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Dependency matrix describes the logical relationship between fault and test. Insulation matrix describes demand relationship between fault insulation and test in a given test sequence. The effect of test sequence on diagnostic cost can be calculated by isolation matrix. So solving the isolation matrix is a basic condition for optimizing diagnostic cost. There are several problems of isolation matrix, such as easy to confuse with dependency matrix and difficult to solve with traditional analysis method. In this paper, the concept and generation principle of isolation matrix are detailed, and the generation algorithm based on the breadth first search is designed. At last, an example shows that the algorithm is feasible and effective.