אין מימון להכריז.

<ol>
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Determined by neutron activation analysis on needle biopsy specimens.</a> Scand J Clin Lab Invest (England). 14, (1962).</li><li>Bergstrom, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Percutaneous+needle+biopsy+of+skeletal+muscle+in+physiological+and+clinical+research).">Percutaneous needle biopsy of skeletal muscle in physiological and clinical research).</a> Scand J Clin Lab Invest. 35, 609-616 (1975).</li><li>Bergstrom, J., Hultman, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+study+of+the+glycogen+metabolism+during+exercise+in+man.">A study of the glycogen metabolism during exercise in man.</a> Scand J Clin Lab Invest. 19, 218-228 (1967).</li><li>Evans, W. J., Phinney, S. D., Young, V. 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C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+carbohydrate+ingestion+on+immune+changes+after+2+h+of+intensive+resistance+training.">Influence of carbohydrate ingestion on immune changes after 2 h of intensive resistance training.</a> J Appl Physiol. 96, 1292-1298 (2004).</li><li>Utter, A. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Carbohydrate+supplementation+and+perceived+exertion+during+prolonged+running.">Carbohydrate supplementation and perceived exertion during prolonged running.</a> Med Sci Sports Exerc. 36, 1036-1041 (2004).</li><li>Nieman, D. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Muscle+cytokine+mRNA+changes+after+2.5+h+of+cycling:+influence+of+carbohydrate.">Muscle cytokine mRNA changes after 2.5 h of cycling: influence of carbohydrate.</a> Med Sci Sports Exerc. 37, 1283-1290 (2005).</li><li>Nieman, D. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+quercetin+and+EGCG+on+mitochondrial+biogenesis+and+immunity.">Effects of quercetin and EGCG on mitochondrial biogenesis and immunity.</a> Med Sci Sports Exerc. 41, 1467-1475 (2009).</li><li>Nieman, D. 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L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Muscle+glycogen+utilization+during+prolonged+strenuous+exercise+when+fed+carbohydrate.">Muscle glycogen utilization during prolonged strenuous exercise when fed carbohydrate.</a> J Appl Physiol. 61, 165-172 (1985).</li></ol>

The authors have no conflicts of interest.

וידאו זה מספק סיכום צעד אחר צעד של הליך ביופסיה של שריר השלד המשמש במעבדה ביצועי אדם ASU. הליך זה, עם שינויים קטנים, נעשה שימוש כדי לאסוף כ 1,600 ביופסיות שריר במהלך 13 השנים האחרונות. דגימות הביופסיה של השריר סיפקו נתונים חשובים בחקירות מבוססות תזונת ספורט, שהובילו לתגליו?...

טכניקת הביופסיה מלעורית או &#39;פתוחה למחצה &quot;משמשת כדי להשיג דגימות רקמת שריר שלד מחולים אנושיים ונושאי מחקר. דושן (1806-1875) הוא זוכה כראשון לבניית מחט עם נקז כדי להשיג שרירי שלד מנושאים המתגוררים דרך הביופסיה מלעורית 1. בשנתי ה -1960, Bergstrom הציג מחט ביופסיה מלעורית דומה לזה שתואר על ידי דושן 2-4. עשרים שנה מאוחר יותר, אוונס et al. 5 הותאם על ידי יישום טכניקת יניקה דרך נקז החיתוך של מחט Bergstrom. שינוי זה יכול לשפר את התשואה של רקמה 3 עד 5 פי 6,7, והוא מועסק בהגדרות מחקר קליניות וביו. טכניקה זו יש ותמשיך לטפח את האבחנה של myopathies וההבנה של המבנה והתפקוד של שרירי שלד שלנו. טכניקת הביופסיה של שריר מלעורית היא straightforward. כאשר נעשו בצורה נכונה, עם הקפדה על טכניקת ספטית, הסיכונים הכרוכים בכך הם מינימאליים. הליך הביופסיה של השריר נחשב ככלי מחקרי לפרויקטי מחקר. חלק מהמוסדות האקדמיים מאפשרים מאומנים חוקרי פקולטה עם תואר דוקטור, עם פיקוח רופא ישיר, כדי להשיג ביופסיות שריר ואחרים דורשים רופא מוסמך לוח כדי לבצע את הטכניקה. צוות הביופסיה ASU ביצע בהצלחה מעל 1,600 ביופסיות שריר במהלך 13 השנים האחרונות 8-15. המטרה של הווידאו הזה היא לתאר את טכניקת הביופסיה של שריר מלעורית מחט Bergstrom הותאם 5 להשיג דגימות רקמת שריר שלד מlateralis vastus של בני אדם.

