Cytokine secretion

Guide from Exploration to Precise Validation

CAR-T cell activation following antigen recognition induces cytokine secretion.
Measurement of these cytokines provides a widely used functional readout of CAR-T activity, from early exploration to final validation.

Explore → Profile → Quantify → Validate
Different assay formats correspond to different stages of analysis. Combining these approaches enables efficient identification, characterization, and validation of CAR-T activity.

1. Explore

Broad screening of dozens to thousands of cytokines simultaneously → Cytokine arrays

Cytokine networks involved in CAR-T activity are complex and highly interconnected.
Single-target assays provide limited insight, especially at early stages of evaluation. Cytokine arrays enable the simultaneous detection of multiple cytokines, allowing a broader characterization of immune responses and identification of relevant biomarkers.

  • Semi-quantitative detection of multiple cytokines in one sample (~50 µL)
  • Multi-species panels available (Human to Rhesus macaque)
  • Useful when targets are not yet defined 
Series Array format Detection Quantification Key advantage Best use
C-Series™ Membrane array Chemiluminescent Semi-quantitative Simple, fast workflow Initial cytokine screening
G-Series™ Glass slide array Fluorescent Semi-quantitative Higher multiplex, low sample input Standard cytokine profiling
L-Series™ Large-scale array Fluorescent / Chemiluminescent Semi-quantitative Ultra-high plex (up to 8,000 targets) Biomarker discovery / deep screening

2. Profile

Profile Cytokine patterns → Multiplex immunoassays 

Once candidate cytokines are identified, multiplex assays enable parallel measurement within a defined panel.

  • Simultaneous measurement of multiple cytokines
  • Suitable for comparative and condition-based studies 

It allows to compare CAR-T responses across conditions, analyze cytokine signatures and refine target selection.

3. Quantify

Cytokine Arrays to quantify multiple cytokine levels → Quantibody Multiplex ELISA

This approach is suitable for cytokine release profiling and quantitative comparison of multiple markers, with performance approaching that of single-target ELISA.

  • Quantify multiple cytokines simultaneously (pg/mL)
  • Analyze up to 40 proteins from ~50 µL sample 

Multiplex cytokine arrays enable simultaneous analysis of multiple immune markers to better understand biological responses in CAR-T workflows. Use multiplex approaches to explore and quantify complex cytokine responses and high-sensitivity ELISA for precise single-target measurement. 

4. Validate

Accurate assessment of CAR-T activation requires precise quantification of key cytokines, making single-target ELISA the preferred method for functional validation.
Key advantages:

  • Detect low-abundance cytokines in the pg/mL range (LLOQ ~1–2 pg/mL)
  • Maintain high specificity and reproducibility (≤10–20% CV)
  • Perform reliably across serum, plasma, and cell culture media 

Examples of key cytokines providing a functional readout of CAR-T activity:

  • IFN-γ: antigen-specific activation and cytotoxic response
  • IL-2: T-cell proliferation and persistence
  • TNF-α: pro-inflammatory effector function 

Preclinical models? Contact us for mouse or other equivalents.

Not sure what fits your workflow?

Get expert guidance from our team to identify the right approach for your CAR-T workflow.

What Sets ProMab’s CAR Engineering Tools Apart

  • Ready-to-use cryopreserved CAR-T cells for rapid functional studies
  • 100+ CAR constructs available (DNA, mRNA, LNP, or engineered cells)

mRNA-LNP delivery :

  • No cloning. No viral packaging. Fast, streamlined workflows
  • 78–99% transfection efficiency in primary T, NK, iPSC, and dendritic cells
  • Integrated functional assays, including real-time killing, binding, and cytokine analysis
  • Clinically validated technology, supported by 20+ patents and 4 clinical-stage programs (up to Phase II)

Prefer to outsource? 

Send your samples, our CRO delivers high-quality, publication-ready results using multiplex, ELISA, and array-based platforms.

  • References


    Article content created by Tebubio using courtesy materials provided by Abnova and Raybiotech.