Cell-free DNA-based prenatal screening via rolling circle amplification: Identifying and resolving analytic issues.

OBJECTIVE: A rolling circle amplification (RCA) based commercial methodology using cell-free (cf)DNA to screen for common trisomies became available in 2018. Relevant publications documented high detection but with a higher than expected 1% false positive rate. Preliminary evidence suggested assay variability was an issue. A multi-center collaboration was created to explore this further and examine whether subsequent manufacturer changes were effective. METHODS: Three academic (four devices) and two commercial (two devices) laboratories provided run date, chromosome 21, 18, and 13 run-specific standard deviations, number of samples run, and reagent lot identifications. Temporal trends and between-site/device consistency were explored. Proportions of run standard deviations exceeding pre-specified caps of 0.4%, 0.4% and 0.6% were computed. RESULTS: Overall, 661 RCA runs between April 2019 and July 30, 2022 tested 39,756 samples. In the first 24, subsequent 9, and final 7 months, proportions of capped chromosome 21 runs dropped from 39% to 22% to 6.0%; for chromosome 18, rates were 76%, 36%, and 4.0%. Few chromosome 13 runs were capped using the original 0.60%, but capping at 0.50%, rates were 28%, 16%, and 7.6%. Final rates occurred after reformulated reagents and imaging software modifications were fully implemented across all devices. Revised detection and false positive rates are estimated at 98.4% and 0.3%, respectively. After repeat testing, failure rates may be as low as 0.3%. CONCLUSION: Current RCA-based screening performance estimates are equivalent to those reported for other methods, but with a lower test failure rate after repeat testing.

Non-Invasive prenatal testing with rolling circle amplification: Real-world clinical experience in a non-molecular laboratory.

BACKGROUND: Non-invasive prenatal testing (NIPT) using cell-free DNA (cfDNA) circulating in maternal blood provides a sensitive and specific screening technique for common fetal aneuploidies, but the high cost and workflow complexity of conventional methodologies limit its widespread implementation. A unique rolling circle amplification methodology reduces cost and complexity, providing a promising alternative for increased global accessibility as a first-tier test. METHODS: In this clinical study, 8160 pregnant women were screened on the Vanadis system for trisomies 13, 18, and 21, and positive results were compared to clinical outcomes where available. RESULTS: The Vanadis system yielded a 0.07% no-call rate, a 98% overall sensitivity, and a specificity of over 99% based on available outcomes. CONCLUSION: The Vanadis system provided a sensitive, specific, and cost-effective cfDNA assay for trisomies 13, 18, and 21, with good performance characteristics and low no-call rate, and it eliminated the need for either next-generation sequencing or polymerase chain reaction amplification.

Migraine diagnosis and treatment: A knowledge and needs assessment of women's healthcare providers.

BACKGROUND: Studies suggest that migraine is often underdiagnosed and inadequately treated in the primary care setting, despite many patients relying on their primary care provider (PCP) to manage their migraine. Many women consider their women's healthcare provider to be their PCP, yet very little is known about migraine knowledge and practice patterns in the women's healthcare setting. OBJECTIVE: The objective of this study was to assess women's healthcare providers' knowledge and needs regarding migraine diagnosis and treatment. METHODS: The comprehensive survey assessing migraine knowledge originally developed for PCPs was used in this study, with the addition of a section regarding the use of hormonal medications in patients impacted by migraine. Surveys were distributed online, and primarily descriptive analyses were performed. RESULTS: The online survey was completed by 115 women's healthcare providers (response rate 28.6%; 115/402), who estimated that they serve as PCPs for approximately one-third of their patients. Results suggest that women's healthcare providers generally recognize the prevalence of migraine, but experience some knowledge gaps regarding migraine management. Despite 82.6% (95/115) of survey respondents feeling very comfortable or somewhat comfortable with diagnosing migraine, only 57.9% (66/114) reported routinely asking patients about headaches during annual visits. Very few were familiar with the American Academy of Neurology guidelines on preventative treatment (6.3%; 7/111) and the Choosing Wisely Campaign recommendations on migraine treatment (17.3%; 19/110), and many prescribed medications known to contribute to medication overuse headache. In addition, only 24.3% (28/115) would order imaging for a new type of headache, 35.7% (41/115) for worsening headache, and 47.8% (55/115) for headache with neurologic symptoms; respondents cited greater tendency with sending patients to an emergency department for the same symptoms. Respondents had limited knowledge of evidence-based, non-pharmacological treatments for migraine (i.e., biofeedback or cognitive behavioral therapy), with nearly none placing referrals for these services. Most providers were comfortable prescribing hormonal contraception (mainly progesterone only) to women with migraine without aura (80.9%; 89/110) and with aura (72.5%; 79/109), and followed American College of Obstetricians and Gynecologists (ACOG) guidelines to limit combination hormonal contraception for patients with aura. When queried, 6.3% or less (5/79) of providers would prescribe estrogen-containing contraception for women with migraine with aura. Only 37.3% (41/110) of respondents reported having headache/migraine education. Providers indicated interest in education pertaining to migraine prevention and treatment (96.3%; 105/109), migraine-associated disability (74.3%; 81/109), and diagnostic testing (59.6%; 65/109). CONCLUSION: Women's healthcare providers appear to have several knowledge gaps regarding the management of migraine in their patients. These providers would likely benefit from access to a headache-specific educational curriculum to improve provider performance and patient outcomes.