<table><tbody><tr><td>Bergstr&amp;ouml;m biopsy needle, 5 x 100 mm</td><td>Stille Surgical Inc.</td><td>119-29187-50</td><td>1 ea</td></tr><tr><td>Durasorb blue pad</td><td>Fisher</td><td>22-031-340</td><td>case of 300</td></tr><tr><td>Prep razor disposable</td><td>Moore Medical</td><td>89760</td><td>1 ea</td></tr><tr><td>Shave cream</td><td>Moore Medical</td><td>92869</td><td>1 can</td></tr><tr><td>Alcohol swab</td><td>Moore Medical</td><td>98721</td><td>box/200</td></tr><tr><td>Povidone-iodine swabsticks (3 pack)</td><td>Moore Medical</td><td>90691</td><td>pack of 50</td></tr><tr><td>Face masks</td><td>Fisher</td><td>19-039-690</td><td>case of 50</td></tr><tr><td>Sterile gloves; sz 6.5</td><td>Moore Medical</td><td>68128</td><td>box of 50 pairs</td></tr><tr><td>Sterile gloves; sz 7.0</td><td>Moore Medical</td><td>68129</td><td>box of 50 pairs</td></tr><tr><td>Sterile gloves; sz 8.5</td><td>Moore Medical</td><td>68132</td><td>box of 50 pairs</td></tr><tr><td>Fenestrated towel drape</td><td>Moore Medical</td><td>92712</td><td>case of 50</td></tr><tr><td>Lidocaine (1% HCl, w/o Epi)</td><td>Dealmed</td><td>427902</td><td>30 cc vial</td></tr><tr><td>Ethyl chloride spray</td><td>Dealmed</td><td>386020</td><td>3.5 fl. Oz. bottle</td></tr><tr><td>5 ml syringe w/ 21 G needle</td><td>Fisher</td><td>14-827-48</td><td>case of 100</td></tr><tr><td>25 G 5/8&quot; needle</td><td>Fisher</td><td>14-826AA</td><td>case of 100</td></tr><tr><td>25 G 1 1/2&quot; needle</td><td>Fisher</td><td>14-826-49</td><td>case of 100</td></tr><tr><td>Single-use scalpels w/ #11 blade</td><td>Fisher</td><td>0-8927-5B</td><td>box of 20</td></tr><tr><td>60 ml Syringe</td><td>Fisher</td><td>13-689-8</td><td>case of 40</td></tr><tr><td>~30 inch extension tubing</td><td>Fisher</td><td>14-169-7A</td><td>roll of 50 ft</td></tr><tr><td>Metal stopcock</td><td>Fisher</td><td>01-290-38</td><td>each</td></tr><tr><td>Polypropylene tubing connector w/ tapered ends, 3.2 to 5.5 mm</td><td>Fisher</td><td>15-315-28A</td><td>case of 100</td></tr><tr><td>Yellow pipette tips (1-200 &amp;micro;l)</td><td>Fisher</td><td>02-681-2</td><td>cut off ~15-18 mm of the tip</td></tr><tr><td>Topical surgical adhesive</td><td>Medline</td><td>MSC091076</td><td>case of 36 tubes</td></tr><tr><td>Topical surgical adhesive</td><td>Medline</td><td>MSC091076Z</td><td>box of 6 tubes</td></tr><tr><td>Butterfly stitches (adhesive strips) 1/2&quot; x 4&quot; (6 per pack)</td><td>Moore Medical</td><td>93050</td><td>box/300</td></tr><tr><td>Bandadge, Strip, 3/4 x 3 in.</td><td>Fisher</td><td>19-027-202</td><td>pack of 100</td></tr><tr><td>Adhesive bandage (5 m rolls)</td><td>Fisher</td><td>19-027-768</td><td>case of 36</td></tr><tr><td>1&quot; paper surgical tape</td><td>Fisher</td><td>19-027-761</td><td>case of 12</td></tr><tr><td>Sutures (4-0 reverse cutting, FS2)</td><td>Moore Medical</td><td>82926</td><td>box of 36</td></tr><tr><td>4 x 4 Sterile gauze pads</td><td>Moore Medical</td><td>66949</td><td>pkg/50</td></tr><tr><td>2 x 2 Non-sterile gauze pads</td><td>Moore Medical</td><td>37336</td><td>pkg/200</td></tr><tr><td>3 x 4 Non-adherent pads</td><td>Moore Medical</td><td>74389</td><td>bx/100</td></tr><tr><td>Autoclave bags 3.5 x 9</td><td>Moore Medical</td><td>40390</td><td>box of 200</td></tr><tr><td>Autoclave bags 5.25 x 10</td><td>Moore Medical</td><td>40391</td><td>box of 200</td></tr><tr><td>Autoclave indicator tape</td><td>Moore Medical</td><td>31608</td><td>box of 250</td></tr><tr><td>Lab coats&amp;mdash;small</td><td>Fisher</td><td>23-900-520A&amp;nbsp;</td><td>10/pk</td></tr><tr><td>Lab coats&amp;mdash;medium</td><td>Fisher</td><td>23-900-520B&amp;nbsp;</td><td>10/pk</td></tr><tr><td>Lab coats&amp;mdash;large</td><td>Fisher</td><td>23-900-520C&amp;nbsp;</td><td>10/pk</td></tr><tr><td>Germicidal Disposable Cloth Wipes</td><td>Moore Medical</td><td>62879</td><td>Can/160</td></tr></tbody></table>

