Evaluation of two real-time multiplex PCR screening assays detecting fetal RHD in plasma from RhD negative women to ascertain the requirement for antenatal RhD prophylaxis

Frederik Banch Clausen; Grethe Risum Krog; Klaus Rieneck; Emma Elin Frida Råsmark; Morten Hanefeld Dziegiel

doi:10.1159/000321347

Evaluation of two real-time multiplex PCR screening assays detecting fetal RHD in plasma from RhD negative women to ascertain the requirement for antenatal RhD prophylaxis

Frederik Banch Clausen, Grethe Risum Krog, Klaus Rieneck, Emma Elin Frida Råsmark, Morten Hanefeld Dziegiel

16 Citations (Scopus)

Abstract

Objective: To evaluate two different multiplex real-time PCR assays detecting fetal RHD for screening of RhD negative women in relation to antenatal RhD prophylaxis. Methods: We designed a duplex assay for the detection of RHD exon 7 and 10 and a triplex assay for the detection of RHD exon 7, 10 and 5. We used the same fluorescent dye for the exon 7 and 10 probes to increase sensitivity; exon 5 was VIC labeled. We evaluated possible inhibition of DNA amplification with dilution experiments. We then tested the two multiplex assays with DNA extracted from 97 plasma samples from 38 RhD negative women in gestational weeks 6-37. Results: Dilution experiments revealed no inhibition of amplification in the multiplex assays. For plasma samples, the duplex assay was significantly more sensitive than the triplex assay (p < 0.0001). For the duplex assay (exon 7/10), accuracy was 99.0%. For the triplex assay (exon 7/10), accuracy was 94.2%. Detection of exon 5 was less reliable. Conclusion: The duplex assay using exon 7/10 was the most reliable for prenatal prediction of fetal RhD type as a candidate assay for screening of RhD negative women in relation to antenatal RhD prophylaxis. The triplex assay needs further optimization.

Original language	English
Journal	Fetal Diagnosis and Therapy
Volume	29
Issue number	2
Pages (from-to)	155-63
Number of pages	9
ISSN	1015-3837
DOIs	https://doi.org/10.1159/000321347
Publication status	Published - Mar 2011

Keywords

Female
Fetal Diseases
Humans
Mass Screening
Polymerase Chain Reaction
Pregnancy
Rh Isoimmunization
Rh-Hr Blood-Group System
Sensitivity and Specificity

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Evaluation of two real-time multiplex PCR screening assays detecting fetal RHD in plasma from RhD negative women to ascertain the requirement for antenatal RhD prophylaxis. / Clausen, Frederik Banch; Krog, Grethe Risum; Rieneck, Klaus et al.

In: Fetal Diagnosis and Therapy, Vol. 29, No. 2, 03.2011, p. 155-63.

Research output: Contribution to journal › Journal article › Research › peer-review

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abstract = "Objective: To evaluate two different multiplex real-time PCR assays detecting fetal RHD for screening of RhD negative women in relation to antenatal RhD prophylaxis. Methods: We designed a duplex assay for the detection of RHD exon 7 and 10 and a triplex assay for the detection of RHD exon 7, 10 and 5. We used the same fluorescent dye for the exon 7 and 10 probes to increase sensitivity; exon 5 was VIC labeled. We evaluated possible inhibition of DNA amplification with dilution experiments. We then tested the two multiplex assays with DNA extracted from 97 plasma samples from 38 RhD negative women in gestational weeks 6-37. Results: Dilution experiments revealed no inhibition of amplification in the multiplex assays. For plasma samples, the duplex assay was significantly more sensitive than the triplex assay (p < 0.0001). For the duplex assay (exon 7/10), accuracy was 99.0%. For the triplex assay (exon 7/10), accuracy was 94.2%. Detection of exon 5 was less reliable. Conclusion: The duplex assay using exon 7/10 was the most reliable for prenatal prediction of fetal RhD type as a candidate assay for screening of RhD negative women in relation to antenatal RhD prophylaxis. The triplex assay needs further optimization.",

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Evaluation of two real-time multiplex PCR screening assays detecting fetal RHD in plasma from RhD negative women to ascertain the requirement for antenatal RhD prophylaxis

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Keywords

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