.
Nature of specimens that this section usually test on are those such as serum (most common), sputum, stool, blood culture,bile, GI fluid and different types of aspirate taken during operation.
One of the test which i frequently did was the Widal Weil Felix Test - a test used to detect and measure levels of warm and cold agglutinins in blood.
Agglutinins are basically antibodies that causes haemagglutination to occur in the body. Cold agglutinins are active at low temperature (IgM) whereas warm agglutinins are active at warm temperatures (IgG).
This test is done on a patient's serum( in a plain tube) by testing it against 11 different reagents consisting of different commercial antigens (mainly different types of Salmonella).
Normal Values:
Warm agglutinins: no agglutination in titres at or below 1:80
Cold agglutinins: no agglutination in titres at or below 1:160
Procedure:
1) 20ul of serum is decanted onto each of 11 individual areas of test card
2) 1 drop of each reagent added to serum on respective test areas (reagent 1 into test circle 1, reagent 2 into test circle 2, etc)
3) Mix the serum and reagent on each circle
3) Rotate the test card for 1 min
4) Check for agglutination
Reagent-serum mixture with agglutination will then be tested again, this time with lower concentrations of serum (10ul and 5ul).
5) Check for agglutination for the 10ul and 5ul serum concentrations
6) Using a standard Widal Test reference table, we can calculate the titre of the agglutinins in the serum (eg. 1:40,1:80,1:160,1:320)
Abnormal values of cold and warm agglutinins may cause hemolytic anemia and therapeutic actions are then taken by the doctor.
High levels of Warm Agglutinins (~1:160,1:320) can be due to:
-Salmonella infections
-Lymphoma
-Systemic Lupus Erythomatosus
High levels of cold agglutinins (~ 1:320) can be due to:
-Mycoplasma and viral infections
-Multiple myeloma.
This test is not normally a routine test but is usually needed for urgent specimens.
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Who are you? Please identify yourself. Please also remove the name of the hospital that you are attached to. Forgot all my instructions!!!!!!
ReplyDeleteHi Siti!
ReplyDeleteCan I ask what is VDRL and TPHA? Also, how do these two test for Syphilis ?
Siew Ming
TG O1
0702862D
Hello Siti!! ((:
ReplyDeleteask you ah, if my test is positive, den how do i noe is it warm or cold agglutinins that causes the positive reaction?
Thanks!
Joanna Yeo
0702054H
Hi Siti.
ReplyDeleteYour very own groupmate here. Would like to know, what does AFB stand for in AFB stain?
The purpose of the test is to detect and measure levels of Ab, but what is the significance? Is this test used for confirmatory or presumptive diagnosis? And when is this test appropriate to be conducted?
Tng Wess Lee
TG02
Grp 10
0702570C
This comment has been removed by the author.
ReplyDeleteHi everybody! Thanks for your questions =)
ReplyDeleteReply to Siew Ming's Qn:
TPHA is basically an indirect hemagglutination test used for the detection antibodies Treponema pallidum. This bacteria is commonly found on the genital tract and it is the the causative agent of Syphillis.
Usually commercial sheep RBC coated with T.pallidum antigens are used to bind with specific Ab present in the patient's serum.
Hemagglutination observed in the microtiter plate wells will indicate a positive result- hence serum is syphyllitic.
VDRL is a hemagglutination test similar to TPHA however is less accurate due to high chances of getting false negative results. Hence VDRL is usually done alongside TPHA, which on the other hand, is a confirmatory test.
Reply to Joanna's Qn:
ReplyDeleteAny reaction occuring at low temperatures (0 - 20 degrees celsius) are induced by IgM which are cold agglutinins, whereby reactions occuring at normal body temperatures (37 degrees celsius) are induced by IgG which are warm agglutinins.
Reply to Wess' Qn:
ReplyDeleteAFB stain refers to 'Acid Fast Bacilli' stain.
Widal Weil Felix Test is a presumptive test for detection of Typhoid Fever.It basically demonstrates presence of agglutinins to Salmonella typhii in patient's serum by testing it against somatic(O) and flagellar(H) antigens. These antigens are commercially available in the Widal Weil Felix Test kit. The test itself is to be conducted when blood culture of patients are positive for Salmonella typhi.
This comment has been removed by the author.
ReplyDeleteOh, I understand now. THanks, siti! (:
ReplyDeleteSiew Ming
TG 02
0702862D
arh...okok! got it. thanks. =D
ReplyDeletewess
heylo Siti!
ReplyDeleteHAHA! may i know why is the Widal Weil Felix Test not done as a routine test but only on urgent samples?
Loves,
Renee
TG02
0703634F
Reply to renee's Qn:
ReplyDeleteHey Renee! =)
Its not 'normally' done on routine specimens..so far about 90% of widal weil felix tests that i did are for urgent specimens...
The reason is because the test should only be done when blood cultures are reported positive for S. typhi. In such cases, Widal Weil Felix test must immediately be done.
Thanks SITI!
ReplyDeleteHi Siti!
ReplyDeleteJust wanted to clarify something. Does it mean that different antigens from different disease will result in the production of different nature of antibodies (cold/warm)? Thanks!
Zi shuang
0703383J
Just wanna ask. Do you do the Widal Weil Felix in the tubes as well? My lab uses only the tube method. Do you know if there is any advantage over the tube method?
ReplyDeletethanks.
Muna
Group 7
0703791D
Hello Muna!! =D
ReplyDeleteNope my lab didnt carry out the tube method but we only did the slide agglutination method.
From what i know tube method is more accurate than slide method as you can go up to 1:1280 titre whereas slide method can only measure up to 1:320 titre.
However an advantage of the slide method is that it is more rapid and convenient...results can be derived in within 3-5 minutes.
Hope that clarifies ur doubt..
Thanks for e qn! =D
Hi Zi Shuang =D
ReplyDeleteIt depends on the type of infection/ disease that occurs. For example, in multiple myeloma, there is an increased level of abnormal IgG and IgM by the plasma tumour cells. In this disease,however, IgG are the type that is highly secreted, followed by IgA and finally IgM.Hence more than 1 class of antibodies is produced.
It is incorrect to say that each disease only result in the production of a particular class of antibody. Instead, each disease may vary in the level of a particular class of Ig it produces.
Hope that helped..
Thanks for e qn!
=)