Anti-IL12A Antibody Picoband®

Western blot analysis of IL12A using anti-IL12A antibody (A00918-1).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 50ug of sample under reducing conditions.
Lane 1: human Hela cell lysates,
Lane 2: human SW620 cell lysates.
After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-IL12A antigen affinity purified polyclonal antibody (Catalog # A00918-1) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for IL12A at approximately 35KD. The expected band size for IL12A is at 25KD.

IL-25 inhibited the production of IL-12, IL-23 in amodel of of neutrophilia-dominant airway inflammation. ( A - D ) Measurement of Il12a , Il12b , Il23a and Il25 mRNA levels in lung tissue using RT-PCR. ( E ) Representative images of IL-12B and CD80 immunofluorescence staining in lung sections. Scale bar, 50 μm. ( F ) The proportion of CD80 staining-positive cells in lung sections in different groups. ( G ) The proportion of IL-12B staining-positive cells in lung sections in different groups. ( H ) The proportion of CD80 and IL-12B staining-positive cells in lung sections in different groups. There were 4–6 mice in each group. One-way ANOVA was used for statistical analysis (* P < 0.05; ** P < 0.01; *** P < 0.001)
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Exogenous IL-25 inhibited LPS-induced M1 polarization and the expression of IL-12 and IL-23 in mouse pulmonary cells. ( A ) Representative Western blots showing IL-12 A and β-Tubulin protein in primary culture of mouse pulmonary macrophages. ( B ) Quantitative analysis of IL-12 A in mouse pulmonary macrophages using ImageJ. Values are expressed in arbitrary units (a.u.). ( C ) Representative Western blots showing IL-12B, IL-23 A, and β-Tubulin protein in mouse pulmonary macrophages. ( D - E ) Quantitative analysis IL-12B, and IL-23A in mouse pulmonary macrophages using ImageJ. ( F - J ) Detection of Il12a, Il12b, Il23a, Cd80 , and Il-1β mRNA level in mouse pulmonary macrophages using RT-PCR. The experiment was repeated 3 times independently, and a similar trend was obtained. One-way ANOVA was used for statistical analysis (* P < 0.05; ** P < 0.01; *** P < 0.001)
Index in PubMed under a CC BY license. PMID: 37898756

IL-25 inhibited LPS-induced translocation of c-Rel in STAT3-dependent manner in pulmonary macrophages. ( A ) The level of STAT3 phosphorylation was detected using Western blot. ( B ) Quantitative analysis of STAT3 phosphorylation in primary culture of pulmonary macrophages using ImageJ. Values are expressed in arbitrary units (a.u.). ( C - D ) Western blot was performed to measure the protein levels of c-Rel in the cytoplasm ( C ) and nucleus ( D ) of pulmonary macrophages. ( E - F ) Quantitative analysis of c-Rel expresion in cytoplasm ( E ) and nucleus ( F ) of pulmonary macrophages using ImageJ. ( G ) Representative images of c-Rel immunofluorescence staining in primary culture of macrophage from lung. Scale bar, 50 μm. ( H ) The protein levels of IL-12 A and IL-23 A of pulmonary macrophages was measured using Western blot. ( I - J ) Quantitative analysis of IL-12 A and IL-23 A in pulmonary macrophages using ImageJ. Values are expressed in arbitrary units (a.u.). ( K ) The graphical abstract of this study, which was drawn by Figdraw. The experiment was repeated 3 times independently, and the similar trend was obtained. One way ANOVA was used for statistical analysis (* P < 0.05; ** P < 0.01; *** P < 0.001)
Index in PubMed under a CC BY license. PMID: 37898756

The expression of IL-25 was decreased whereas IL-12, IL-23, and M1 macrophage markers were increased in severe of non-eosinophilic asthmatics. ( A ) IL-25 protein levels were decreased in sputum from a cohort of non-smoker severe asthma in the U-BIOPRED study. Based on the analysis of the data provided by Takahashi et al. [ ], 158 downregulated and 187 upregulated proteins were identified in the supernatant of induced sputum from non-smoker severe asthma patients (n = 37) compared to controls (n = 18) by proteomic assay and were shown in the volcano plot. The protein level of IL-25 (indicated by the red arrow) was decreased in the supernatant of induced sputum from non-smoker severe asthma patients compared to controls (log2 of fold change = -0.274, P = 0.019). ( B ) Detection of IL-25 mRNA level in airway brushings of eosinophilic asthma (n = 20) and non-eosinophilic asthma (n = 14) by RT-PCR. ( C - E ) Detection of IL-12 A, IL-12B and IL-23 A mRNA level in induced sputum of eosinophilic asthma (n = 17) and non-eosinophilic asthma (n = 12) by RT-PCR. ( F - J ) Detection of I L-1β, IFN-γ, CD80, iNOS , and CD206 mRNA level in induced sputum of eosinophilic asthma (n = 17) and non-eosinophilic asthma (n = 12) by RT-PCR. ( K ) Representative images of IL-12B and CD80 immunofluorescence staining in BALF cells of control, eosinophilic asthma, and non-eosinophilic asthma. Scale bar, 5 μm. ( L ) Detection of IL-25 mRNA level in induced sputum of eosinophilic asthma (n = 17) and non-eosinophilic asthma (n = 12) by RT-PCR. One way ANOVA was used for statistical analysis. (* P < 0.05; ** P < 0.01; *** P < 0.001)
Index in PubMed under a CC BY license. PMID: 37898756

IHC analysis of IL12A using anti-IL12A antibody (A00918-1).
IL12A was detected in paraffin-embedded section of mouse small intestine tissue. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-IL12A Antibody (A00918-1) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

IHC analysis of IL12A using anti-IL12A antibody (A00918-1).
IL12A was detected in paraffin-embedded section of mouse spleen tissue. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-IL12A Antibody (A00918-1) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.