Proteomic Profiling of Rat Thyroarytenoid Muscle Purpose Proteomic methodologies offer promise in elucidating the systemwide cellular and molecular processes that characterize normal and diseased thyroarytenoid (TA) muscle. This study examined methodological issues central to the application of 2-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (2D SDS-PAGE) to the study of the TA muscle proteome using a ... Research Article
Research Article  |   June 01, 2006
Proteomic Profiling of Rat Thyroarytenoid Muscle
 
Author Affiliations & Notes
  • Nathan V. Welham
    University of Wisconsin—Madison
  • Gerard Marriott
    University of Wisconsin—Madison
  • Diane M. Bless
    University of Wisconsin—Madison
  • Contact author: Gerard Marriott, Department of Physiology, University of Wisconsin—Madison, 2385 Medical Sciences Center, 1300 University Avenue, Madison, WI 53706. Email: gm@physiology.wisc.edu
Article Information
Speech, Voice & Prosody / Speech / Research Articles
Research Article   |   June 01, 2006
Proteomic Profiling of Rat Thyroarytenoid Muscle
Journal of Speech, Language, and Hearing Research, June 2006, Vol. 49, 671-685. doi:10.1044/1092-4388(2006/049)
History: Received May 21, 2005 , Accepted November 10, 2005
 
Journal of Speech, Language, and Hearing Research, June 2006, Vol. 49, 671-685. doi:10.1044/1092-4388(2006/049)
History: Received May 21, 2005; Accepted November 10, 2005
Web of Science® Times Cited: 6

Purpose Proteomic methodologies offer promise in elucidating the systemwide cellular and molecular processes that characterize normal and diseased thyroarytenoid (TA) muscle. This study examined methodological issues central to the application of 2-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (2D SDS-PAGE) to the study of the TA muscle proteome using a rat model.

Method 2D SDS-PAGE was performed using 4 chemically skinned rat TA muscle samples. Gel images were analyzed and compared. Protein spot detection and matching were performed using computational image analysis algorithms only and computational image analysis followed by visual inspection and manual error correction. A synthetic master gel, constructed to control for uninteresting biological variation and technical artifact due to differences in protein loading and staining, was evaluated against its constituent gels.

Results Manual error correction resulted in a consistent increase in the number of protein spots detected (between 5.8% and 40.9%) and matched (from 25.8% to 70.8%) across all gels. Sensitivity and specificity of the automatic (computational) spot detection procedure, evaluated against the manual correction procedure, were 74.1% and 97.9%, respectively. Evaluation of protein quantitation parameter values revealed statistically significant differences (p < .0001) in optical density, area, and volume for matched protein spots across gels. The synthetic master gel successfully compensated for these intergel differences.

Conclusions Valid and reliable proteomic data are dependant on well-controlled manageable variability and well-defined unmanageable variability. Manual correction of spot detection and matching errors and the use of a synthetic master gel appear to be useful strategies in addressing these issues. With these issues accounted for, 2D SDS-PAGE may be applied to quantitative experimental comparisons of normal and disease conditions affecting voice, speech, and swallowing function.

Acknowledgments
This study was supported by National Institute on Deafness and Other Communication Disorders Grant R01 DC004428, National Heart, Lung, and Blood Institute Grant R01 HL069970, and the Robert Zeek Memorial Laryngeal and Voice Research Fund in the Division of Otolaryngology, Department of Surgery, University of Wisconsin—Madison. We acknowledge laboratory services provided by Kendrick Laboratories, Madison, WI; statistical consultation provided by Alejandro Muñoz del Río and Glen Leverson, Department of Surgery, University of Wisconsin—Madison; and interjudge agreement measurements performed by Ichiro Tateya, Division of Otolaryngology, Department of Surgery, University of Wisconsin—Madison.
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