Construction of circRNA-miRNA-mRNA Network Reveal Functional circRNAs and Key Genes in Acute Spinal Cord Injury.
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The raw data of Western Blot.
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BV2 cells were lysed using RIPA (Solarbio) with 1% protease and 2% phosphatase inhibitor (Sigma). Protein concentration was measured using a BCA Protein Assay Kit (Solarbio, Beijing, China). And then the proteins were transferred to a polyvinylidene fluoride (PVDF) membrane (Millipore) after being separated using SDS-PAGE gels (Millipore, Billerica, MA). The membrane was incubated with primary antibodies at 4 °C overnight after being blocked for 60 min with 5% BSA (Biofroxx) in TBST (Solarbio) at room temperature. The following primary antibodies and dilutions were utilized: rabbit anti-IL-1β (1:1000; Abcam), rabbit anti-IL-6 (1:1000; Abcam), rabbit anti-TNFa (1:1000; Abcam), rabbit anti-iNOS (1:1000; Abcam), rabbit anti-Bax (1:1000; Abcam), rabbit anti-Bcl-2 (1:1000; Abcam) and rabbit anti-Cleaved-Caspase3 (1:1000; Abcam).The membrane was then treated with peroxidase-conjugated anti-rabbit after being cleaned three times with TBST (1:5000; CST). As an endogenous control, an rabbit anti-β-actin antibody (1:1000; Abcam) was employed. Signals were observed by using enhanced chemiluminescence (ECL) substrates (Seven, Beijing, China).