Comparison of diagnostic performances of HDV-RNA quantification assays used in clinical practice: Results from a national quality control multicenter study

A reliable quantification of hepatitis D virus (HDV) RNA is of paramount importance for monitoring patients under antiviral therapy. This quality control study compares the diagnostic performances of quantitative HDV-RNA assays used in clinical practice. Two HDV-RNA sample panels were quantified in...

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Published in:Journal of clinical virology Vol. 180; p. 105850
Main Authors: Salpini, Romina, Piermatteo, Lorenzo, Caviglia, Gian Paolo, Bertoli, Ada, Brunetto, Maurizia Rossana, Bruzzone, Bianca, Callegaro, Annapaola, Caudai, Cinzia, Cavallone, Daniela, Chessa, Luchino, Coghe, Fernando, Coppola, Nicola, Cuomo, Nunzia, D'Anna, Stefano, Di Stefano, Mariantonietta, Facchetti, Floriana, Farina, Claudio, Ferraro, Donatella, Franchin, Elisa, Francisci, Daniela, Galli, Silvia, Garbuglia, Anna Rosa, Gennari, William, Ghisetti, Valeria, Lampertico, Pietro, Lo Caputo, Sergio, Marascio, Nadia, Menzo, Stefano, Micheli, Valeria, Niro, Grazia Anna, Olivero, Antonella, Paba, Pierpaolo, Palermo, Concetta Ilenia, Palmieri, Orazio, Paolucci, Stefania, Pisaturo, Mariantonietta, Pollicino, Teresa, Raffa, Giuseppina, Santantonio, Teresa, Torre, Giulia, Turriziani, Ombretta, Uzzau, Sergio, Uceda Renteria, Sara Colonia, Vatteroni, Marialinda, Zazzi, Maurizio, Craxì, Antonio, Ceccherini-Silberstein, Francesca, Svicher, Valentina, Arosio, Marco, Bastianelli, Sabrina, Gentile, Annamaria, Giardina, Federica A.M., Gidari, Anna, Govoni, Rosalba, Ibba, Gabriele, Loglio, Alessandro, Lombardi, Alessandra, Mascarella, Chiara, Maggi, Fabrizio, Matera, Giovanni, Mazzei, Chiara, Milia, Maria Grazia, Quirino, Angela, Raddi, Adriana, Scioscia, Rosetta, Tagliazucchi, Sara, Totaro, Michele, Valaperta, Rea
Format: Journal Article
Language:English
Published: Netherlands Elsevier B.V 01.10.2025
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ISSN:1386-6532, 1873-5967, 1873-5967
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Abstract A reliable quantification of hepatitis D virus (HDV) RNA is of paramount importance for monitoring patients under antiviral therapy. This quality control study compares the diagnostic performances of quantitative HDV-RNA assays used in clinical practice. Two HDV-RNA sample panels were quantified in 30 centers by RoboGene (N = 9 laboratories), EurobioPlex (N = 7), RealStar (N = 4), AltoStar (N = 1), Bosphore (N = 3), Bosphore-on-InGenius (N = 1), Dia.Pro (N = 2), Nuclear-Laser-Medicine (N = 1) and 3 in-house assays. Panel A and B comprised 8 serial dilutions of WHO/HDV standard (range: 0.5–5.0 log10 IU/ml) and 20 clinical samples (range: 0.5–6.0 log10 IU/ml), respectively. The following parameters were determined: sensitivity by 95 % LOD (limit of detection), precision by intra- and inter-run CV (coefficient of variation), accuracy by the differences between expected-observed HDV-RNA, linearity by linear regression analysis. 95 % LOD varied across assays and centers underlining heterogeneous sensitivities: AltoStar had the lowest 95 % LOD (3 IU/ml) followed by RealStar (10 [min–max: 3–316] IU/ml), Bosphore-on-InGenius (10 IU/ml), RoboGene (31 [3–316] IU/ml), Nuclear-Laser-Medicine (31 IU/ml) and EuroBioplex (100 [100–316] IU/ml). Moreover, 6 assays (RoboGene, EurobioPlex, RealStar, AltoStar, Nuclear-Laser-Medicine and In-house) showed <0.5 log10 IU/ml differences between expected and observed HDV-RNA for all dilutions while other assays had >1 log10 IU/ml underestimations. RealStar, Bosphore-on-InGenius and EurobioPlex had the highest precision (mean intra-run CV < 20 %). Inter-run CV was higher for all assays, with CVs < 25 % for RealStar, AltoStar, Nuclear-Laser-Medicine and EurobioPlex. Seven assays (RoboGene/AltoStar/RealStar/EurobioPlex/Nuclear-Laser-Medicine/In-house) showed a good linearity (R2 > 0.90), but for HDV-RNA < 1000 IU/ml only Bosphore-on-InGenius, AltoStar, RealStar and Robogene showed a R2 > 0.85. This study underlines heterogeneous sensitivities (inter- and intraassays), that could hamper proper HDV-RNA quantification, particularly at low viral loads. This raises the need to improve the diagnostic performance of most assays for properly identifying virological response to anti-HDV drugs. ●The assays for HDV-RNA quantification need to optimize the performances at low HDV-RNA concentrations.●The laboratories should mitigate inter-laboratory and inter-assay variability by standardizing procedures and by favouring automation.●The use of sensitive assays is important for the proper monitoring of virological response to anti-HDV drugs and for optimizing therapeutical approaches aimed at setting-up a finite course of anti-HDV treatment.
