Tartrate-Resistant Acid Phosphatase Staining: An In Vitro Technique to Detect TRAP Enzyme-Containing Cultured Osteoclasts

In This Article

Abstract

Source: Dai, Q., et al. A RANKL-based Osteoclast Culture Assay of Mouse Bone Marrow to Investigate the Role of mTORC1 in Osteoclast Formation. J. Vis. Exp. (2018).

This video demonstrates the tartrate-resistant acid phosphatase staining technique to detect the presence of TRAP-containing granules inside osteoclasts. The technique forms magenta-colored dye granule deposits in the cytoplasm of osteoclasts, rendering them visible under an inverted microscope.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Preparation

Generate osteoclast specific Raptor deletion mice (Raptor^fl/fl; Ctsk-cre, hereafter Rap^Ctsk) by mating Raptor^fl/fl mice with Ctsk-cre mice. Use the Raptor^fl/fl mice as the WT control in this study.
Have a container with ice to keep isolated bone.
Prepare culture medium.
1. Prepare α-MEM culture mediu....

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Results

Microscopy imaging of cell morphology with highlights, diagram, cell analysis, 500 μm and 200 μm scale.
Figure 1: Representative view of osteoclasts. (A) Giant osteoclasts observed in bright field on day 6. The red line outlined the border of osteoclasts. (B) A typically giant, multinucleated, TRAP positive (wine red) osteoclast. The black a.......