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Reproductive Sciences
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Progesterone Receptor or Cytoskeletal Protein?

Gemma Madsen, MSc

Mothers and Babies Research Centre, Hunter Medical Research Institute, University of Newcastle and John Hunter Hospital, Newcastle, NSW, Australia, gemma.madsen{at}newcastle.edu.au, Department of Reproductive Biology, Case Western Reserve University, Cleveland, Ohio

David A. MacIntyre, BSc (Biotech) Hons

Mothers and Babies Research Centre, Hunter Medical Research Institute, University of Newcastle and John Hunter Hospital, Newcastle, NSW, Australia, gemma.madsen{at}newcastle.edu.au, Department of Reproductive Biology, Case Western Reserve University, Cleveland, Ohio

Sam Mesiano, PhD

Department of Reproductive Biology, Case Western Reserve University, Cleveland, Ohio

Roger Smith, MBBS, PhD

Mothers and Babies Research Centre, Hunter Medical Research Institute, University of Newcastle and John Hunter Hospital, Newcastle, NSW, Australia, Department of Reproductive Biology, Case Western Reserve University, Cleveland, Ohio

Immunoblotting is used to characterize the various nuclear progesterone receptor (nPR) isoforms present in tissues; however, the success of this technique is dependent on the specificity of the primary nPR antibody. The authors investigate the specificity of a frequently used nPR antibody, sc-538, in total protein from human myometrium and a myometrial cell line (PHM1-31). Using immunoblotting, 2 sc-538 immunoreactive bands at 100 and 55 kDa were detected. The bands were extracted and identified by 1-dimensional liquid chromatography mass spectrometry. The predominant protein in the 100-kDa band was {alpha}-actinin. The dominant proteins in the smaller band were vimentin (57 kDa) and desmin (53 kDa). Myometrial lysate was immunoprecipitated with sc-538, and immunoblotting of the immunoprecipitate with antibodies to {alpha}-actinin, desmin, and vimentin confirmed the presence of these proteins. The sc-538 nPR antibody therefore cross-reacts with cytoskeletal proteins that could be misinterpreted as nPR isoforms. Such misinterpretation has confused the progesterone response literature.

Key Words: Progesterone receptor • immunoblotting • cytoskeletal proteins.

Reproductive Sciences, Vol. 14, No. 3, 217-222 (2007)
DOI: 10.1177/1933719107302380
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