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. 2024 Sep 30;14(19):2194.
doi: 10.3390/diagnostics14192194.

Evaluation of C-C Motif Chemokine Receptor 5 (CCR5) as a Sample Adequacy Control in HPV Molecular Diagnostics

Ruth C Njoku  1   2 , Marianna Martinelli  2 , Chiara Giubbi  2 , Sofia De Marco  2 , Barbara Torsello  2 , Morena d'Avenia  3 , Manuela Sironi  2 , Cristina Bianchi  2 , Clementina E Cocuzza  2
Affiliations

Evaluation of C-C Motif Chemokine Receptor 5 (CCR5) as a Sample Adequacy Control in HPV Molecular Diagnostics

Ruth C Njoku et al. Diagnostics (Basel). .

Abstract

Background: Reliable Human Papillomavirus (HPV) testing and genotyping are essential for quality assurance in HPV-based primary screening, disease management and for monitoring the impact of HPV vaccination. The clinical validation of HPV molecular diagnostic assays has significantly contributed to these objectives; however, little emphasis has been placed on assuring sample quality. This study aimed to evaluate the accuracy of sample cellularity assessment using the C-C Motif Chemokine Receptor 5 (CCR5) gene target as a marker of sample adequacy in molecular diagnostics. Methods: Jurkat cell line samples were counted using both a Thoma cell-counting chamber and Fluorescence-Activated Cell Sorting (FACS). Jurkat cell line samples at three different concentrations were subsequently evaluated using the OncoPredict HPV Quality Control (QC) real-time PCR assay, employing CCR5 for molecular cellularity quantification. Results: The cellularity values obtained were comparable across the three different methods for all dilutions of the cell line tested. Conclusions: The results obtained from this study show that CCR5 represents a promising molecular marker for the accurate quantification of sample cellularity, confirming its use as a reliable sample adequacy control, thus reducing the risk of "false-negative" results.

Keywords: C-C Motif Chemokine Receptor 5 (CCR5); Human Papillomavirus (HPV); OncoPredict HPV Quality Control (QC) assay; sample adequacy control (SAC).

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Conflict of interest statement

CEC received research funding and consumables to support research from the following commercial entities in the last 3 years: BD, Seegene, Copan, Novosanis and Hiantis; CEC is a minority shareholder of Hiantis. SRL., R.C.N., M.M., C.G., S.D.M., B.T., M.D.A., M.S. and C.B have no conflicts of interest to declare.

Figures

Figure 1
Figure 1
Study workflow.
Figure 2
Figure 2
Gating strategy of Flow Cytometry: The gating strategy of Flow Cytometry for the sorting of Jurkat cells was: (i) “SINGLE” cells to exclude aggregates (ii) “CELLS” to sort the desired cell number. SSC: Side Scatter. FSC: Forward Scatter.
Figure 3
Figure 3
CCR5 standard curve plot.
Figure 4
Figure 4
Graphic representation of the QC CCR5 cellularity value. Cellularity values (Log10 CCR5 cellularity, cells/sample) are presented for three replicates (a, b, c) across initial cell numbers (103, 104 and 105).
See this image and copyright information in PMC

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