Membranous HLA class I expression analysis by four-color multiparameter flow cytometry of colorectal tumor cell suspensions. Fluorescence intensities of HLA class I with antibodies W6/32 (HC), BBM.1 (B2M), and alloantigen-specific antibodies against single A and B alleles (A.1 – B.2; see Table 2) are displayed in filled curves; corresponding negative controls are in non-filled curves. A, keratin positive (ker+) epithelial cells are distinguished from vimentin positive (vim+) stroma cells and infiltrating leukocytes. DNA ploidy analysis of case 122 reveals an aneuploid ker+ tumor cell fraction (black) compared to the normal diploid vim+ cells (light gray). B, Expression of all HLA molecules tested in case 122 of both ker+, tumor (black), and vim+ 'normal' (gray) cells. C, loss of the single A.1 (HLA-A2) allele was observed for tumor cells in case 191. D, loss of expression of all 4 HLA-A and -B alleles in tumor cells, but retention of HC and B2M, in case 179. The patient is homozygous for HLA-A3 (--;see Table 2).