FIG. 5.

Regulation of CrRelish expression by secondary bacterial challenge. (A) Western blot analysis of CrRelish protein expression after first bacterial infection and secondary bacterial infection. Twelve hours after the first infection, the horseshoe crabs were rechallenged with the same dose of P. aeruginosa used in the primary infection and the hemocyte lysates were collected at the indicated time points up to 12 hpi after both the primary and secondary infections. Horseshoe crab actin served as a loading control. (B) RT-PCR for examination of CrRelish mRNA expression at time points after primary and secondary infections was as indicated for the Western blot. Horseshoe crab actin mRNA was the loading control. (C) Quantitation of Western blot analysis and RT-PCR bands by densitometric scanning. The relative intensities for primary and secondary infections were presented separately.