166Ho-1,4,7,10-tetraazacyclododecane-1,4,7,10- tetramethylenephosphonate (DOTMP) is a tetraphosphonate molecule radiolabeled with 166Ho that localizes to bone surfaces. This study evaluated pharmacokinetics and radiation-absorbed dose to all organs from this β-emitting radiopharmaceutical. Methods: After two 1.1-GBq administrations of 166Ho-DOTMP, data from whole-body counting using a γ-camera or uptake probe were assessed for reproducibility of whole-body retention in 12 patients with multiple myeloma. The radiation-absorbed dose to normal organs was estimated using MIRD methodology, applying residence times and S values for 166Ho. Marrow dose was estimated from measured activity retained after 18 h. The activity to deliver a therapeutic dose of 25 Gy to the marrow was determined. Methods based on region-of-interest (ROI) and wholebody clearance were evaluated to estimate kidney activity, because the radiotracer is rapidly excreted in the urine. The dose to the surface of the bladder wall was estimated using a dynamic bladder model. Results: In clinical practice, γ-camera methods were more reliable than uptake probe-based methods for whole-body counting. The intrapatient variability of dose calculations was less than 10% between the 2 tracer studies. Skeletal uptake of 166Ho-DOTMP varied from 19% to 39% (mean, 28%). The activity of 166Ho prescribed for therapy ranged from 38 to 67 GBq (1,030-1,810 mCi). After high-dose therapy, the estimates of absorbed dose to the kidney varied from 1.6 to 4 Gy using the whole-body clearance-based method and from 8.3 to 17.3 Gy using the ROI-based method. Bladder dose ranged from 10 to 20 Gy, bone surface dose ranged from 39 to 57 Gy, and doses to other organs were less than 2 Gy for all patients. Repetitive administration had no impact on tracer biodistribution, pharmacokinetics, or organ dose. Conclusion: Pharmacokinetics analysis validated γ-camera whole-body counting of 166Ho as an appropriate approach to assess clearance and to estimate radiation-absorbed dose to normal organs except the kidneys. Quantitative g-camera imaging is difficult and requires scatter subtraction because of the multiple energy emissions of 166Ho. Kidney dose estimates were approximately 5-fold higher when the ROI-based method was used rather than the clearance-based model, and neither appeared reliable. In future clinical trials with 166Ho-DOTMP, we recommend that dose estimation based on the methods described here be used for all organs except the kidneys. Assumptions for the kidney dose require further evaluation.