NlaIII restriction analysis of the 94-bp FcγRIIIA-specific fragment, containing the polymorphic nucleotide 559. A 1.2-kb FcγRIIIA-specific fragment was amplified from genomic DNA, followed by a nested PCR. The sense primer of this nested PCR contained a mismatch that introduced a NlaIII restriction site only in the FcγRIIIA-158V (559G) allele. Homozygous FcγRIIIA-158FF fragments were not digested (lanes 1 through 3). Three bands (94 bp, 61 bp, and 33 bp) were visible in heterozygous individuals, whereas homozygous FcγRIIIA-158VV fragments were maximally digested (lanes 7 through 9). A 92-bp fragment of low intensity remained in homozygous 559G fragments (lanes 7 through 9).