ELISpot (Enzyme-Linked ImmunoSpot) is a sensitive immunological assay closely related to the ELISA technique that was developed to detect and quantify secreted proteins, such as cytokines, growth factors, or antibodies, at the single-cell level. ELISpot assays allow visualization of the secretory product of individually activated or responding cells. Each spot that develops in the assay represents a single reactive cell. They provide valuable insights into the immune response at the single-cell level and are essential in immunology and vaccine research. We offer a range of ready to use T-cell ELISpot kits as well as ELISpot kits with uncoated plates for more flexibility.

How do ELISpot work?

The ELISpot assay is based on the sandwich ELISA technique but adapted to measure secretion at the single-cell level:

• A microplate well covered in a PVDF membrane is coated with a specific capture antibody against the target analyte (eg cytokine, immunoglobulin).

• Cells are added to the wells and cultured under stimulating or resting conditions. As they secrete the analyte, it's immediately captured by the antibody coated on the well.

• After removing the cells and washing the well, a biotinylated detection antibody is added that binds to the capture analyte. This is followed by an enzyme-conjugated reagent (commonly streptavidin–alkaline phosphatase).

• A substrate is added, producing an insoluble colored precipitate at the site of secretion. Each spot corresponds to a single secreting cell.

• The number of spots is counted, providing a direct measure of the frequency of analyte-secreting cells in the sample.

How are ELISpot results read?

There are two options to read ELISpot results:

• Use a specialized ELISpot reader with dedicated software that will automatically identify and count the spots in each well through high-resolution imaging.

• Use a microscope for manual reading (this option is more time-consuming and less reproducible).

Each spot corresponds to a single cell secreting the targeted cytokine. The frequency of spots reflects the number of antigen-secreting T-cells in the sample. Spot size and intensity can also provide qualitative information about secretion strength.

What research applications are ELISpot used in?

T-cell ELISpot assays are mainly used to measure antigen-specific T-cell responses in a variety of applications:

Vaccine research and development

Quantify how well candidate vaccines stimulate antigen‑specific T cells. Provide sensitive readouts of protective cellular immunity.

Infectious diseases studies

Detect T-cell responses to pathogens such as viruses (HIV, influenza, SARSCoV2), bacteria (tuberculosis), and parasites.

Cancer immunology and immunotherapy

Measure tumor-specific T-cell activity. Evaluate patient responses to checkpoint inhibitors, CART therapies, and cancer vaccines.

Basic immunology studies

Investigate cytokine secretion profiles (IFNγ, IL-2, IL-10, TNFα) from individual T cells. Enables frequency analysis of rare antigen-specific populations that are difficult to detect with other methods.

Why use ELISpot assays?

ELISpot assays provide an accurate and sensitive method to assess the T-cell responses. Their key benefits are:

• High sensitivity: Detect rare antigen-specific T-cells.

• Single-cell resolution: Each spot corresponds to one secreting cell.

Our T‑cell ELISpot assays provide sensitive, single‑cell resolution for key cytokines such as IFN Gamma, IL‑2, IL‑4, and IL‑10, enabling precise immune monitoring across research and clinical applications.