Detection of endogenous avian leukosis virus envelope in chicken plasma using R2 antiserum

Immunologic tolerance to oncogenic avian leukosis virus (ALV) is mediated, in part, by the interaction of endogenous ALV (EV) envelope and immune competent cells. A flow cytometry method is described for detecting the EV envelope in chicken plasma or serum. The method employs two types of target red blood cells (RBC) obtained from chickens lacking EV; RBC susceptible to EV infection (containing EV receptors), and those resistant to EV infection (lacking EV receptors). RBC from susceptible chickens will bind EV envelope glycoprotein (gp85) when present in plasma. The gp85-bound RBC are subsequently incubated with a highly specific chicken alloantibody, termed R2. Using flow cytometry, gp85 is detected indirectly with a fluoresceine-tagged antibody to chicken immunoglobulin; plasmas lacking gp85 are nonreactive and fluorescence remains at a background level. Because RBC from resistant chickens are nonreactive regardless of the presence or absence of EV gp85, a specific binding index was calculated to compare relative binding of EV gp85 on susceptible and resistant RBC, and thus identify chickens that express EV gp85. The specificity of the assay was demonstrated using plasma from chickens of 14 standard laboratory lines previously defined for EV envelope expression including two sets of highly congenic lines that differ in EV expression. This assay detects differences attributable to EV gp85 in chickens of commercial breeding lines of White Leghorns and broilers. Moreover, if chickens lack EV, the R2 plasma assay can differentiate between EV-susceptible and EVresistant siblings.