Isolation and characterization of mitochondria-rich cell types from the gill of freshwater rainbow trout

A magnetic cell separation technique (MACS) was developed for isolating and characterizing peanut lectin agglutinin positive ([PNA.sup.+]) cells from rainbow trout gills. Percoll density separated mitochondria-rich (MR) cells were serially labeled with PNA-FITC and an anti-FITC antibody covalently coupled to a 50-nm iron particle and then applied to a magnetic column. [PNA.sup.+] MR cells were enriched to >95% purity. Transmission electron microscopy analysis of both the [PNA.sup.+] and PNA negative ([PNA.sup.-]) fraction showed that PNA binds to MR chloride cells while the [PNA.sup.-] cell fraction is comprised of MR cells with features characteristic of pavement cells. Western blotting demonstrated that both [PNA.sup.+] and [PNA.sup.-] fractions had high levels of [Na.sup.+]-[K.sup.+]-ATPase and Sco1 expression; however, relative expression of [H.sup.+]-ATPase in [PNA.sup.+] and [PNA.sup.-] cells demonstrated that untreated fish had twofold higher [H.sup.+]-ATPase levels in [PNA.sup.-] cells relative to the [PNA.sup.+] cells. Furthermore, hypercapnic acidosis significantly increased the relative [H.sup.+]-ATPase expression on [PNA.sup.-] cells only, whereas metabolic alkalosis had no significant effect. peanut lectin; pavement cell; fish; hypercapnia; hydrogen-adenosine-triphosphatase