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During the on-line inspection of flat enameled wire surface defects, the jitter of the sample line caused jitter interference when the background was separated using the inter-frame difference method. Common jitter interference processing methods had the problems of too long processing time and false elimination of real defects, so an improved method was proposed. Firstly, the distribution area of the jitter interference was analyzed, and the property that the jitter interference is not on the same line as the real defect and the number of foreground pixels on the line where the jitter interference exists is large. Then, in the distribution area, the rows with the number of foreground pixels exceeding a certain threshold were set to zero to eliminate the jitter interference. The selection of the threshold was obtained by using an improved Otsu processing distribution map. The map was based on the number of foreground pixels on the line for all lines of the image, and the frequency of line appearance was counted. Experiments show that compared with common methods, the improved method with short processing time and does not mistakenly eliminate the real defects connected with jitter interference, and the IoU of the obtained processing results and the ideal results is more than 88%, which has a good application prospect.