Loss of DNA copy number at the short arm of chromosome 3 is one of the most common genetic changes in human lung cancer, suggesting the existence of one or more tumor suppressor genes (TSG) at 3p. To identify most frequently deleted regions and candidate TSGs within these regions, a recently developed single-nucleotide polymorphism (SNP)-mass spectrometry-genotyping (SMSG) technology was applied to investigate the loss of heterozygosity (LOH) in 30 primary non-small-cell lung cancers. A total of 386 SNP markers that spanned a region of 70 Mb at 3p, from 3pter to 3p14.1, were selected for LOH analysis. The average intermarker distance in the present study is approximately 180 kb. Several frequently deleted regions, including 3p26.3, 3p25.3, 3p24.1, 3p23, and 3p21.1, were found. Several candidate TSGs within these frequently detected LOH regions have been found, including APG7L at 3p25.3, CLASP2 at 3p23, and CACNA2D3 at 3p21.1. This study also showed that SMSG technology is a very useful approach to rapidly define the minimal deleted region and to identify target TSGs in a given cancer.