Assessing sample adequacy and clinical performance of self-collected and clinician-collected HPV specimens using internal control Ct values.

BACKGROUND

Human papillomavirus (HPV) testing is the primary method for cervical cancer screening, but reliable detection depends on adequate sample cellularity. Cycle threshold (Ct) values for the assay's internal control (IC), such as β-globin, are commonly used as proxies for adequacy, yet standardized Ct cut-offs are lacking. We aimed to contribute evidence-based thresholds for sample adequacy using real-world data.

METHODS

We analyzed 237,853 clinician-collected and self-collected samples tested with the BD Onclarity™ HPV Assay between 2022 and 2024. β-globin Ct values were assessed by HPV status to evaluate adequacy. Histologically confirmed CIN2+ outcomes were linked via the National Cervical Screening Registry to assess clinical performance.

RESULTS

Among 110,482 clinician-taken samples, 73.63 % (81,350) were HPV negative; 74.32 % (60,457) of these had β-globin Ct ≤28, and only 1.28 % exceeded Ct 32.1. In 127,390 self-collected samples, 83.47 % (106,329) were HPV negative; 99.66 % (105,967) had Ct ≤28 and only 0.06 % exceeded Ct 32.1. HPV positivity declined gradually beyond Ct 26 and more markedly above Ct 28. CIN2+ cases (n = 5546) were rarely HPV negative (n = 73), and these showed low β-globin Ct values, indicating adequate cellularity. Self-collected samples had significantly lower Ct values than clinician-taken ones (median 21.5 vs. 26.5; p < 2.2e-16), likely due to lower resuspension volume.

CONCLUSIONS

Both clinician- and self-collected samples showed adequate cellularity, with potentially false negative HPV results from low cellular content appearing rare. Observed patterns suggest Ct <26 as optimal and Ct <28 as a minimum for program-level quality assurance with the BD Onclarity™ HPV Assay.