Lysis buffer (Biosharp, Hefei, China) including 5 g/L sodium deoxycholate, 0.2 g/L NaN3,10 mL/l NP-40, 150 mmol/L NaCl 100 μg/mL, 0.1 g/LSDS, phenylmethylsulfonyl fluoride, 1 μg/mL aprotinin, and 50 mmol/L Tris-HCl (pH 8.5) was used to lyse the cells or tissues according to the manufacturer’s instructions. We used 12% SDS-PAGE to purify protein samples and then transferred them to nitrocellulose membranes (GE Healthcare, Milan, Italy). To avoid unspecific binding, Tris-buffered saline/Tween-20 (0.1%, Bioeasy, Shanghai, China) including 5% non-fat milk (Merck, Darmstadt, Germany) was used to incubate with the membrane at room temperature for 2 h. Subsequently, monoclonal antibodies against DROSHA (anti-DROSHA antibody, 1:1000, RT, 2h, Abcam, Boston, MA) were incubated with the blot, and at the same time a monoclonal antibody against β-actin (anti-β-actin antibody, 1:10000, RT, 1 h, Abcam, Boston, MA) were used as an internal control. After washing with PBS (Invitrogen, CA), secondary antibody conjugated to HRP (1:10000, RT, 1 h, Abcam, Boston, MA) and the ECL Western Blotting Kit (Promega, Milan, Italy) were used for signal detection according to the manufacturers’ protocols.
Anti drosha antibody
Manufactured by Abcam
Sourced in United States
Anti-Drosha antibody is a laboratory reagent used to detect the presence and expression levels of the Drosha protein in biological samples. Drosha is an enzyme involved in the initial processing of microRNA precursors. This antibody can be used in various applications, such as Western blotting, immunohistochemistry, and immunoprecipitation, to study Drosha and its role in cellular processes.
Automatically generated - may contain errors
Lab products found in correlation
Lysis buffer, by Biosharp (1 mentions)
Nitrocellulose membrane, by GE Healthcare (1 mentions)
Ecl western blotting kit, by Promega (1 mentions)
Phosphate buffered saline (pbs), by Thermo Fisher Scientific (1 mentions)
Anti β actin antibody, by Abcam (1 mentions)
Non fat milk, by Merck Group (1 mentions)
Trizol reagent, by Thermo Fisher Scientific (1 mentions)
Protein g plus protein a agarose beads, by Merck Group (1 mentions)
3 protocols using anti drosha antibody
1
DROSHA Protein Expression Analysis
2
Elucidating RNA Regulatory Interactions
Check if the same lab product or an alternative is used in the 5 most similar protocols
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RIP assay was carried out to explore the interaction between Drosha, pri-miR-195 or pri-miR-4668, and uc.372 as previously described63 (link). Briefly, cells were seeded at a density of 1 × 105 cells/ml for 24 h. Then, the cells were transfected with Ad-NC or Ad-uc.372m for 48 h. After that the cells were incubated with glycine after fixing by formaldehyde (0.3%). The cells were washed with phosphate-buffered saline (PBS) for three times and cell suspension was reserved in RIP buffer. Then, anti-Drosha antibody (Abcam, USA) was added and incubated with the cell suspension at 37 °C overnight. TRIzol reagent (Invitrogen) was used to isolate the precipitated RNA and the level of pri-miR-195 or pri-miR-4668 was further analyzed using real-time PCR.
3
Chromatin Immunoprecipitation and RNA Immunoprecipitation
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