R&D Systems
Mouse YM1/Chitinase 3-like 3 Antibody
AF2446
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Our projects require the testing of various inflammatory stimuli or agonists on peritoneal macrophages from mice that are either drug-treated or untreated. To confirm or test our hypotheses on what targets may have been modulated upon inflammatory stimuli and drug treatment, we utilize best practice western blot techniques to analyze and measure the protein/target of interest in our experiments.
Western Blot
Mouse peritoneal macrophages (see results for more details)
1/2000 of Ab in 5% milk in TBST, overnight, 4 deg C
Non-fat dry milk proteins in TBS
1/5000 of HRP-Ab in 5% milk in TBST, 1 hour, room temperature
None
X-ray Development using ECL substrate and photofilm
The peritoneal macrophages were sorted using flow cytometry and the drug's inherent fluorescent signature. See - https://www.ncbi.nlm.nih.gov/pubmed/26109497. Post-sorting, the cells were plated for 24-48 hours before the agonists or inflammatory stimuli were applied. 8-24 hours post stimulus, cells were collected and lysed, measure for protein concentration and best practice western blot techniques were used as discussed above. As seen in the figure, the repeatable band production of Ym-1 in three different mice samples was easily verified. As noted, there was sample variability across mice samples and the detection will be quantified via densitometry and normalized using B-actin as a loading control to validate these findings.
10.1002/cyto.a.22706
Consistent and repeatable
A very effective Ab for detection of Ym-1 via western blot analysis of mouse peritoneal macrophages.