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      Muscle Fiber Typing in Bovine and Porcine Skeletal Muscles Using Immunofluorescence with Monoclonal Antibodies Specific to Myosin Heavy Chain Isoforms

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          Abstract

          The aim of this study was to optimize staining procedures for muscle fiber typing efficiently and rapidly in bovine and porcine skeletal muscles, such as longissimus thoracis, psoas major, semimembranosus, and semitendinosus muscles. The commercially available monoclonal anti-myosin heavy chain (MHC) antibodies and fluorescent dye-conjugated secondary antibodies were applied to immunofluorescence histology. Two different procedures, such as cocktail and serial staining, were adopted to immunofluo-rescence analysis. In bovine muscles, three pure types (I, IIA, and IIX) and one hybrid type, IIA+IIX, were identified by the cocktail procedure with a combination of BA-F8, SC-71, BF-35, and 6H1 anti-MHC antibodies. Porcine muscle fibers were typed into four pure types (I, IIA, IIX, and IIB) and two hybrid types (IIA+IIX and IIX+IIB) by a serial procedure with a combination of BA-F8, SC-71, BF-35, and BF-F3. Unlike for bovine muscle, the cocktail procedure was not recommended in porcine muscle fiber typing because of the abnormal reactivity of SC-71 antibody under cocktail procedure. Within the four antibodies, combinations of two or more anti-MHC antibodies allowed us to distinguish pure fiber types or all fiber types including hybrid types. Application of other secondary antibodies conjugated with different fluorescent dyes allowed us to get improved image resolution that clearly distinguished hybrid fibers. Muscle fiber characteristics differed depending on species and muscle types.

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          Most cited references 31

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          Rapid Determination of Myosin Heavy Chain Expression in Rat, Mouse, and Human Skeletal Muscle Using Multicolor Immunofluorescence Analysis

          Skeletal muscle is a heterogeneous tissue comprised of fibers with different morphological, functional, and metabolic properties. Different muscles contain varying proportions of fiber types; therefore, accurate identification is important. A number of histochemical methods are used to determine muscle fiber type; however, these techniques have several disadvantages. Immunofluorescence analysis is a sensitive method that allows for simultaneous evaluation of multiple MHC isoforms on a large number of fibers on a single cross-section, and offers a more precise means of identifying fiber types. In this investigation we characterized pure and hybrid fiber type distribution in 10 rat and 10 mouse skeletal muscles, as well as human vastus lateralis (VL) using multicolor immunofluorescence analysis. In addition, we determined fiber type-specific cross-sectional area (CSA), succinate dehydrogenase (SDH) activity, and α-glycerophosphate dehydrogenase (GPD) activity. Using this procedure we were able to easily identify pure and hybrid fiber populations in rat, mouse, and human muscle. Hybrid fibers were identified in all species and made up a significant portion of the total population in some rat and mouse muscles. For example, rat mixed gastrocnemius (MG) contained 12.2% hybrid fibers whereas mouse white tibialis anterior (WTA) contained 12.1% hybrid fibers. Collectively, we outline a simple and time-efficient method for determining MHC expression in skeletal muscle of multiple species. In addition, we provide a useful resource of the pure and hybrid fiber type distribution, fiber CSA, and relative fiber type-specific SDH and GPD activity in a number of rat and mouse muscles.
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            Three "myosin adenosine triphosphatase" systems: the nature of their pH lability and sulfhydryl dependence.

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              Phenotypic and genetic parameters for longissimus muscle fiber characteristics in relation to growth, carcass, and meat quality traits in large white pigs.

              A total of 383 barrows and gilts from a French Large White experimental herd were slaughtered at 100 kg BW. Samples of longissimus muscle were taken to categorize myofibers according to their contractile (I, IIA, and IIB) and metabolic (oxidative and nonoxidative) properties. Myofiber percentages, cross-sectional areas (CSA), and relative areas were measured. Growth rate, carcass composition, muscle chemical composition, metabolic enzyme activities, and meat quality traits were also measured to estimate phenotypic and genetic correlations between these traits and myofiber characteristics. Genetic parameters were estimated using a REML procedure applied to an individual animal model. Heritabilities of fiber traits were moderate to high (h2 = .20 to .59). Highest heritabilities were found for type I fiber percentage (h2 = .46 +/- .11), type IIBw fiber percentage (h2 = .58 +/- .11), and type I fiber cross-sectional area (h2 = .59 +/- .10). For a given fiber type, the relative area was phenotypically and genetically more closely related to the percentage than to the CSA. Phenotypic correlations between fiber type composition and other traits were low. Genetically, growth rate, carcass leanness, and loin eye area were positively related to fiber CSA. Intramuscular fat content was not related to fiber type composition (r(g) = -.05 to .06), whereas it was positively related to fiber CSA (r(g) = .68). Type IIBw fiber percentage was related to pH at 30 min (r(g) = -.46), pH at 24 h (r(g) = -.62), glycolytic potential (r(g) = .31), and lightness of color (r(g) = .55) of longissimus muscle.
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                Author and article information

                Journal
                Food Sci Anim Resour
                Food Sci Anim Resour
                Food Sci Anim Resour
                kosfa
                Food Science of Animal Resources
                Korean Society for Food Science of Animal Resources
                2636-0772
                2636-0780
                January 2020
                01 January 2020
                : 40
                : 1
                : 132-144
                Affiliations
                [1 ]Graduate School of International Agricultural Technology, Seoul National University , Pyeongchang 25354, Korea
                [2 ]Institutes of Green Bio Science & Technology, Seoul National University , Pyeongchang 25354, Korea
                Author notes
                [* ]Corresponding author: Gap-Don Kim, Graduate School of International Agricultural Technology, Seoul National University, Pyeongchang 25354, Korea, Tel: +82-33-339-5778, Fax: +82-33-339-5779, E-mail: gapdonkim@ 123456snu.ac.kr
                Article
                kosfa-40-1-132
                10.5851/kosfa.2019.e97
                6957451
                © Korean Society for Food Science of Animal Resources

                This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License ( http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

                Funding
                Funded by: CrossRef http://dx.doi.org/10.13039/501100003725, National Research Foundation of Korea;
                Award ID: NRF-2019R1C1C1011056
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                Custom metadata
                2020-01-31

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