Characteristics of a Specific Radioimmunoassay for Measurement of Ferritin on the Surface of Peripheral Mononuclear White Blood Cells in Cancer Patients

Using ¹²⁵I-labeled rabbit anti-Hodgkin's spleen ferritin antibody (RHF), a simple radioimmunoassay has been developed for quantitation of ferritin on the surface of peripheral blood mononuclear white blood cells (PBM). This method makes use of a % specific binding determination (%SP) by measuring the amount of ¹²⁵I-labeled RHF bound to 1 × 10⁶ PBM in the presence and absence of soluble ferritin. To standardize this procedure, artificial ferritin positive control cells were prepared by covalently coupling ferritin to cultured acute lymphoblastic luekemia cells. These cells were tested on a daily basis in parallel with patient PBM's to ensure inter and intra-assay precision and remained stable for over two years. Characteristics of ¹²⁵I-labeled RHF binding to control and patient PBM's were evaluated to determine the specificity of interaction and optimum binding parameters. %SP was linear in the range of 1 × 10⁵ - 1 × 10⁶ PBM's and was progressively inhibited by graded concentrations of soluble ferritin. F(ab')₂ preparations of RHF were equally as effective as intact RHF in blocking ¹²⁵I-labeled RHF binding confirming that ¹²⁵I-labeled RHF was not binding non-specifically to PBM Fc receptors. Additional experiments describing kinetics and methods of standardization of new lots of ¹²⁵I-labeled RHF are also described.