Pre-osteoblastic proliferation assessed with BrdU in undecalcified, Epon-embedded adult rat trabecular bone

J Histochem Cytochem. 1997 Sep;45(9):1189-95. doi: 10.1177/002215549704500902.

Abstract

We evaluated bromodeoxyuridine (BrdU) immunohistochemistry in undecalcified adult rat tibiae to study cell kinetics in various bone compartments: primary and secondary spongiosae, periosteum, and bone marrow. Several regimens of BrdU administration were tested (i.p. injections and osmotic minipumps). We compared LR White resin, methylmethacrylate, and Epon-araldite embedding, microwave irradiation for antigen retrieval, several concentrations of sodium ethoxide for deplastification, and various DNA denaturation procedures. Paraffin-embedded decalcified tibiae and Epon-embedded bowel were used as positive controls. The best results were obtained in rats labeled with 40 mg of BrdU for 72 hr using osmotic minipumps. The procedure using a Microprobe system in Eponembedded bone tissue with a sodium ethoxide concentration of 50% for two intervals of 20 min provided the best staining quality and tissue preservation. Labeled pre-osteoblastic cells and bone marrow cells could be counted. Epon embedding allowed preservation of tetracycline double labeling performed 1 to 5 days before sacrifice. The number of labeled pre-osteoblastic cells was correlated with the double-labeled surface area measured histomorphometrically.

MeSH terms

  • Animals
  • Bone and Bones / chemistry*
  • Bromodeoxyuridine / analysis
  • Calcification, Physiologic
  • Cell Division*
  • Immunohistochemistry / methods*
  • Male
  • Osteoblasts / chemistry*
  • Rats
  • Rats, Wistar
  • Tissue Embedding / methods*

Substances

  • Bromodeoxyuridine