Md Raish MLT

Interpretation of Widal Test-Tube Method
If you have typhoid fever and there is some agglutination, you will see that the 9th tube (positive control) will look similar to one of the eight other tubes. If there is no enteric fever, there will be no change in the normal range of the eight tubes and widal test.

If the tube that showed agglutination has a titre of more than 1:100 in case of O and 1: 200 in H, it will be widal test positive (active infection). Other than this, rest titers are considered the normal range of a widal test.

Limitations of Widal Test
No doubt, the widal test is a quick and effective way to diagnose typhoid fever, but it has some limitations as well, including:

The results of the Widal test can be falsely positive in the case of past vaccination or S. Typhi infection.
The Widal test is time-consuming; until a diagnosis is made, it becomes too late to start the treatment.
A widal test can not distinguish between a patient's past infection, current infection, or a S. Typhi vaccination.
The test results can be falsely positive in typhus, acute falciparum malaria, chronic liver disease, rheumatoid arthritis, nephrotic syndrome and myelomatosis.
Because so many factors can influence the test results, it is better to not just depend on this test for typhoid diagnosis.
However, it is always best to choose trustable labs regarding screening. A delay or ineffective screening can cause a delay in the treatment.

S. Paratyphi is more than or equal to 1:160.
If S. Typhi O is positive, then you have an active fever. However, in the case of positive S. Typhi H, the fever is a past infection. Moreover, the Widal test normal range is when the titre is equal to or below 1:80 in both O and H antigens.

Tube Widal Test
Apart from qualitative and quantitative Widal tests, there is another test method; the standard tube method. Here, eight tubes are taken, diluted and observed. The first test can screen for typhoid fever; however, a tube Widal test effectively confirms a fever.

Originally, two types of tubes were used to detect typhoid fever:

Dreyer’s tube for H agglutination
Felix tube for O agglutination
Nowadays, 3 x 0.5 ml Kahn tubes are used for both O and H agglutination.

Process of Standard Tube Method:

To prepare for a Widal test, we would require the following objects:

Patient’s serum
O, H, AH and BH Antigens
Normal Saline
Pipette
Test Tube Rack
Test Tubes
Water Bath
Before beginning this test, it is essential to note that the tube method is a dilution technique that has to be done correctly. Let’s understand this test in a simpler language.

First, take nine tubes and arrange them in the rack. In the case of O, mark the tubes in numbers from 1 to 9.
Add and mix 0.1 ml normal saline and 0.9 ml serum in the first test tube. On the other hand, add 0.5 normal salines to each remaining tube.
Next, take 0.5 ml from the 1st tube and add in the second. This will result in 0.5 ml remaining solution in test tube 1 and 1 ml in test tube 2nd.
Repeat this process i.e., take 0.5 ml from the last tube and add it to the next tube to make it 1 ml. With the 8th tube, take 0.5 ml and keep that in another separate tube.
Mix all the tubes properly. This will give us primary serial dilution of all the tubes from 1st to 8th as 1:10, 1:20, 1:40, 1:80, 1:160, 1:320, 1:640, and 1:1280, respectively.
Take a new (9th) test tube and add positive control.
Next, add 0.5 ml of respective antigen (O, H, AH, BH) in all eight tubes. This will make the final volume of each tube 1 ml.
After adding a reagent to all the tubes, we will have the final serial dilution of all the tubes from 1st to 8th as 1:20, 1:40, 1:80, 1:160, 1:320, 1:640, 1:1280, 1:2580.
Now, mix well, cover and incubate the tubes at 37° C overnight (18 to 24 hours).