Non-invasive fetal genotyping for maternal alleles with droplet digital PCR: A comparative study of analytical approaches
OBJECTIVES: To develop a flexible droplet digital PCR (ddPCR) workflow to perform non-invasive prenatal diagnosis via relative mutation dosage (RMD) for maternal pathogenic variants with a range of inheritance patterns, and to compare the accuracy of multiple analytical approaches. METHODS: Cell free DNA (cfDNA) was tested from 124 archived maternal plasma samples: 88 cases for sickle cell disease and 36 for rare Mendelian conditions. Three analytical methods were compared: sequential probability ratio testing (SPRT), Bayesian and z-score analyses. RESULTS: The SPRT, Bayesian and z-score analyses performed similarly well with correct prediction rates of 96%, 97% and 98%, respectively. However, there were high rates of inconclusive results for each cohort, particularly for z-score analysis which was 31% overall. Two samples were incorrectly classified by all three analytical methods; a false negative result predicted for a fetus affected with sickle cell disease and a false positive result predicting the presence of an X-linked IDS variant in an unaffected fetus. CONCLUSIONS: ddPCR can be applied to RMD for diverse conditions and inheritance patterns, but all methods carry a small risk of erroneous results. Further evaluation is required both to reduce the rate of inconclusive results and explore discordant results in more detail.
Citation
@article{j.2023,
author = {Shaw, J. and Scotchman, E. and Paternoster, B. and Ramos, M.
and Nesbitt, S. and Sheppard, S. and Snowsill, T. and Chitty, L. S.
and Chandler, N.},
title = {Non-Invasive Fetal Genotyping for Maternal Alleles with
Droplet Digital {PCR:} {A} Comparative Study of Analytical
Approaches},
journal = {Prenatal Diagnosis},
date = {2023-02-09},
url = {https://tristansnowsill.co.uk/non-invasive-fetal-genotyping-for-maternal-alleles.html},
doi = {10.1002/pd.6333},
langid = {en},
abstract = {OBJECTIVES: To develop a flexible droplet digital PCR
(ddPCR) workflow to perform non-invasive prenatal diagnosis via
relative mutation dosage (RMD) for maternal pathogenic variants with
a range of inheritance patterns, and to compare the accuracy of
multiple analytical approaches. METHODS: Cell free DNA (cfDNA) was
tested from 124 archived maternal plasma samples: 88 cases for
sickle cell disease and 36 for rare Mendelian conditions. Three
analytical methods were compared: sequential probability ratio
testing (SPRT), Bayesian and z-score analyses. RESULTS: The SPRT,
Bayesian and z-score analyses performed similarly well with correct
prediction rates of 96\%, 97\% and 98\%, respectively. However,
there were high rates of inconclusive results for each cohort,
particularly for z-score analysis which was 31\% overall. Two
samples were incorrectly classified by all three analytical methods;
a false negative result predicted for a fetus affected with sickle
cell disease and a false positive result predicting the presence of
an X-linked IDS variant in an unaffected fetus. CONCLUSIONS: ddPCR
can be applied to RMD for diverse conditions and inheritance
patterns, but all methods carry a small risk of erroneous results.
Further evaluation is required both to reduce the rate of
inconclusive results and explore discordant results in more detail.}
}