- In previous studies classification algorithms were employed successfully for the detection of unknown malicious code. Most of these studies extracted features based on byte n-gram patterns in order to represent the inspected files. In this study we represent the inspected files using OpCode n-gram patterns which are extracted from the files after disassembly. The OpCode n-gram patterns are used as features for the classification process. The classification process main goal is to detect unknown malware within a set of suspected files which will later be included in antivirus software as signatures. A rigorous evaluation was performed using a test collection comprising of more than 30,000 files, in which various settings of OpCode n-gram patterns of various size representations and eight types of classifiers were evaluated. A typical problem of this domain is the imbalance problem in which the distribution of the classes in real life varies. We investigated the imbalance problem, referring to several real-life scenarios in which malicious files are expected to be about 10% of the total inspected files. Lastly, we present a chronological evaluation in which the frequent need for updating the training set was evaluated. Evaluation results indicate that the evaluated methodology achieves a level of accuracy higher than 96% (with TPR above 0.95 and FPR approximately 0.1), which slightly improves the results in previous studies that use byte n-gram representation. The chronological evaluation showed a clear trend in which the performance improves as the training set is more updated.