Automated Erythropoietin Testing on Beckman Coulter’s Family of Access® Immunoassay Systems.

Background: Erythropoietin (EPO) is a ~30,400 dalton glycoprotein hormone produced primarily by the kidneys. EPO is the principal factor stimulating bone marrow cells to differentiate into red blood cells (RBCs). Normally, EPO levels increase when oxygen or RBC levels become abnormally low, or decrease if RBC concentrations become abnormally high. The evaluation of EPO is useful in distinguishing between disease etiologies in erythrocytosis or polycythemia (overproduction of RBCs). Primary polycythemia or polycythemia vera is caused by EPO-independent growth of erythrocytic progenitors from neoplastic bone marrow stem cells, and in most cases decreased levels of EPO are found in the serum of affected patients. Various types of secondary polycythemias are associated with the production of elevated levels of EPO. EPO evaluation has been utilized recently to aid in predicting the responsiveness of anemic cancer patients to recombinant human EPO (rhEPO) therapy. An elevated EPO level (>200 mIU/mL) in an anemic patient may predict a low response to rhEPO therapy.

Objectives and Methods: This study was designed to determine the performance characteristics of the EPO assay on Beckman Coulter’s family of Access Immunoassay Systems. This automated sandwich assay combines a polyclonal anti-EPO alkaline phosphatase conjugate and 75 microliters of sample (serum or plasma). Following a 10-minute incubation, paramagnetic particles coated with goat anti-mouse IgG / mouse anti-rhEPO monoclonal antibody are added. A second 20-minute incubation allows for antigen-antibody complex formation and is followed by wash and substrate addition steps. Light generated by the reaction is measured with a luminometer. The light production is directly proportional to the concentration of EPO in the sample. The EPO concentration is determined from a stored multipoint calibration curve. The time to first result is < 45 minutes.

Results: The dynamic range of the EPO assay is 0.6–750 mIU/mL with an analytical sensitivity of less than or equal to 0.6 mIU/mL. When evaluated using control material in a total of 20 tests with 2 replicates per test, the assay demonstrated a total imprecision of less than or equal to 10% at concentrations from approximately 9.5–475 mIU/mL. Method comparison to a commercially available EPO microtiter assay with 103 samples (range approximately 3–200 mIU/mL) yielded a correlation coefficient of 0.988 and a slope of 1.051. The expected range of 2.59–18.50 mIU/mL (range 1.48–31.88 mIU/mL) was based on a 95% non-parametric analysis of 122 normal samples. When tested using related compounds and common serum and plasma components, no significant cross reactivity or interference was observed.

Conclusion: With the expanding uses for EPO testing, including the evaluation of cancer patients who may be candidates for rhEPO therapy, the rapid turnaround time provided by this automated assay for EPO on the Access Immunoassay Systems provides the laboratory with a rapid, sensitive and precise assay for measuring EPO.

Pending submission to and clearance by the United States FDA; not yet available for diagnostic use.