Are estradiol assays accurate enough for clinical testing?

Accurate estradiol measurements are important at both low and high concentrations, e.g., for diagnosis of estrogen-producing ovarian and testicular tumors and monitoring anti-estrogen therapy for estrogen receptor-positive breast cancer. Based on CAP survey data, estradiol is measured using immunoassays in > 99% of US clinical laboratories, and laboratory developed tests (LDT) using tandem mass spectrometry in < 1% (1). Estradiol was recently added to the list of analytes specifically required by the CLIA’88 regulations for proficiency testing (PT) (2). PT events are conventionally done using modified human serum sample pools, or similar matrices, for which the commutability is unknown. Therefore, participants’ results are peer-grouped, and the mean values are used as targets. Conventional PT, therefore, can only assess whether a laboratory’s analysis meet acceptance limits relative to its peers using the same method, and it cannot differentiate matrix bias from calibration bias or other causes of inaccuracy. In contrast, accuracy-based PT uses unaltered human samples and target values determined by a reference method measurement procedure and can reliably assess proficiency in the context of clinical needs.

We conducted an accuracy-based PT event via the New York State Department of Health (NYSDOH) proficiency testing program in 2016 using 5 authentic serum samples from single-donors. They were distributed frozen overnight to 76 NYSDOH-certified laboratories, 2 of 76 used LC-MS/MS methods. Participants analyzed samples for total estradiol and reported results within two weeks post receipt. Accuracy performance as assessed using the CDC-defined targets against: [1] the reported results from participant laboratories, and [2] the method/instrument group means (n ≥ 4) of 9 FDA-approved assays. We observed biases (range) of 34% (-17% to 175%), 40% (-33% to 386%), 16% (-45% to 193%), 5% (-27% to 117%), and -4% (-31% to 21%), for samples at estradiol of 24.1, 28.4, 61.7, 94.1 and 127 pg/mL, respectively, revealing greater biases at low estradiol concentrations than high concentrations. The highest value was about 7-times higher than the lowest value for the sample at estradiol 28.4 pg/mL. Two of the 76 participants used LDT LC-MS/MS methods and had results with about a two-fold difference, i.e., 19 vs 39 pg/mL, and biases of -33.1% and 37%, respectively, for the same. These data support that LDTs for estradiol tests can be inaccurate (3).

We estimated the participant laboratories’ PT performance using the CDC-defined targets and each of the 3 acceptance criteria: i.e., target ±25% or ±15 pg/mL (whichever was greater, NYSDOH), ±30% (CLIA), or ±26% (minimal limit based on biological variability). Results showed that 59%, 69% and 87% of laboratories, respectively, would receive a PT event passing score. However, when the PT evaluation was done using peer method/system means as targets, > 95% laboratories would obtain PT passing score regardless which criterion was used.

We concluded that some immunoassays and LC-MS/MS-based methods did not accurately measure estradiol at low concentrations, possibly impacting the quality of patient care. Future efforts would be best focused on lower concentrations. Accuracy-based PT identified IVD products that need improvement in the accuracy performance.

References

1. Sex Hormones Y-B 2023 Participant Summary. Survey and Anatomic Pathology Education Programs. College of American Pathologists.
2. Clinical Laboratory Improvement Amendments of 1988 (CLIA) Proficiency Testing Regulations Related to Analytes and Acceptable Performance. Fed Reg 2022;87:41194-242. https://www.govinfo.gov/content/pkg/FR-2022-07-11/pdf/2022-14513.pdf(accessed Dec 2024)
3. Cao TZ, Rej R, Vesper H, Rex Astles J. Accuracy-based proficiency testing for estradiol measurements. Clin Biochem. 2023 Dec 1:110700. doi: 10.1016/j.clinbiochem.2023.110700. Epub ahead of print. PMID: 38043696.

Disclaimer

The findings and conclusions in this manuscript are those of the authors and do not necessarily represent the official views or positions of the Centers for Disease Control and Prevention/Agency for Toxic Substances and Disease Registry and Wadsworth Center of New York State Department of Health. Use of trade names and commercial sources is for identification only and does not constitute endorsement by the U.S. Department of Health and Human Services, the U.S. Centers for Disease Control and Prevention, or New York State Department of Health.