Figure 4

Double-probe Fluorescent in situ Hybridization Using Digoxigenin-labeled ATPase 6 and Fluorescein-labeled SP-B Riboprobes. Three micrometer-thick sections of normal rat lung (NRL) (A and G) and Pc-infected rat lung (PcIRL) (B and H) were processed for double-probe fluorescent in situ hybridization. Panels A (NRL) and B (PcIRL) are composite images of sections reacted with a mixture of anti-sense digoxigenin-labeled ATPase 6 and fluorescein-labeled SP-B probes. Panels C and E are split images of the composite A, and panels D and F are split images of the composite B, showing concurrence of red (ATPase 6) and green (SP-B) signal in the same cells. Panels G (NRL) and H (PcIRL) are composite images of sections reacted with both ATPase 6 and SP-B sense probes showing very little background signal. Arrows in panel B indicate the punctate DAPI-stained (blue) nuclei of P. carinii organisms.