Fig 1: Treatment of chicken macrophage cell line HD11 with LTC stimulated significant increases in the release of IFNa, IL-6, NO, and ROS. Chicken HD11 macrophages were plated in 24-well plates and treated with either nothing, LPS (300 ng/mL), or 1, 5, 10, 25, or 50 µL/mL LTC for 36 h. LTC treatment stimulated significant increases in chicken IFNa release and INFß mRNA expression from HD11 cells (A,B). Primer pairs for reverse transcribing and generating cDNA from chicken IFNß mRNA transcripts were amplified by qRT-PCR. Quantitative comparisons of transcripts encoding IFNß were determined and compared in either untreated, LPS-treated, or LTC-treated HD11 (B). Differences in IL-6 production were determined by chicken IL-6 ELISA (C). NO release (D) and ROS production (E) were assessed using the Griess reagent and H2DCFDA assays as described in Methods. Significant differences in the release of these mediators were determined using a one-way ANOVA with p-values of **** = 0.001, *** = 0.005, and ** = 0.01.
Fig 2: Treatment of bovine PBMC with LTC stimulates the production of pro-inflammatory cytokines with potential potent antiviral and antibacterial activity. Bovine PBMC were plated at 4.0 × 106 cells per well in 24-well tissue culture plates and treated for 36 h. Supernatants were subsequently harvested and assayed for cytokine/chemokine release. ELISA for release of IFN? (A), TNFa (B), IL-6 (C), IL-17A (D), IL-1a (E), IFNa (F), IL-36RA (G) MCP-1 (H), IL-8 (I), and IL-10 (J). Significant differences in cytokine/chemokine release were determined using one-way ANOVA with p-values of ** = 0.01 and * = 0.05.
Fig 3: Chicken PBMC treated with LTC demonstrated significant increases in the secretion of IFN?, INFa, IL-6, NO, and ROS. PBMC from chickens were seeded in 24-well plates in 1 mL cultures at 4.0 × 106 cells per well. Cells were either untreated or treated with either 300 ng/mL LPS, 1, 5, 10, and 25 µL/mL LTC for 36 h at 42 °C. Medium control assays for cytokines were performed in cultures without cells to determine any cross-reaction of the ELISA components to the medium used in the cultures. Specific chicken cytokine ELISA was performed for IFN? (A), IFNa (B), and IL-6 (C). NO (D) and ROS (E) were assessed using the Griess reagent and H2DCFDA assays as described above. Symbols (?) for each bar represent the mean of triplicate cultures from each of 3–4 different animals. Significant differences in cytokine production via ELISA, NO, or ROS were determined using a one-way ANOVA with p-values of **** = 0.001, *** = 0.005, ** = 0.01, and * = 0.05.
Fig 4: Treatment of bovine nasal turbinate cells (ATCC # CRL-1390) with increasing dosages of LTC stimulates significantly higher secretion of pro-inflammatory cytokines IL-6 and TNFa. Turbinate cells were seeded in 24-well plates and treated with the indicated concentrations of LPS or LTC for 36 h. Secretion of IL-6 (A) and TNFa (B) was determined by ELISA. Significant differences were determined using a one-way ANOVA with p-values of **** = 0.001 and *** = 0.005.
Fig 5: Treatment of the chicken macrophage cell line MQ-NCSU with LTC stimulated significant increases in the release of IFNa, IL-6, and NO but inhibited ROS production. The chicken early monocyte/macrophage cell line MQ-NCSU cells were plated in 24-well plates and treated with 1, 5, 10, 25, or 50 µL/mL LTC or LPS (300 ng/mL) for 36 h, and supernatants assayed for cytokine and NO/ROS production. LTC treatment stimulated significant increases in IFNa release and INFß mRNA expression (A,B). IFNß transcription was assessed in either untreated, LPS-treated, or LTC-treated MQ-NCSU cells (B). IL-6 production was determined by specific IL-6 ELISA (C). NO release (D) and ROS production (E) were assessed using the Griess reagent and H2DCFDA assays as described above. Significant differences in the release of cytokines, mRNA expression, NO, or ROS were determined using a one-way ANOVA with p-values of **** = 0.001, *** = 0.005, ** = 0.01, and * = 0.05.
Supplier Page from Thermo Fisher Scientific for Chicken IL-6 ELISA Kit