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e for X-ray or CBCT for pretreatment verification for treatment of anatomical sites that are rigid. 3D3D using CBCT had the best image registration accuracy among all the tested methods. 2D3D method was slightly inferior to the 3D3D method but was still acceptable as a treatment position verification device. 3DS is comparable to 2D3D technique and could be a substitute for X-ray or CBCT for pretreatment verification for treatment of anatomical sites that are rigid.Environmental conditions experienced by developing animals have an impact on the dev