Search results for: Rabbit Anti-Myostatin) Antibodies
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Production of a polyclonal anti-myostatin antibody and the effects of in ovo administration of the antibody on posthatch broiler growth and muscle mass.
In this study, we produced a polyclonal antibody against unprocessed chicken myostatin and examined the effect of in ovo administration of the antibody on posthatch chicken growth and muscle mass. A PCR-amplified unprocessed chicken myostatin cDNA was cloned into an Escherichia coli expression vector, and myostatin proteins were expressed. Recombinant myostatin purified by electro-elution of the SDS-PAGE fractionated myostatin band was used as an immunogen to produce rabbit polyclonal antimyostatin antibody (pAb-AVM46). In Western blot analysis, the pAb-AVM46 showed high affinity to the myostatin propeptide, but little affinity to the mature myostatin. Two experiments examined the effect of in ovo administration of the pAb-AVM46 on posthatch chicken growth and skeletal muscle mass. In experiment 1, broilers from eggs injected once with 35 microg of the antibody into the yolk on d 3 of incubation had significantly lower combined thigh and leg weight at 4 wk posthatch than the controls that received no injection, or the broilers from eggs received the same dose of antibody into the albumen. In experiment 2, 2 different doses of the antibody (9 or 70 microg) were injected into the yolk, and the effects on body and muscle weight were examined at 5 wk posthatch. Birds from eggs injected with 70 microg of the antibody had significantly lighter (11.6%) combined thigh and leg weight than the control birds. The percentage of the combined thigh and leg weight to BW of the 70-microg group was also significantly lower than that of the control group (20.95 vs. 23.08%). The results of this study indicate that unprocessed full-length myostatin as an immunogen produced antibody populations having affinity mostly to the propeptide with little to the mature form. The decreased muscle weight observed in broilers injected with the antibody in the yolk indicates that myostatin activity was probably elevated by the binding of the antibody to the propeptide, and provides evidence that myostatin propeptide inhibits the biological activity of myostatin in broilers.Y S Kim, N K Bobbili, Y K Lee, H J Jin, M A Dunn
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[Functional characterization of recombinant myostatin and its inhibitory role to chicken muscle development].
Myostatin is a recently discovered member of transforming growth factor beta (TGFbeta) superfamily and shares similar structure features with other members of TGFbeta superfamily. For a better understanding of molecular mechanism of myostatin function, the production of C-terminal truncated form of recombinant myostatin protein (rMSTN) in E. coli was previously reported. Herein, the functional role of the recombinant myostatin in regulating myogenesis in a chicken embryonic myoblasts (CEMs) system was determined. By using flow cytometric analysis, the myostatin was found to inhibit cell cycle transition from G1 to S phase and result in a cell cycle arrest at G1. In addition, myostatin blocked the multi-nucleus myotube formation and caused a decreased expression of the muscle cell differentiation markers (myogenin and MHC) in CEMs. In this study, a rabbit polyclonal antibody against myostatin was produced and high affinity and specificity of this anti-myostatin antibody to recombinant and endogenous myostatin were assayed by Western blot analysis. Further studies showed that the antibody could also recognize the tissue endogenous myostatin of human, mouse and rat. A specific 40 kD band was detected in chicken muscle, which suggested that chicken myostatin might have different splicing pattern. Immunofluorescence assay indicated that myostatin predominantly existed in the cytosol in C2C12 cells. Taken together, the results show that myostatin inhibits chicken muscle cells proliferation and differentiation and down-regulates expression of two differentiation marker gene in CEMs. Remarkably, production of functional recombinant myostatin protein and its specific antibody provides important reagents for unraveling molecular mechanisms underlying myostatin action during myogenesis.Wei Yang, Kun Wang, Yan Chen, Yong Zhang, Bo Huang, Da-Hai Zhu