ELISpot Assay: A Powerful Tool for Measuring Cellular Immune Responses

BioAgilytix offers Enzyme-Linked Immunosorbent Spot (ELISpot) assays at multiple global locations. With the ability to qualify and validate assays for nonregulated or regulated work, BioAgilytix’s ELISpot platform provides robust data for monitoring cell-mediated immune responses to biologics, cell therapies, and gene therapies during clinical and preclinical testing.  

t cell

BioAgilytix’s ELISpot expertise

The ELISpot platform provides a method for accurately detecting and enumerating antigen-specific immune cells with a high degree of sensitivity. BioAgilytix’s ELISpot assays can be qualified and validated to support regulated studies, and our team has a proven track record of leveraging this platform to deliver high-quality results. Speak with one of our ELISpot experts about your scientific requirements and learn how the features of this platform can be optimally leveraged for bioanalysis in support of your therapeutic program.  

cancer cells 3d illustration

ELISpot applications for determining cell-mediated immunity  

The ELISpot platform has become a leading technology for monitoring cell-mediated immunity in the regulated bioanalytical space. ELISpot assays provide quantitative information regarding the number of cells secreting specific cytokines as part of a cellular immune response directed toward therapeutic proteins or peptides. 

We use ELISpot to visualize individual cells that secrete cytokines after stimulation. By adding a known concentration of a patient’s immune cells, such as purified peripheral blood mononuclear cells (PBMCs), the frequency of antigen-specific cells can be determined after in vitro stimulation with the antigen of interest. For cellular immunogenicity, proteins or peptides derived from the therapeutic can be used for stimulation, which provides information on the drug-specific immune response. 

At BioAgilytix, we most frequently utilize ELISpot in our work with cellular immune responses or T cell responses to cell and gene therapy products. However, we have also supported determination of antigenic epitopes, and antigen-specific B cell or plasma cell prevalence. For bioanalytical assessment of gene therapies, ELISpot can be used to assess cellular immune responses against the vector and transgene. With cell therapy products, the response against unique sequences in the CAR construct or other novel proteins expressed on the therapeutic can be evaluated.   

Key ELISpot Benefits:

Exceptional sensitivity and single cell detection  

The ELISpot platform offers several distinct advantages over other methods. First, the cells used in the assay can be isolated and cryopreserved, negating the need for fresh samples at the point of analysis. Stimulation of the cells then takes place in the final assay plate, which streamlines the laboratory procedure.  

A cell that is activated by the proteins or peptides added to the wells during stimulation will produce cytokines, which are secreted into the surrounding matrix. The wells of the assay plate have a membrane coated with antibodies that captures the secreted cytokines. When the cells are removed from the plate at the end of the assay, each activated cell can then be visualized as spots of cytokines left behind on the membrane. By visualizing each individual activated cell, this methodology offers exceptional sensitivity and provides a direct, quantitative readout of antigen-specific cell frequency. 

Selecting the specific cytokine — or cytokines for multiplexed assays — detected in this assay format can also provide an indication of the immune cell types that are activated in response to a stimulant. As such, ELISpot assays are able to provide informative data sets for evaluating and predicting the type of immune responses occurring in the patient. 

Benefits of ELISpot assays: 
  • High sensitivity 
  • Single cell detection and enumeration 
  • Scalability for higher throughput 

How the ELISpot platform works 

BioAgilytix utilizes ImmunoSpot® ELISpot plate readers equipped with top-end optics to enable high-resolution detection. These systems support traditional enzymatic assays as well as fluorescence and bright-field imaging. The ImmunoSpot software can accommodate single- or double-color enzymatic assays (blue spot/single analyte or blue and red spot/dual analyte) as well as FluoroSpot assays (single or multiplex up to four analytes). An automated plate loader also enables scaling up for higher throughput applications. The instruments and software at BioAgilytix have been installed and validated to be 21 CFR Part 11 compliant, enabling full use of the platform for regulated assay support.