Spinal cord injury (SCI) has severe impact on millions of people around the world. Finding a cure for the SCI has been difficult but recent research has shown some hope for a cure for patients with SCI. It is currently known that neurons have ability to regenerate their own injured axons. However, the local environment in the lesion is not hospitable for regeneration due to the inhibitory molecules derived from several cell types in the glial scar. After the Injury, the Chondroitin Sulfate Proteoglycan (CSPG), an extracellular matrix produced by various cells in the area, is the major type of inhibitory molecule in the glial scar. The axonal regeneration may be enhanced if the amount of CSPG produced after the SCI can be decreased. Since the enzyme chondroitinase (cABC) can degrades CSPG molecules, cABC's controlled release in the region of the spinal cord injury may be of considerable therapeutic value. Chondroitinase loaded PLGA nanospheres were fabricated using single emulsion method. BSA was not added during the fabrication process, however, small amount of BSA was already exist in the stock enzyme which was purchased from Sigma Aldrich Co. St. Louis MO. Two different total initial cABC concentrations were considered, namely, 300 microliters of 5 unit and 150 microliters of 10 unit cABC. Fabricated nanospheres were freeze dried overnight prior to the release experiments. A typical release experiment was done by adding 5 mg of cABC loaded nanospheres to 1000 microliters of Phosphate Buffer Solution (PBS) and incubating at 37 °C. Each set of experiments were done in triplicates. Periodically, 100 microliters of sample from each tube was taken and mixed with 1 ml of 50 μg/ml of Chondroitin Sulfate solution to perform digestion experiments. 100 microliters of fresh PBS were added to the release tubes to compensate the lost volume by removal of the sample. This routine procedure was continued about 30 days. Digestion experiments were performed by mixing 100 microliters of sample (taken from release experiments) with lml of 50 μg/ml of CS solution. At certain time intervals, 100 microliters of sample was taken from digestion tubes and analyzed using DMMB essay. The concentration of cABC released from nanospheres was calculated using the slope of CS digestion profile which was obtained after each sample of release experiment and calibration curve (between slope and cABC concentration) which was prepared earlier. This procedure can be seen in Figure 1. The cumulative cABC release was calculated for both sets of experiments as shown in Figures 2 and 3. The amount of enzyme released from the first set of experiments (300 microliters of 5 unit cABC) is 7.03×10-03 units and for the second set of experiments is 7.16×10-03 unit. The maximum amount of enzyme that can be possibly released was 6.25×10-02 unit as calculated from the total enzyme added during the fabrication process of nanospheres. This indicates that approximately 11% of the original cABC was released from nanospheres and the enzyme was still active. The remaining 89% either remained in the supernatant during the fabrication process or 89% of the incorporated enzyme was not active. A two step diffusion model has been used to describe the release of cABC from the nanospheres. This model postulates an initial quick (burst) release of enzyme either by diffusion of the enzyme from liquid filled macropores in the nanospheres or desorption of enzyme from the nanosphere surface, followed by a much slower release of enzyme from the nanospheres by diffusion through the PLGA polymer making up the nanospheres. A good fit of the release data to this model was obtained (Figure 4) for burst release of 38% of the total enzyme released, followed by release by diffusion of enzyme through the polymer characterized by an enzyme diffusivity of 3×10-17 cm2/sec.