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J Histochem Cytochem. 2011 Sep;59(9):807-12. doi: 10.1369/0022155411411713. Epub 2011 May 23.

The sodium-dependent inorganic phosphate transporter SLC34A1 (NaPi-IIa) is not localized in the mouse brain: a case of tissue-specific antigenic cross-reactivity.

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  • 1Department of Anatomy and Centre for Molecular Biology and Neuroscience, University of Oslo, Oslo, Norway.

Abstract

The sodium-dependent inorganic phosphate transporter NaPi-IIa is expressed in the kidney. Here, the authors used a polyclonal antiserum raised against NaPi-IIa- and NaPi-IIa-deficient mice to characterize its expression in nervous tissue. Western blots showed that a NaPi-IIa immunoreactive band (~90 kDa) was only present in wild-type kidney membranes and not in kidney knockout or wild-type brain membranes. In the water-soluble fraction of wild-type and knockout brains, another band (~50 kDa) was observed; this band was not detected in the kidney. Light and electron microscopic immunohistochemistry using the NaPi-IIa antibodies showed immunolabeling of kidney tubules in wild-type but not knockout mice. In the brain, labeling of presynaptic nerve terminals was present also in NaPi-IIa-deficient mice. This labeling pattern was also produced by the NaPi-IIa preimmune serum. The authors conclude that the polyclonal antiserum is specific toward NaPi-IIa in the kidney, but in the brain, immunolabeling is caused by a cross-reaction of the antiserum with an unknown cytosolic protein that is not present in the kidney. This tissue-specific cross-reactivity highlights a potential pitfall when validating antibody specificity using knockout mouse-derived tissue other than the specific tissue of interest and underlines the utility of specificity testing using preimmune sera.

PMID:
21606201
[PubMed - indexed for MEDLINE]
PMCID:
PMC3201171
Free PMC Article

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