Widal Test Using Tube Agglutination Reaction

The tube test is performed for quantifying the agglutinating antibodies in the serum of a patient suffering from typhoid. Add suspension of dead typhoid bacterial cells to a series of tubes containing the diluted patient serum. This technique confirms the actual causal organism of typhoid. This test is based on the principle if higher the titer, the greater is the response of the reaction product. Successive daily tests on a patient serum reveal no or low antibody titer from day to day, the organism is not Salmonella. It is possible only in case when titer increases daily.

Requirements

• Samples of blood serum
• Serological tubes
• Physiological saline (0.85% NaCl)
• Pipette
• Incubator
• Glass marker.

Procedure

1. Take a blood sample from patient suffering from typhoid in a serological tube and collect
2. few more tubes for making the sample.
3. Dispense 0.5 ml of saline to each tube of blood serum sample (1:10) as well as to other
4. tubes (Fig. 15.2).
5. Transfer 0.5 ml blood serum from tube 1 to next tube, mix it and transfer 0.5 ml from this tube to the next, mix it and again transfer into the next tube subsequently. The last tube should not have serum and will contain only saline and can be used as control.
6. Transfer 0.5 ml of S. typhimurium antigen in each tube with the help of a pipette, mix the antigen thoroughly and keep on a water bath at 37ºC for 30 minutes.
7. Examine each tube for agglutination after centrifugation and record the titer in which
8. agglutination has occurred.

Results

Agglutination will show that patient is suffering from typhoid.

Reference:

Dr. R. C. Dubey – Practical Microbiology