AbstractList A reliable quantification of hepatitis D virus (HDV) RNA is of paramount importance for monitoring patients under antiviral therapy. This quality control study compares the diagnostic performances of quantitative HDV-RNA assays used in clinical practice. Two HDV-RNA sample panels were quantified in 30 centers by RoboGene (N = 9 laboratories), EurobioPlex (N = 7), RealStar (N = 4), AltoStar (N = 1), Bosphore (N = 3), Bosphore-on-InGenius (N = 1), Dia.Pro (N = 2), Nuclear-Laser-Medicine (N = 1) and 3 in-house assays. Panel A and B comprised 8 serial dilutions of WHO/HDV standard (range: 0.5–5.0 log10 IU/ml) and 20 clinical samples (range: 0.5–6.0 log10 IU/ml), respectively. The following parameters were determined: sensitivity by 95 % LOD (limit of detection), precision by intra- and inter-run CV (coefficient of variation), accuracy by the differences between expected-observed HDV-RNA, linearity by linear regression analysis. 95 % LOD varied across assays and centers underlining heterogeneous sensitivities: AltoStar had the lowest 95 % LOD (3 IU/ml) followed by RealStar (10 [min–max: 3–316] IU/ml), Bosphore-on-InGenius (10 IU/ml), RoboGene (31 [3–316] IU/ml), Nuclear-Laser-Medicine (31 IU/ml) and EuroBioplex (100 [100–316] IU/ml). Moreover, 6 assays (RoboGene, EurobioPlex, RealStar, AltoStar, Nuclear-Laser-Medicine and In-house) showed <0.5 log10 IU/ml differences between expected and observed HDV-RNA for all dilutions while other assays had >1 log10 IU/ml underestimations. RealStar, Bosphore-on-InGenius and EurobioPlex had the highest precision (mean intra-run CV < 20 %). Inter-run CV was higher for all assays, with CVs < 25 % for RealStar, AltoStar, Nuclear-Laser-Medicine and EurobioPlex. Seven assays (RoboGene/AltoStar/RealStar/EurobioPlex/Nuclear-Laser-Medicine/In-house) showed a good linearity (R2 > 0.90), but for HDV-RNA < 1000 IU/ml only Bosphore-on-InGenius, AltoStar, RealStar and Robogene showed a R2 > 0.85. This study underlines heterogeneous sensitivities (inter- and intraassays), that could hamper proper HDV-RNA quantification, particularly at low viral loads. This raises the need to improve the diagnostic performance of most assays for properly identifying virological response to anti-HDV drugs. ●The assays for HDV-RNA quantification need to optimize the performances at low HDV-RNA concentrations.●The laboratories should mitigate inter-laboratory and inter-assay variability by standardizing procedures and by favouring automation.●The use of sensitive assays is important for the proper monitoring of virological response to anti-HDV drugs and for optimizing therapeutical approaches aimed at setting-up a finite course of anti-HDV treatment.