human skeletal muscle biopsy procedures using the modified bergstr&#246;m technique

טכניקת הביופסיה מלעורית מאפשרת לחוקרים ולרופאים כדי לאסוף דגימות רקמת שריר שלד. הטכניקה היא בטוחה ויעילה ביותר. וידאו זה מתאר את טכניקת הביופסיה מלעורית באמצעות מחט Bergstrom שונה כדי להשיג דגימות רקמת שריר שלד מlateralis vastus של בני אדם. מחט Bergstrom מורכבת מצינורית חיצונית עם פתח קטן (&#39;חלון&#39;) בצד של הקצה ונקז פנימי עם להב חיתוך בקצה הדיסטלי. בתנאי הרדמה וספטית מקומיים, המחט התקדמה לתוך שריר השלד באמצעות חתך בעור, הרקמה התת עורית, וfascia. בשלב הבא, היניקה מוחלת על הנקז הפנימי, הנקז החיצוני משוך לאחור, רקמת שריר שלד הוא נשאבה לתוך החלון של הצינורית החיצונית על ידי היניקה, והנקז הפנימי סגור במהירות, ובכך חיתוך או גזיר דגימת רקמת שריר השלד . המחט מסובבת 90 °60; וקיצוץ נוסף הוא עשה. תהליך זה ניתן לחזור עוד שלוש פעמים. טכניקת חיתוך מרובה זה בדרך כלל מייצרת מדגם של 100-200 מ&quot;ג או יותר בנבדקים בריאים ויכולה להיעשות באופן מיידי לפני, במהלך, ואחרי התקף של פעילות גופנית או התערבות אחרת. בעקבות הלבשה לאחר ביופסיה של אתר החתך, נושאים בדרך כלל לחדש את פעילותם של חיים היום יום באופן מיידי ויכולים להשתתף באופן מלא בפעילות גופנית נמרצת בתוך 48-72 שעה. נושאים צריכים להימנע מפעילות גופני התנגדות כבדה ל48 שעות כדי להפחית את הסיכון של פריצת של השריר דרך החתך בfascia.

הליך הביופסיה של השריר כפי שתואר לעיל מאפשר לחוקר לאסוף במהירות ובאופן עקבי דגימות רקמת שריר שלד. התשואה האופיינית בנבדקים בריאים, אתלטי היא 200 מ&quot;ג או יותר במעבר אחד עם 3-4 סרטונים. ההליך אורך 15-20 דקות, שרובו בילו בהכנה לחתך. במחקרים מבוססי תרגיל, דגימות שריר לעתים קרו...

Watch this Scientific Journal Video about Human Skeletal Muscle Biopsy Procedures Using the Modified BergstrÃƒÂ¶m Technique at JoVE.com

Human Skeletal Muscle Biopsy Procedures Using the Modified Bergstr&#246;m Technique

The purpose of this video is to present the modified Bergstr&#246;m skeletal muscle biopsy technique on human subjects.

נהלי ביופסיה של שריר שלד אנושי באמצעות טכניקת Bergstrom השתנה

The percutaneous biopsy technique enables researchers and clinicians to collect skeletal muscle tissue samples. The technique is safe and highly ...

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39.8K Views.  Appalacian State University. The purpose of this video is to present the modified Bergström skeletal muscle biopsy technique on human subjects. 

