title: False-positive beta-galactosidase staining in osteoclasts by endogenous enzyme: studies in neonatal and month-old wild-type mice.
abstract: Escherichia coli beta-galactosidase (beta-gal), encoded by the lacZ gene, has become an essential tool in studies of gene expression and function in higher eukaryotes. lac-Z is widely used as a marker gene to detect expression of transgenes or Cre recombinase driven by tissue-specific promoters. The timing and location of promoter activity is easily visualized in whole embryos or specific tissues using the cleavable, chromogenic substrate, 5-bromo-4-chloro-3-indolyl-D-galactopyranoside (X-gal). The tissue specificity of promoters in transgenic constructs is routinely tested by using a promoter of choice to drive lacZ. Alternatively, the targeted expression of Cre recombinase to perform in vivo recombination of loxP sites can be visualized by beta-gal staining in mice carrying a Cre-activated lacZ transgene, such as the ROSA26 strain. In the course of our investigations, we examined beta-gal activity in bone tissue from genetically normal mice using standard detection methodology and found very high endogenous activity in bone-resorbing osteoclasts. This was true in frozen, paraffin, and glycol methacrylate sections. X-gal staining colocalized with the osteoclast marker, tartrate-resistant acid phosphatase (TRAP). beta-gal activity was present in osteoclasts in long bones, in the mandible, and in both neonatal and more mature animals. We present this brief article as a caution to those testing genetic models of skeletal gene expression using beta-gal as a marker gene.
authors.affiliation: Department of Cell Biology, University of Massachusetts Medical School, Worcester, Massachusetts 01655, USA. paul.odgren@umassmed.edu
authors.forename: Paul R
authors.initials: PR
authors.lastname: Odgren
authors.forename: Carole A
authors.initials: CA
authors.lastname: MacKay
authors.forename: April
authors.initials: A
authors.lastname: Mason-Savas
authors.forename: Meiheng
authors.initials: M
authors.lastname: Yang
authors.forename: Geneviève
authors.initials: G
authors.lastname: Mailhot
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Birnbaum
mesh_terms: D000135:Acid Phosphatase / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D001842:Bone and Bones / Q000201:enzymology; D005189:False Positive Reactions; D006651:Histocytochemistry / Q000379:methods*; D007527:Isoenzymes / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D010010:Osteoclasts / Q000166:cytology / Q000201:enzymology*; D000071681:Tartrate-Resistant Acid Phosphatase; D001616:beta-Galactosidase / Q000378:metabolism*
chemical_list: D007527:Isoenzymes; D000135:Acid Phosphatase; C000605511:Acp5 protein, mouse; D000071681:Tartrate-Resistant Acid Phosphatase; D001616:beta-Galactosidase