Optimization of Capture ELISAs for Chicken Cytokines Using Commercially Available Antibodies

Cytokines like interferon (IFN)-γ, interleukin (IL)-2, IL-6, IL-10, and IL-12p40 are important biomarkers for characterizing the nature and strength of immune responses. It is important to be able to quantify the cytokines at the protein level in biological samples. Quantification of chicken cytokin...

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Bibliographic Details
Published in:Animals (Basel) 2022-11, Vol.12 (21), p.3040
Main Authors: Krzysica, Paulina, Verhoog, Loes, de Vries, Sonja, Smits, Coen, Savelkoul, Huub F J, Tijhaar, Edwin
Format: Article
Language:English
Subjects:
Antibodies
Biological properties
Biological samples
Biomarkers
Cell culture
Chickens
cytokine
Cytokines
DNA-directed RNA polymerase
ELISA
Immune response
Immune system
Immunoassay
Immunology
Interferon
interleukin
Interleukin 10
Interleukin 12
Interleukin 2
Interleukin 6
mRNA
Optimization
Pathogens
Plasma
Polymerase chain reaction
Poultry
Proteins
Reagents
Tumor necrosis factor-TNF
γ-Interferon
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Summary:Cytokines like interferon (IFN)-γ, interleukin (IL)-2, IL-6, IL-10, and IL-12p40 are important biomarkers for characterizing the nature and strength of immune responses. It is important to be able to quantify the cytokines at the protein level in biological samples. Quantification of chicken cytokines is generally performed on the level of messenger RNA (mRNA) by quantitative polymerase chain reaction (qPCR) because very few capture ELISAs for the quantification of chicken cytokine proteins are commercially available. Here, we describe the optimization and validation of capture ELISAs for chicken IL-2, IL-6, IL-10, IL-12p40, and IFN-γ using commercially available antibodies and reagents. First, we determined the optimal concentrations of the antibodies. We then verified the ELISAs’ performance and established that the lower limit of detection (LLOD) for all cytokines was below 32 pg/mL. The ELISAs show the same binding characteristics for recombinant and native cytokines (parallelism was
ISSN:2076-2615
2076-2615
DOI:10.3390/ani12213040