Abstract

In attempting to find a secondary way (the primary being ELISA) to test for the presence of monoclonal antibodies against Ehrlichia canis, a test has been conducted to examine the effectiveness of Western Blot analysis. Although previous attempts to blot monoclonals of E. canis have failed, a different procedure has been obtained for the Western Blot. The primary difference between the two procedures is that the new procedure uses dry milk reconstituted in a Net Buffer solution as its primary blocking agent and the base for the dilution of its primary and conjugate antibodies, as opposed to the old procedure which used goat serum. A trial was run using a sample of E. canis infected dog bone marrow cells whose proteins were separated using Gel Electrophoresis and transferred onto a nitrocellulose paper membrane for the Western Blot. Both the old and new procedures were tested using samples from the same gel and the same primary and conjugate antibodies. The test was run on both known polyvalent and known monoclonal antibodies. The results were mixed. The monoclonal antibodies still failed to blot, but, upon initial analysis, it does appear that the new procedure is more clear even for the detection of polyvalent antibodies as it eliminates much of the background staining present in the old procedure.