A reliable quantification of hepatitis D virus (HDV) RNA is of paramount importance for monitoring patients under antiviral therapy. This quality control study compares the diagnostic performances of quantitative HDV-RNA assays used in clinical practice. Two HDV-RNA sample panels were quantified in 30 centers by RoboGene (N = 9 laboratories), EurobioPlex (N = 7), RealStar (N = 4), AltoStar (N = 1), Bosphore (N = 3), Bosphore-on-InGenius (N = 1), Dia.Pro (N = 2), Nuclear-Laser-Medicine (N = 1) and 3 in-house assays. Panel A and B comprised 8 serial dilutions of WHO/HDV standard (range: 0.5-5.0 log10 IU/ml) and 20 clinical samples (range: 0.5-6.0 log10 IU/ml), respectively. The following parameters were determined: sensitivity by 95 % LOD (limit of detection), precision by intra- and inter-run CV (coefficient of variation), accuracy by the differences between expected-observed HDV-RNA, linearity by linear regression analysis. 95 % LOD varied across assays and centers underlining heterogeneous sensitivities: AltoStar had the lowest 95 % LOD (3 IU/ml) followed by RealStar (10 [min-max: 3-316] IU/ml), Bosphore-on-InGenius (10 IU/ml), RoboGene (31 [3-316] IU/ml), Nuclear-Laser-Medicine (31 IU/ml) and EuroBioplex (100 [100-316] IU/ml). Moreover, 6 assays (RoboGene, EurobioPlex, RealStar, AltoStar, Nuclear-Laser-Medicine and In-house) showed <0.5 log10 IU/ml differences between expected and observed HDV-RNA for all dilutions while other assays had >1 log10 IU/ml underestimations. RealStar, Bosphore-on-InGenius and EurobioPlex had the highest precision (mean intra-run CV < 20 %). Inter-run CV was higher for all assays, with CVs < 25 % for RealStar, AltoStar, Nuclear-Laser-Medicine and EurobioPlex. Seven assays (RoboGene/AltoStar/RealStar/EurobioPlex/Nuclear-Laser-Medicine/In-house) showed a good linearity (R > 0.90), but for HDV-RNA < 1000 IU/ml only Bosphore-on-InGenius, AltoStar, RealStar and Robogene showed a R > 0.85. This study underlines heterogeneous sensitivities (inter- and intraassays), that could hamper proper HDV-RNA quantification, particularly at low viral loads. This raises the need to improve the diagnostic performance of most assays for properly identifying virological response to anti-HDV drugs.
A reliable quantification of hepatitis D virus (HDV) RNA is of paramount importance for monitoring patients under antiviral therapy. This quality control study compares the diagnostic performances of quantitative HDV-RNA assays used in clinical practice.INTRODUCTIONA reliable quantification of hepatitis D virus (HDV) RNA is of paramount importance for monitoring patients under antiviral therapy. This quality control study compares the diagnostic performances of quantitative HDV-RNA assays used in clinical practice.Two HDV-RNA sample panels were quantified in 30 centers by RoboGene (N = 9 laboratories), EurobioPlex (N = 7), RealStar (N = 4), AltoStar (N = 1), Bosphore (N = 3), Bosphore-on-InGenius (N = 1), Dia.Pro (N = 2), Nuclear-Laser-Medicine (N = 1) and 3 in-house assays. Panel A and B comprised 8 serial dilutions of WHO/HDV standard (range: 0.5-5.0 log10 IU/ml) and 20 clinical samples (range: 0.5-6.0 log10 IU/ml), respectively. The following parameters were determined: sensitivity by 95 % LOD (limit of detection), precision by intra- and inter-run CV (coefficient of variation), accuracy by the differences between expected-observed HDV-RNA, linearity by linear regression analysis.METHODSTwo HDV-RNA sample panels were quantified in 30 centers by RoboGene (N = 9 laboratories), EurobioPlex (N = 7), RealStar (N = 4), AltoStar (N = 1), Bosphore (N = 3), Bosphore-on-InGenius (N = 1), Dia.Pro (N = 2), Nuclear-Laser-Medicine (N = 1) and 3 in-house assays. Panel A and B comprised 8 serial dilutions of WHO/HDV standard (range: 0.5-5.0 log10 IU/ml) and 20 clinical samples (range: 0.5-6.0 log10 IU/ml), respectively. The following parameters were determined: sensitivity by 95 % LOD (limit of detection), precision by intra- and inter-run CV (coefficient of variation), accuracy by the differences between expected-observed HDV-RNA, linearity by linear regression analysis.95 % LOD varied across assays and centers underlining heterogeneous sensitivities: AltoStar had the lowest 95 % LOD (3 IU/ml) followed by RealStar (10 [min-max: 3-316] IU/ml), Bosphore-on-InGenius (10 IU/ml), RoboGene (31 [3-316] IU/ml), Nuclear-Laser-Medicine (31 IU/ml) and EuroBioplex (100 [100-316] IU/ml). Moreover, 6 assays (RoboGene, EurobioPlex, RealStar, AltoStar, Nuclear-Laser-Medicine and In-house) showed <0.5 log10 IU/ml differences between expected and observed HDV-RNA for all dilutions while