Video: Human Skeletal Muscle Biopsy Procedures Using the Modified Bergström Technique

Isolation and Quantitative Immunocytochemical Characterization of Primary Myogenic Cells and Fibroblasts from Human Skeletal Muscle

The repair and regeneration of skeletal muscle requires the action of satellite cells, which are the resident muscle stem cells. These can be isolated from human muscle biopsy samples using enzymatic digestion and their myogenic properties studied in culture. Quantitatively, the two main adherent cell types obtained from enzymatic digestion are: (i) the satellite cells (termed myogenic cells or muscle precursor cells), identified initially as CD56+ and later as CD56+/desmin+ cells and (ii) muscle-derived fibroblasts, identified as CD56&#8211; and TE-7+. Fibroblasts proliferate very efficiently in culture and in mixed cell populations these cells may overrun myogenic cells to dominate the culture. The isolation and purification of different cell types from human muscle is thus an important methodological consideration when trying to investigate the innate behavior of either cell type in culture. Here we describe a system of sorting based on the gentle enzymatic digestion of cells using collagenase and dispase followed by magnetic activated cell sorting (MACS) which gives both a high purity (&#62;95% myogenic cells) and good yield (~2.8 x 106 &#177; 8.87 x 105 cells/g tissue after 7 days in vitro) for experiments in culture. This approach is based on incubating the mixed muscle-derived cell population with magnetic microbeads beads conjugated to an antibody against CD56 and then passing cells though a magnetic field. CD56+ cells bound to microbeads are retained by the field whereas CD56&#8211; cells pass unimpeded through the column. Cell suspensions from any stage of the sorting process can be plated and cultured. Following a given intervention, cell morphology, and the expression and localization of proteins including nuclear transcription factors can be quantified using immunofluorescent labeling with specific antibodies and an image processing and analysis package.

The main adherent cell types derived from human muscle are myogenic cells and fibroblasts. Here, cell populations are enriched using magnetic-activated cell sorting based on the CD56 antigen. Subsequent immunolabelling with specific antibodies and use of image analysis techniques allows quantification of cytoplasmic and nuclear characteristics in individual cells.

בידוד וכמוני Immunocytochemical אפיון של תאים ראשוניים myogenic וFibroblasts משרירי שלד אדם

The repair and regeneration of skeletal muscle requires the action of satellite cells, which are the resident muscle stem cells. These can be isolated ...

Developmental Biology

Myo-mechanical Analysis of Isolated Skeletal Muscle

To assess the in vivo effects of therapeutic interventions for the treatment of muscle disease 1,2,3, quantitative methods are needed that measure force generation and fatigability in treated muscle. We describe a detailed approach to evaluating myo-mechanical properties in freshly explanted hindlimb muscle from the mouse. We describe the atraumatic harvest of mouse extensor digitorum longus muscle, mounting the muscle in a muscle strip myograph (Model 820MS; Danish Myo Technology), and the measurement of maximal twitch and tetanic tension, contraction time, and half-relaxation time, using a square pulse stimulator (Model S48; Grass Technologies). Using these measurements, we demonstrate the calculation of specific twitch and tetanic tension normalized to muscle cross-sectional area, the twitch-to-tetanic tension ratio, the force-frequency relationship curve and the low frequency fatigue curve 4. This analysis provides a method for quantitative comparison between therapeutic interventions in mouse models of muscle disease 1,2,3,5, as well as comparison of the effects of genetic modification on muscle function 6,7,8,9.

To assess the in vivo effects of therapeutic interventions for muscle disease, methods are needed to quantitate force generation and fatigability in treated muscle. We detail an approach to evaluating myo-mechanical properties in explanted mouse hindlimb muscle. This analysis provides a robust approach to quantitating the effects of genetic modification on muscle function, as well as comparison of therapies in mouse models of muscle disease.

מיו מכני ניתוח בשרירי השלד מבודדת

To assess the in vivo effects of therapeutic interventions for the treatment of muscle disease 1,2,3, quantitative methods are needed that measure force ...

X-ray Diffraction of Intact Murine Skeletal Muscle as a Tool for Studying the Structural Basis of Muscle Disease

Transgenic mouse models have been important tools for studying the relationship of genotype to phenotype for human diseases including those of skeletal muscle. Mouse skeletal muscle has been shown to produce high quality X-ray diffraction patterns on third generation synchrotron beamlines providing an opportunity to link changes at the level of the genotype to functional phenotypes in health and disease by determining the structural consequences of genetic changes. We present detailed protocols for preparation of specimens, collecting the X-ray patterns and extracting relevant structural parameters from the X-ray patterns that may help guide experimenters wishing to perform such experiments for themselves.

