Strong Correlation between Liver and Serum Levels of Hepatitis C
Virus Core Antigen and RNA in Chronically Infected Patients
V. Descamps,a,bA. Op de Beeck,cC. Plassart,a,bE. Brochot,a,bC. François,a,bF. Helle,bM. Adler,dN. Bourgeois,dD. Degré,d,e
G. Duverlie,a,band S. Castelaina,b
Virology Department, Amiens University Hospital, South Hospital, Amiens, Francea; Jules Verne University of Picardy, Amiens, Franceb; Laboratory of Molecular Virology,
Medicine Faculty, Université libre de Bruxelles, Brussels, Belgiumc; Department of Gastroenterology, Hepatopancreatology and Digestive Oncology, Erasme Hospital,
Université libre de Bruxelles, Brussels, Belgiumd; and Laboratory of Experimental Gastroenterology, Université libre de Bruxelles, Brussels, Belgiume
The standard virologic diagnosis of infection is based on the de-
tection of specific anti-HCV antibodies and cannot distinguish
between past and active infections. Therefore, assays of the HCV
RNA load in serum or plasma have become essential for confirm-
ing active infections and guiding the initiation and continuation
of treatment with pegylated interferon and ribavirin or other an-
detecting and quantitating HCV RNA in clinical practice, with
limits of detection of 10 to 15 IU/ml (4, 7, 15). However, these
assays require high levels of technical skill and are labor-intensive
in routine use.
Hepatitis C virus particles contain a core antigen (Ag) which
free core protein structures (18). Recently, a highly sensitive and
tect HCV Ag test; Abbott Diagnostics, Rungis, France). It has a
cutoff of 3 fmol/liter (0.06 pg/ml). Previous studies using this
assay have indicated that (i) the time course of HCV Ag levels is
similar to those of HCV RNA for all phases of infection (11, 17)
correlated (8–10, 12, 17).
the measurement of intrahepatic HCV Ag levels. The liver is the
primary site of HCV replication, and previous work revealed a
direct correlation between liver and serum HCV RNA levels (16,
20–22). Here, we sought to investigate the applicability of the as-
say to quantitation of HCV Ag in paired serum and liver biopsy
investigated the relationship between the intrahepatic viral load
and the serum viral load, as determined by Ag and RNA assays.
We retrospectively evaluated liver biopsy specimens and sera
collected from 22 chronically HCV-infected patients and 3 HCV-
negative patients. For each patient, a percutaneous liver biopsy
specimen and a serum sample were simultaneously collected and
stored at ?80°C. Epidemiological, clinical, and biochemical data
(age, gender, HCV genotype, fibrosis, and alanine aminotransfer-
ase levels [IU/liter]) were also recorded (Table 1).
epatitis C virus (HCV) is an important human pathogen;
worldwide, over 170 million people are chronically infected.
and homogenized in passive lysis buffer (Promega France, Char-
SAS, Paris, France). The total protein yield was determined on a
NanoDrop spectrophotometer (Thermo Scientific, Courtaboeuf,
was dissociated in RNeasy lysis buffer (Qiagen, Courtaboeuf,
France), as described above. After centrifugation, total RNA was
purified using an RNeasy minikit (Qiagen). The total RNA yield
was determined with a spectrophotometer at 260 nm. Total RNA
was extracted from serum as described previously (3).
The Architect HCV Ag immunoassay was used for the quanti-
tative determination of HCV Ag in paired liver tissue and sera.
The serum concentration of HCV Ag was expressed in pg/ml.
Specimens with concentration values of ?0.06 pg/ml were con-
sidered nonreactive for HCV Ag. For liver tissue, the results were
expressed as pg HCV Ag per mg of total extracted protein. HCV
RNA in paired liver and serum samples was quantitated with a
real-time RT-qPCR assay as previously described (1, 3). Three
negative sera and three negative liver biopsy specimens were in-
cluded as controls.
Statistical analysis was performed using GraphPad Prism5 for
coefficients for serum and liver levels of HCV RNA and Ag were
calculated using Spearman’s rank test. Intergroup comparisons
were performed with the Mann-Whitney U test. A two-sided P
value of ?0.05 was considered statistically significant.
