Generation and Characterization of Ptf1a Antiserum and Localization of Ptf1a in Relation to Nkx6.1 and Pdx1 During the Earliest Stages of Mouse Pancreas Development
Jacob Hald 1, Anne Ejrnæs Sprinkel 1, Michael Ray 1, Palle Serup 1, Chris Wright 1 and Ole D. Madsen 1*
1 Department of Developmental Biology (JH,PS,ODM) and Antibody Core (JH,AES,ODM), Hagedorn Research Institute, Gentofte, Denmark; Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, Tennessee (MR,CW); and Antibody Technology, Novo Nordisk A/S, Måløv, Denmark (AES)
* To whom correspondence should be addressed. E-mail: odm{at}hagedorn.dk.
Submitted on December 20, 2007
Accepted on 5 March 2008
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Abstract |
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Ptf1a and Pdx1 are critical transcription factors of early pancreatic development, as shown by loss of function studies where lack of each gene alone causes almost complete pancreas agenesis. Ptf1a is particularly interesting as it is linked to a recently reported signature gene expression profile associated with the multipotent condition (Zhou et al. 2007). Few useful antibody reagents have been available for consistent and reliable immunohistochemical visualization of Ptf1a protein expression in the early developing pancreas, in which the level of production of this critical regulator seems to be very low. We describe a novel rabbit antibody raised against the c-terminal portion of the mouse Ptf1a protein and report for the first time the immunodetection as early as e8.5 - e8.75 in the dorsal and ventral buds of the mouse pancreas as well as in the neural tube at e10.0. Detailed confocal analysis identifies an abundant triple-positive (Ptf1a+/Nkx6.1+/Pdx1+) putative early multipotent pancreatic progenitor cell that marks the e9.5 dorsal pancreas and e10.5 ventral pancreas. Furthermore, the expression patterns of Nkx6.1 versus Ptf1a subsequently segregate during branching morphogenesis (trunk versus tip), ending up marking two distinct cell populations of progenitors at e12.5. From day e15.5 (mouse) and in adult pancreas (mouse, rat, human) the Ptf1a antibody marks only acinar cell nuclei, as expected for its subsequent role in committing/maintaining cells in this differentiated state. In summary, this antibody is a novel tool to further characterize important early steps of pancreas differentiation. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.
Key Words:
Ptf1a, p48, pancreas, whole mount, antibody, Nkx6.1, Pdx1, progenitor, stem cell
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