We present detailed protocols for performing small-angle X-ray diffraction experiments using intact mouse skeletal muscles. With the wide availability of transgenic mouse models for human diseases, this experimental platform can form a useful test bed for elucidating the structural basis of genetic muscle diseases

צילום רנטגן של שריר השלד Murine שלמים ככלי לחקר בסיס מבניים של מחלת שרירים

Transgenic mouse models have been important tools for studying the relationship of genotype to phenotype for human diseases including those of skeletal ...

Isolation, Culture, Characterization, and Differentiation of Human Muscle Progenitor Cells from the Skeletal Muscle Biopsy Procedure

The use of primary human tissue and cells is ideal for the investigation of biological and physiological processes such as the skeletal muscle regenerative process. There are recognized challenges to working with human primary adult stem cells, particularly human muscle progenitor cells (hMPCs) derived from skeletal muscle biopsies, including low cell yield from collected tissue and a large degree of donor heterogeneity of growth and death parameters among cultures. While incorporating heterogeneity into experimental design requires a larger sample size to detect significant effects, it also allows us to identify mechanisms that underlie variability in hMPC&#160;expansion capacity, and thus allows us to better understand heterogeneity in skeletal muscle regeneration. Novel mechanisms that distinguish the expansion capacity of cultures have the potential to lead to the development of therapies to improve skeletal muscle regeneration.

We present techniques for isolating, culturing, characterizing, and differentiating human primary muscle progenitor cells (hMPCs) obtained from skeletal muscle biopsy tissue. hMPCs obtained and characterized through these methods can be used to subsequently address research questions related to human myogenesis and skeletal muscle regeneration.

בידוד, תרבות, אפיון, והבידול של תאים האדם שריר ביופסיה של השלד שרירי השגרה נוהל

The use of primary human tissue and cells is ideal for the investigation of biological and physiological processes such as the skeletal muscle ...

Performing Human Skeletal Muscle Xenografts in Immunodeficient Mice

Treatment effects observed in animal studies often fail to be recapitulated in clinical trials. While this problem is multifaceted, one reason for this failure is the use of inadequate laboratory models. It is challenging to model complex human diseases in traditional laboratory organisms, but this issue can be circumvented through the study of human xenografts. The surgical method we describe here allows for the creation of human skeletal muscle xenografts, which can be used to model muscle disease and to carry out preclinical therapeutic testing. Under an Institutional Review Board (IRB)-approved protocol, skeletal muscle specimens are acquired from patients and then transplanted into NOD-Rag1null IL2r&gamma;null (NRG) host mice. These mice are ideal hosts for transplantation studies due to their inability to make mature lymphocytes and are thus unable to develop cell-mediated and humoral adaptive immune responses. Host mice are anaesthetized with isoflurane, and the mouse tibialis anterior and extensor digitorum longus muscles are removed. A piece of human muscle is then placed in the empty tibial compartment and sutured to the proximal and distal tendons of the peroneus longus muscle. The xenografted muscle is spontaneously vascularized and innervated by the mouse host, resulting in robustly regenerated human muscle that can serve as a model for preclinical studies.

Complex human diseases can be challenging to model in traditional laboratory model systems. Here, we describe a surgical approach to model human muscle disease through the transplantation of human skeletal muscle biopsies into immunodeficient mice.

ביצוע שריר השלד האנושי Xenografts בעכברים חיסוני

Treatment effects observed in animal studies often fail to be recapitulated in clinical trials. While this problem is multifaceted, one reason for this ...

Quantification of Subcellular Glycogen Distribution in Skeletal Muscle Fibers using Transmission Electron Microscopy

With the use of transmission electron microscopy, high-resolution images of fixed samples containing individual muscle fibers can be obtained. This enables quantifications of ultrastructural aspects such as volume fractions, surface area to volume ratios, morphometry, and physical contact sites of different subcellular structures. In the 1970s, a protocol for enhanced staining of glycogen in cells was developed and paved the way for a string of studies on the subcellular localization of glycogen and glycogen particle size using transmission electron microscopy. While most analyses interpret glycogen as if it is homogeneously distributed within the muscle fibers, providing only a single value (e.g., an average concentration), transmission electron microscopy has revealed that glycogen is stored as discrete glycogen particles located in distinct subcellular compartments. Here, the step-by-step protocol from tissue collection to the quantitative determination of the volume fraction and particle diameter of glycogen in the distinct subcellular compartments of individual skeletal muscle fibers is described. Considerations on how to 1) collect and stain tissue specimens, 2) perform image analyses and data handling, 3) evaluate the precision of estimates, 4) discriminate between muscle fiber types, and 5) methodological pitfalls and limitations are included.

