Pre-osteoblastic proliferation assessed with BrdU in undecalcified, Epon-embedded adult rat trabecular bone.
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.
Barou
O
O
Laboratoire de Biologie du Tissu Osseux, University of Saint-Etienne, France.
Laroche
N
N
Palle
S
S
Alexandre
C
C
Lafage-Proust
M H
MH
D000818:Animals; D001842:Bone and Bones / Q000737:chemistry*; D001973:Bromodeoxyuridine / Q000032:analysis; D002113:Calcification, Physiologic; D002455:Cell Division; D007150:Immunohistochemistry / Q000379:methods*; D008297:Male; D010006:Osteoblasts / Q000737:chemistry*; D051381:Rats; D017208:Rats, Wistar; D016610:Tissue Embedding / Q000379:methods*
D001973:Bromodeoxyuridine