In order to segment reaction holes from the card, a segmentation algorithm of reaction holes based on Lab and HSV color spaces was proposed in this pepper. The method uses the means of OTSU threshold segmentation in Lab color space and global threshold segmentation in HSV according to the characteristic of reaction holes image, then combines two segmentation parts together as a whole and filters the noise, the target area was acquired finally. Experimental showed that this method can segment reaction holes precisely and restrain the influence of noise or uneven illumination.