![]() ![]() Taken together, our study suggests that β-actin whose expression increases following SCI is not a suitable internal control for Western blot analysis of spinal cord tissues following a traumatic injury. ![]() AM1021b was diluted at 1:1000 at each lane. ![]() The CV of β-tubulin expression following SCI was 14.3 β 3.96%, significantly less than that of the β-actin expression (43.79 ± 4.67% p < 0.01). Western blot analysis of lysates from HepG2, HL-60 cell line (from left to right), using Beta-actin Antibody(Cat. In contrast, no statistically significant difference was found in β- tubulin expression following SCI, compared with sham-operated controls. Coefficient of variation (CV) analysis showed that the CV of β-actin expression was 43.79 ± 4.67%, significantly higher than that of six loadings from a single sample (6.5 ± 0.9%, p < 0.01), indicating that increased expression of β-actin was a result of SCI, instead of a loading error. Using Western blot analysis, we demonstrated that spinal cord injury (SCI) induced a significant increase in β-actin expression which peaked at 7 days post-SCI (2.48-fold). Rated 5 out of 5 by ElenaS from SC-69879 - Anti -b Actin AC-15 in western blot We are really satisfied of the Cod. If so, their use as an internal control may have a negative impact on data acquisition, analysis, and interpretation. However, no information is available concerning the stability of their expressions in response to a traumatic injury to the central nervous system (CNS). Actin Antibody (C4) is a high quality monoclonal beta-Actin antibody suitable for the detection of the beta-Actin protein of mouse, rat, human, avian, bovine. To date, two housekeeping gene–coded proteins (i.e., β-actin and β-tubulin) are widely used as internal controls in the Western blot analysis. To control and correct for loading error, an internal control is often used. #BETA ACTIN WESTERN BLOT FOR FREE#The experiments were repeated three times.įull paper Login or join for free to view the full paper.Western blot is a widely used method for determining specific protein levels. #BETA ACTIN WESTERN BLOT SOFTWARE#Finally, the membrane was washed six times with TSBT and developed using enhanced chemiluminescent (ECL), with the gray value of the protein bands quantified by ImageJ 1.48u software (Bio-Rad, Hercules, CA, U.S.A.). ![]() The membrane was then incubated with the horseradish peroxidase (HRP)-conjugated secondary antibodies at room temperature for 1 h: goat anti-mouse IgG (1:5000, ab6789, Abcam) or anti-rabbit (1:5000, ab6721, Abcam). Membrane was blocked using 5% skim milk for 1 h at room temperature, followed by incubation at 4☌ overnight with Tris-buffer saline with Tween (TBST)-diluted primary antibodies: anti-p62 (1:1000, ab56416, Abcam), anti-LC3B (1:10000, L7543, Sigma–Aldrich, St. Low antibody specificity can lead to a high background on a fluorescent or chemiluminescent western blot. The cell lysates were then separated using 10% PAGE followed by transfer onto a polyvinylidene fluoride (PVDF) membrane. beta-Actin antibody detects beta-actin, a cytoskeletal protein with a predicted molecular weight of 42 kDa. Non-specific bands aren’t the only issue related to blocking. The PR-M(+) cells were lysed in a lysis buffer containing proteinase and phosphatase inhibitor at 4☌ for 30 min and centrifuged at 10,000 r/min for 15 min. ![]()
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