The purpose of this study was to assess the contribution of lymphatics to the systemic bioavailability of macromolecules following SC administration in a rat model. The rat model included continuous lymph collection from the thoracic lymph duct and concurrent serial blood sampling from freely moving animals. A thoracic lymph duct-jugular vein shunt produced by an implanted connective cannula, and maintained during the recovery period, enabled superior rat survival and prevented lymphatic cannula occlusion. The SC absorption of three macromolecules (bovine insulin, bovine serum albumin, and recombinant human erythropoietin alpha) was assessed in comparison to the non-lymph cannulated control group. For all tested molecules, only minimal amounts (less than 3%) of the SC administered dose were detected in the collected lymph. In the rat model, following SC administration, the macromolecules were absorbed mainly through the blood capillaries with minimal contribution of the lymphatic system to systemic bioavailability. The relatively small elevation in the lymphatic concentration, which occurred in all molecules, may be attributed to the redistribution of the molecules from the blood to the interstitial fluid compartment. These findings are important since rodents are commonly used in preclinical evaluation of macromolecular drugs.