The importance of costimulation on CD4+ T cells has been well documented. However, primary CTLs against many infections including influenza can be generated in the absence of CD4+ T-cell help. The role of costimulation under such helpless circumstances is not fully elucidated. Here, we investigated such a role for CD28 using CTLA4Ig transgenic (Tg) mice. To ensure valid comparison across the genotypes, we showed that all mice had similar naive precursor frequencies and similar peak viral loads. In the absence of help, viral clearance was significantly reduced in CTLA4Ig Tg mice compared with WT mice. CD44+BrdU+influenza-specific CD8+ T cells were diminished in CTLA4Ig Tg mice at days 5 and 8 postinfection. Adoptive transfer of ovalbumin-specific transgenic CD8+ T cells (OT-I)-I cells into WT or CTLA4Ig Tg mice revealed that loss of CD28 costimulation resulted in impairment in OT-I cell division. As shown previously, neither viral clearance nor the generation of influenza-specific CD8+ T cells was affected by the absence of CD4+ T cells alone. In contrast, both were markedly impaired by CD28 blockade of helpless CD8+ T cells. We suggest that direct CD28 costimulation of CD8+ T cells is more critical in their priming during primary influenza infection than previously appreciated.