Types of ELISA

All ELISA relies on the specific interaction or antibody and antigen. There numerous variation of ELISA method. Listed here are the most common ELISA types:

Direct ELISA

  • Advantage

    • Faster because it only uses one antibody
    • It eliminates cross-reactivity between antibodies.
  • Disadvantage

    • Inflexible and expensive: the primary antibody must be labeled individually and cannot be used for multiple experiments
    • The signal is less amplified.
  • Is typically used when: Analyzing immune response to an antigen.

Indirect ELISA

  • Advantage

    • High sensitivity: the primary antibody has several epitopes than can be bound by multiple secondary labeled antibodies, which amplify the signal.
    • Flexible: different primary detection antibodies can be used with a single labeled secondary antibody.
    • Numerous variations of labeled secondary antibodies are available commercially.
  • Disadvantage

    • Cross-reaction might occur with the secondary antibody.
    • Needs extra incubation steps.

The image provides a schematic representation of four ELISA techniques: Direct ELISA shows a primary antibody conjugated to an enzyme binding directly to an antigen; Indirect ELISA uses a secondary antibody conjugate for detection; Sandwich ELISA employs a capture antibody and a secondary antibody to sandwich the antigen; and Competitive ELISA involves an inhibitor antigen competing with the target antigen for antibody binding, affecting the signal output. Each technique is depicted with color-coded elements to illustrate the specific interactions and components involved.

Figure 1: Types of ELISA

  • Is typically used when: Determining total antibody concentration in samples.

Sandwich ELISA

  • Advantage

    • Very high sensitivity: Two antibodies are used so that the antigen is specifically captured and detected. The antigen doesn’t require purification prior to the measurement
    • Flexibility: Both indirect and direct detection can be used.
  • Disadvantage

    • A matched pair is required: Antibody combination optimization can be difficult. Each antibody has to react with one specific epitope on the target protein and cannot cross react with its partner antibody.
  • Is typically used when: Analyzing complex samples. We don't need to purify the antigen prior using this method.

Competitive ELISA

  • Advantage

    • High sensitivity: it can specifically detect antigens in relatively small amount.
    • Requires no sample processing.
    • Greater precision, accuracy and reproducibility.
  • Disadvantage

    • False positive: low specificity.
  • Is typically used when: Detecting small antigens that cannot be bound by two different antibodies such as in the sandwich ELISA technique. This ELISA method is commonly used when only one antibody is available for the antigen.