other assays had >1 log10 IU/ml underestimations. RealStar, Bosphore-on-InGenius and EurobioPlex had the highest precision (mean intra-run CV < 20 %). Inter-run CV was higher for all assays, with CVs < 25 % for RealStar, AltoStar, Nuclear-Laser-Medicine and EurobioPlex. Seven assays (RoboGene/AltoStar/RealStar/EurobioPlex/Nuclear-Laser-Medicine/In-house) showed a good linearity (R2 > 0.90), but for HDV-RNA < 1000 IU/ml only Bosphore-on-InGenius, AltoStar, RealStar and Robogene showed a R2 > 0.85.RESULTS95 % LOD varied across assays and centers underlining heterogeneous sensitivities: AltoStar had the lowest 95 % LOD (3 IU/ml) followed by RealStar (10 [min-max: 3-316] IU/ml), Bosphore-on-InGenius (10 IU/ml), RoboGene (31 [3-316] IU/ml), Nuclear-Laser-Medicine (31 IU/ml) and EuroBioplex (100 [100-316] IU/ml). Moreover, 6 assays (RoboGene, EurobioPlex, RealStar, AltoStar, Nuclear-Laser-Medicine and In-house) showed <0.5 log10 IU/ml differences between expected and observed HDV-RNA for all dilutions while other assays had >1 log10 IU/ml underestimations. RealStar, Bosphore-on-InGenius and EurobioPlex had the highest precision (mean intra-run CV < 20 %). Inter-run CV was higher for all assays, with CVs < 25 % for RealStar, AltoStar, Nuclear-Laser-Medicine and EurobioPlex. Seven assays (RoboGene/AltoStar/RealStar/EurobioPlex/Nuclear-Laser-Medicine/In-house) showed a good linearity (R2 > 0.90), but for HDV-RNA < 1000 IU/ml only Bosphore-on-InGenius, AltoStar, RealStar and Robogene showed a R2 > 0.85.This study underlines heterogeneous sensitivities (inter- and intraassays), that could hamper proper HDV-RNA quantification, particularly at low viral loads. This raises the need to improve the diagnostic performance of most assays for properly identifying virological response to anti-HDV drugs.CONCLUSIONSThis study underlines heterogeneous sensitivities (inter- and intraassays), that could hamper proper HDV-RNA quantification, particularly at low viral loads. This raises the need to improve the diagnostic performance of most assays for properly identifying virological response to anti-HDV drugs.
ArticleNumber 105850
Author Ceccherini-Silberstein, Francesca
Salpini, Romina
Piermatteo, Lorenzo
Caviglia, Gian Paolo
Lo Caputo, Sergio
Totaro, Michele
Matera, Giovanni
Brunetto, Maurizia Rossana
Olivero, Antonella
Coppola, Nicola
Zazzi, Maurizio
Bastianelli, Sabrina
D'Anna, Stefano
Scioscia, Rosetta
Lampertico, Pietro
Cavallone, Daniela
Gentile, Annamaria
Valaperta, Rea
Paolucci, Stefania
Menzo, Stefano
Palmieri, Orazio
Ibba, Gabriele
Lombardi, Alessandra
Franchin, Elisa
Niro, Grazia Anna
Mazzei, Chiara
Francisci, Daniela
Chessa, Luchino
Milia, Maria Grazia
Micheli, Valeria
Raffa, Giuseppina
Callegaro, Annapaola
Arosio, Marco
Santantonio, Teresa
Giardina, Federica A.M.
Ferraro, Donatella
Loglio, Alessandro
Vatteroni, Marialinda
Caudai, Cinzia
Torre, Giulia
Maggi, Fabrizio
Quirino, Angela
Uceda Renteria, Sara Colonia
Gennari, William
Di Stefano, Mariantonietta
Farina, Claudio
Coghe, Fernando
Pollicino, Teresa
Paba, Pierpaolo
Palermo, Concetta Ilenia
Turriziani, Ombretta
Govoni, Rosalba
Mascarella, Chiara
Bruzzone, Bianca
Uzzau, Sergio
Bertoli, Ada
Raddi, Adriana
Cuomo
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Cites_doi 10.1016/j.jhep.2023.05.001
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Keywords HDV-RNA
Hepatitis D
Real-Time PCR
Language English
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Copyright © 2025 The Authors. Published by Elsevier B.V. All rights reserved.
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Snippet A reliable quantification of hepatitis D virus (HDV) RNA is of paramount importance for monitoring patients under antiviral therapy. This quality control study...
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SubjectTerms HDV-RNA
Hepatitis D
Hepatitis D - diagnosis
Hepatitis D - virology
Hepatitis Delta Virus - genetics
Hepatitis Delta Virus - isolation & purification
Humans
Molecular Diagnostic Techniques - methods
Molecular Diagnostic Techniques - standards
Quality Control
Real-Time PCR
Reproducibility of Results
RNA, Viral - blood
RNA, Viral - genetics
Sensitivity and Specificity
Viral Load - methods
Viral Load - standards
Title Comparison of diagnostic performances of HDV-RNA quantification assays used in clinical practice: Results from a national quality control multicenter study
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https://dx.doi.org/10.1016/j.jcv.2025.105850
https://www.ncbi.nlm.nih.gov/pubmed/40784197
https://www.proquest.com/docview/3238429451
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