The application of different staining methods in the morphological observation of non-decalcified bone tissue sections containing implants

Objective To investigate the application and advantages of different staining methods in the morphological observation of non-decalcified bone tissue sections containing implants. Methods Rabbit mandibular bone tissue containing hydroxyapatite were collected and prepared into 50 μm thin sections using a hard tissue grinding system, and after various staining procedures, the sections were sealed with neutral gum and observed under an optical microscope. Results After hematoxylin-eosin staining, the tissue showed a relatively monotonous color, with a red color for bone tissue and an unclear boundary between new bone and original mineralized bone. After Weigert hematoxylin-toluidine blue staining, bone tissue showed a blue color, new bone was deeply stained with a more distinct boundary between new bone and original mineralized bone, and osteoblast nuclei showed a blue color. After Weigert hematoxylin-Ladewig staining, bone tissue showed a blue color, the attached soft tissue showed a yellow color, unmineralized bone and osteoid had a dark yellow color, and osteoblast nuclei appeared yellowish-brown. After Weigert hematoxylin-Van Gieson (VG) staining, bone tissue appeared bright red, new bone was deeply stained, and the attached soft tissue, osteoblasts, osteoid, and unmineralized bone appeared light yellow. After methylene blue-acid fuchsin staining, bone tissue appeared red, new bone was deeply stained, attached soft tissue appeared blue, osteoblast nuclei appeared dark blue, cell matrix appeared light blue, and osteoid appeared purple-gray. After Goldner trichrome staining, bone tissue appeared green, new bone was deeply stained, and osteoid appeared orange-red and orange-yellow alternately. After Von Kossa staining, the areas with calcium salt deposition appeared dark brown, original mineralized bone appeared light gray, and soft tissue, osteoblasts, and loose connective tissue appeared yellowish-brown. Conclusion Weigert hematoxylin-VG staining or Weigert hematoxylin-toluidine blue staining is recommended for high-precision measurement of the ratio of new bone to original mineralized bone and the area of bone integration; methylene blue-acid fuchsin staining or Goldner trichrome staining is recommended for simultaneous observation of the mineralization status of bone tissue (osteoid/new bone/original mi-neralized bone) and the condition of soft tissue/cells; Von Kossa staining is recommended for analyzing the osteogenic activity of bone tissue or observing the calcified area.