The germinal center (GC) develops after antigenic stimulation and is thought to occur at the site of various immune responses. We separated a single GC from chicken spleen after antigenic stimulation. Flow cytometric analysis of the cells derived from a single GC and RT-PCR analysis of Ig mRNA expression in GC was performed. Direct evidence indicates that: (1) there was a considerable difference in the cell population of each GC, (2) the ratio of CD3(+) cells in a GC remains constant at 10-20%, (3) the highest proportion of sIgY(+) cells in a GC occurs 1 week after the time of highest proportion of sIgM(+) cells, and (4) RT-PCR analysis was used to detect IgY mRNA expression in a GC. The continuous existence of CD3(+) cells, the alterations in sIgM(+) and sIgY(+) cell ratios, and the expression of IgY mRNA strongly suggest that Ig class switching occurs in the GC during an immune response.