(A). ELISA results demonstrating reactivity of three primary hybridoma supernatants against zebrafish CYP1A peptide used for immunizations. (B). ELISA results demonstrating that only mAb CRC4 recognizes a conserved peptide sequence common to all vertebrates. (C). ELISA results demonstrating that the conserved CYP1A sequence blocks the binding of mAb CRC4 to ELISA plate-bound peptide used for immunizations. (D). Cross reactivity of mAb CRC4 with induced CYP1A protein in representative vertebrates. MW, molecular weight markers; C (control); T (treated); ZF, zebrafish liver cells; PLHC-1, Poeciliopsis lucida hepatoma cell line; RBT, β-NF induced rainbow trout microsomes; HEPA1C1C7, mouse hepatoma cell line; Rat, liver S9 fractions; frog, XLK-WG cell line; human CYP1A1, recombinant CYP1A1 expressed in HEK cells. Details are found in Methods and Table 2.

Whole-mount immunolocalization of zebrafish CYP1A protein in a representative DMSO vehicle (control) larva (A,B) and 30 µM retene-exposed (C,D) larvae at 48 hpf using mAb CRC2. Each larva was exposed to their respective treatment from 4 hpf through 48 hpf. BA, brachial vasculature; BV, brain vasculature; H, heart; UP, urogenital pore; PVP, posterior vascular plexus; ISV, intersegmental vessel. Details are found in Methods.

Immunohistochemical detection of CYP1A protein expression using mAb CRC4 in liver tissues from representative vertebrates. (A). Control mouse liver; (B). Liver from a PCB-126 treated mouse; (C). Control chicken embryo liver at 18 days post fertilization (DPF); (D). Liver from a crude oil vapor-exposed chicken embryo at 18 DPF; (E). Liver from CYP1A recalcitrant Gulf killifish; (F). Liver from Gulf killifish collected from a crude oil exposed site; (G). Control Rainbow trout liver; (H). Liver from a β-NF-exposed rainbow trout; (I). Control zebrafish liver; (J). Liver from PCB-126 treated zebrafish. (A,B) are 20× magnification, scale bar = 100 µm (C–J) are 40× magnification, scale bar = 50 µm. Details of tissue sources, experimental conditions, and procedures are found in Methods and Table 2.