A modified post-fixation procedure increases the contrast of glycogen particles in tissue. This paper provides a step-by-step protocol describing how to handle the tissue, conduct the imaging, and use stereological methods to obtain unbiased and quantitative data on fiber type-specific subcellular glycogen distribution in skeletal muscle.

כימות של התפלגות גליקוגן תת-תאית בסיבי שריר השלד באמצעות מיקרוסקופיית אלקטרונים שידור

With the use of transmission electron microscopy, high-resolution images of fixed samples containing individual muscle fibers can be obtained. This ...

Biochemistry

Preparation and Respirometric Assessment of Mitochondria Isolated from Skeletal Muscle Tissue Obtained by Percutaneous Needle Biopsy

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining samples of human Vastus lateralis, isolating mitochondria from minimal amounts of skeletal muscle tissue, and plate based respirometric profiling using an extracellular flux (XF) analyzer. Comparison of respirometric profiles obtained using 1.0, 2.5 and 5.0 &#956;g of mitochondria indicate that 1.0 &#956;g is sufficient to measure respiration and that 5.0 &#956;g provides most consistent results based on comparison of standard errors. Western blot analysis of isolated mitochondria for mitochondrial marker COX IV and non-mitochondrial tissue marker GAPDH indicate that there is limited non-mitochondrial contamination using this protocol. The ability to study mitochondrial respirometry in as little as 20 mg of muscle tissue allows users to utilize individual biopsies for multiple study endpoints in clinical research projects.

Methods for biopsy of Vastus lateralis, preparation of purified mitochondria, and respirometric profiling are described. The use of small muscle volume makes this technique suitable for clinical research applications.

ההכנה וRespirometric הערכה של המיטוכונדריה מבודדת מרקמת שריר שלד מתקבלת על ידי Percutaneous מחט ביופסיה

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining ...

MyDoBID - טכניקה חדשנית לזיהוי יעיל של סוגי סיבי שריר

Fiber Type Identification of Human Skeletal Muscle

The technique described here can be used to identify specific myosin heavy chain (MHC) isoforms in segments of individual muscle fibers using dot blotting, hereafter referred to as Myosin heavy chain detection by Dot Blotting for IDentification of muscle fiber type (MyDoBID). This protocol describes the process of freeze-drying human skeletal muscle and isolating segments of single muscle fibers. Using MyDoBID, type I and II fibers are classified with MHCI- and IIa-specific antibodies, respectively. Classified fibers are then combined into fiber type-specific samples for each biopsy.
The total protein in each sample is determined by Sodium Dodecyl-Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE) and UV-activated gel technology. The fiber type of samples is validated using western blotting. The importance of performing protein loading normalization to enhance target protein detection across multiple western blots is also described. The benefits of consolidating classified fibers into fiber type-specific samples compared to single-fiber western blots, include sample versatility, increased sample throughput, shorter time investment, and cost-saving measures, all while retaining valuable fiber type-specific information that is frequently overlooked using homogenized muscle samples. The purpose of the protocol is to achieve accurate and efficient identification of type I and type II fibers isolated from freeze-dried human skeletal muscle samples.
These individual fibers are subsequently combined to create type I and type II fiber type-specific samples. Furthermore, the protocol is extended to include the identification of type IIx fibers, using Actin as a marker for fibers that were negative for MHCI and MHCIIa, which are confirmed as IIx fibers by western blotting. Each fiber type-specific sample is then used to quantify the expression of various target proteins using western blotting techniques.

מחבר זרקור: פענוח המסתורין של סוגי סיבי שריר השלד באמצעות טכניקת MyDoBID 

This protocol demonstrates single-fiber isolation from freeze-dried human skeletal muscle and fiber-type classification according to Myosin heavy chain (MHC) isoform using the dot blotting technique. Identified MHC I and II fiber samples can then be further analyzed for fiber type-specific differences in protein expression using western blotting.

נהלי ביופסיה של שריר שלד אנושי באמצעות טכניקת Bergstrom השתנה

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