In a Sandwich ELISA, how is the antigen captured and detected?

In a Sandwich ELISA, how is the antigen captured and detected?

• A. The antigen is bound by a single antibody and then measured without amplification.
• B. An antibody is bound to the plate and the antigen displaces it for detection.
• C. The antigen is 'pinned' between two antibodies for capture and detection.
• D. The assay uses a labeled antigen to compete with sample antigen.

Answer: (C)

In a Sandwich ELISA, the antigen is captured by a first antibody coated on the plate and then detected by a second antibody that binds to a different part of the same antigen. This creates a scenario where the antigen is effectively pinned between two antibodies—the capture antibody holds it on the surface, and the detection antibody binds to another epitope, allowing a measurable signal (often via an enzyme label on the detection antibody) after adding substrate. This dual-binding arrangement provides high specificity because both antibodies must recognize the antigen for a signal to appear. The other scenarios don’t fit: using a single antibody without amplification wouldn’t constitute a sandwich; expecting the antigen to displace a bound antibody isn’t how ELISA works; and using a labeled antigen in a competitive format describes a different assay type, not a sandwich ELISA.

In a Sandwich ELISA, the antigen is captured by a first antibody coated on the plate and then detected by a second antibody that binds to a different part of the same antigen. This creates a scenario where the antigen is effectively pinned between two antibodies—the capture antibody holds it on the surface, and the detection antibody binds to another epitope, allowing a measurable signal (often via an enzyme label on the detection antibody) after adding substrate. This dual-binding arrangement provides high specificity because both antibodies must recognize the antigen for a signal to appear.

The other scenarios don’t fit: using a single antibody without amplification wouldn’t constitute a sandwich; expecting the antigen to displace a bound antibody isn’t how ELISA works; and using a labeled antigen in a competitive format describes a different assay type, not a sandwich ELISA.