Confirmatory tests for Saliva
Unlike preliminary tests, the confirmatory tests for saliva are specific to salivary α-amylase and can be used to confirm if the origin of the suspected stain is human saliva. The most commonly used confirmatory test for saliva is the RSID-Saliva test, described in detail below:
RSID-Saliva test
The Rapid Stain Identification test for the detection of saliva or RSID-Saliva test is a lateral flow immunochromatographic strip test designed to detect the presence of salivary α-amylase from as little as 1 μL of human saliva. It is designed for fast, easy, and reliable detection of human saliva from various samples encountered at crime scenes, including envelopes, glass bottles, aluminium cans, plastic lids and swabs of possibly contaminated surfaces.
Principle of the RSID- Saliva test:
Figure: Principle of RSID test for the detection of Saliva
Immunochromatographic assays (including the RSID-Saliva test) are based on the principles of capillary action and specific antigen-antibody interactions to enable rapid and visual detection of target analytes.
This test utilizes a test strip containing distinct zones, including a sample application pad, a conjugate pad containing labelled primary antibodies (specific to salivary α-amylase), a reaction or test zone (T) with immobilized primary antibodies (specific to salivary α-amylase), and a control zone (C) with immobilized secondary antibodies (specific to primary antibody).
When a sample is added to the application pad, it begins to migrate along the membrane facilitated by capillary action. If the target antigen (α-amylase) is present in the test sample, it forms an antigen-antibody complex with labelled primary antibodies on the conjugate pad. These complexes are captured at the test zone (T) and the control zone (Z), resulting in the formation of coloured lines at these zones. The appearance or absence of these lines at the test zone (T) indicates the test result. Since the control zone contains immobilized secondary antibodies specific to primary antibodies, a coloured line will be formed at the control zone (C) even if the target antigen is absent in the test sample and the antigen-antibody complex is not formed. This ensures proper flow and functionality of the test.
📌 Read: Immunochromatographic Assay
Procedure:
Combine the suspected saliva stain extract (maximum 20 μL) with RSID-Universal Buffer to bring the test sample to a total volume of 100 μL.
Add the test sample to the sample window and keep it undisturbed for 10 minutes.
Observation:
a) A visible red line at the Control (C) position only, indicates a negative result, i.e. α-amylase could not be detected.
b) Visible red lines at both the Control (C) and Test (T) positions indicate a positive result and confirm the presence of saliva, i.e. α-amylase is detected.
c) A visible red line at the Test (T) position only indicates an inconclusive or failed test.
Figure: Results for RSID-Saliva test