Analysis of subcutaneous fluid in rats

Subcutaneous implantation in rats is a commonly used model for biomaterial calcification studies. Although this model is frequently used, its components have not been characterized with respect to calcification. Exudate from the subcutaneous spaces of 18 young rats was collected using diffusion chambers. These chambers consisted of polymethylmethacrylate tubes with 0.22 micron pore filters covering each end allowing fluid, but not cells, to enter the chambers. Glutaraldehyde treated bovine pericardial strips were implanted subcutaneously, inside the chambers and outside the chambers, to test the calcification inducing abilities of the various environments. The animals were killed on postoperative day 10, and the exudate and materials were collected. The exudate was analyzed for ionic calcium, total calcium, inorganic phosphorus, and albumin, and for cells by a differentiated cell smear. The materials were analyzed for calcification by radiography, histology, and atomic absorption. Calcification was present in the materials inside the chambers where no cells were present and in the materials that were not in chambers. The distinct features of the exudate were elevated ionic calcium, a high Ca x P product, and elevated phosphorus.