Liposome-encapsulated clodronate eliminates more than 80% of blood monocytes. (A) Rats received daily injections of liposome-encapsulated clodronate starting 24 h prior to intra-muscular injection of bupivacaine until euthanasia at 4 d post-injury. Clodronate treatment induces a decrease of the CD3-CD45RA- non-lymphoid cell frequency (A, upper lane), and of the frequency of CD11b+CD68+ monocytes found in the CD3-CD45RA- population (A, lower lane). Gates were set using either omission of primary antibodies (omission of CD3/CD45RA or CD11b) and using an isotype control for CD68. n = 3. (B) When expressed relatively to total PBMCs, the frequency of CD3-CD45RA-CD11b+CD68+ monocytes was decreased by 90% by the clodronate treatment in injured animals, in comparison to uninjured (*) and injured non-depleted (#) animals. Student’s t-test, p < 0.05. n = 3. (C) Using the same gating strategy, we shown that clodronate treatment significantly decreased the frequency of CD3-CD45RA-CD11b+CD68+ monocytes at 2 d and 3 d by 86% and 82%, respectively in injured animals, in comparison to uninjured (*) and injured non-depleted (#) animals. Student’s t-test, p < 0.05. n = 3.