To test his system, Nirenberg used polyU as the mRNA, and [14C]phenylalanine-charged tRNA, which binds to ribosomes in the presence of polyU. Ribosomes were incubated with both polyU and [14C]-phenylalanine tRNA for sufficient time to allow both molecules to bind to the ribosomes; the reaction mixtures were then passed through a nylon membrane. When the membranes were analyzed using a scintillation counter, they contained radioactivity, demonstrating that in this system polyU could recognize phenylalanine-charged tRNA. But was this recognition specific for the proper amino-acylated tRNA? As a control, [14C]-lysine- and [14C]-proline-charged tRNAs also were incubated with polyU and ribosomes. After the reaction mixtures were passed through a nylon filter, no radioactivity was detected on the filter.