(n ? 22), with amounts varying from 1.27 to 3.20 log10pg/mg
total protein (mean: 2.23 ? 0.52) or, relative to the tissue weight,
3.89 ? 0.60 log10pg/g of liver (range, 2.81 to 5.07). These two
parameters showed a strong positive correlation (r ? 0.87; P ?
0.0001). The mean hepatic HCV RNA level was 5.81 ? 0.55 log10
Received 24 November 2011 Accepted 30 November 2011
Published ahead of print 7 December 2011
Address correspondence to S. Castelain, email@example.com.
Copyright © 2012, American Society for Microbiology. All Rights Reserved.
0095-1137/12/$12.00Journal of Clinical Microbiology p. 465–468 jcm.asm.org
IU/?g of total RNA (range, 4.72 to 6.74) or, expressed in terms of
10.26). These two parameters were also positively correlated (r ?
0.73; P ? 0.0001), suggesting that the quantity of total RNA (and
thus total protein) extracted was related to the weight of liver
tissue. We further observed that HCV Ag and RNA levels in liver
biopsy specimens were highly correlated (r ? 0.83; P ? 0.0001)
(Fig. 1A). On this basis, a quantitative relationship between HCV
RNA and HCV Ag was derived: HCV Ag (log10pg/g) ? 0.8044 ?
HCV RNA (log10IU/g) ? 3.555. We calculated that 1 pg of HCV
Ag per gram was equivalent to approximately 26,276 IU/g (95%
confidence interval [CI], 3.8 ? 103to 2.3 ? 105IU/g).
In serum samples, the HCV Ag titer ranged from 0.61 to 2.19
0.58 log10IU/ml). There was a statistically significant correlation
between HCV Ag and HCV RNA levels in sera (r ? 0.90; P ?
0.0001) (Fig. 1B). We calculated that 1 pg of HCV Ag per ml is
equivalent to 9,755 IU/ml (95% [CI], 3.4 ? 103to 3.1 ? 104
To establish whether the HCV Ag level in liver tissue was re-
lated to the HCV Ag level in serum samples, we performed a cor-
TABLE 1 Characteristics of the HCV-infected patients
Median age ? SD (yr)
Gender (no. of males:females)
Median ALT level ? SD (IU/ml)a
No. with liver fibrosis stageb
F0 or F1
F2 or higher
No. with HCV genotype
52 ? 13
65 ? 46
aThe upper limit of normal for alanine aminotransferase (ALT) is 40 IU/ml.
bLiver fibrosis was graded according to the METAVIR scoring system.
FIG 1 Relationship between the Architect HCV core Ag and HCV RNA concentrations determined in 22 paired samples. (A) Liver biopsy samples; (B) serum
Descamps et al.
jcm.asm.orgJournal of Clinical Microbiology
significant correlation was observed (r ? 0.80; P ? 0.0001) (Fig.
2A). The same comparison was made for HCV RNA level in liver
the 22 HCV-infected patients (data not shown).
This study evaluated the use of the Architect HCV Ag assay to
quantitate HCV Ag in paired serum-liver biopsy specimens from
treatment-naïve, chronically infected patients. For the group of
infected patients tested here, we found a mean HCV Ag level of
measurements appear to be suitable, since there was a significant
observed a strong correlation between concentrations of HCV Ag
and HCV RNA in liver biopsy specimens (r ? 0.80) and in serum
(r ? 0.90). In two previous studies using the same Ag assay and a
commercial RT-qPCR assay, the correlation coefficients for con-
centrations of HCV Ag and HCV RNA in sera were 0.90 (n ? 98)
after therapy (19). We also observed a statistically significant cor-
relation between hepatic and serum levels of both HCV Ag (r ?
0.80) and HCV RNA (r ? 0.87). A number of studies have shown
that the HCV viremia in serum mirrors the intrahepatic HCV
Ag concentration in serum also mirrors the intrahepatic concen-
We estimated that 1 pg of total HCV Ag per ml of serum was
been reported that 1 pg of core Ag per ml is equivalent to approx-
FIG 2 (A) Relationship between HCV core Ag levels in 22 liver-serum pairs, expressed in terms of picograms per ml of serum sample and picograms per
milligram of total liver proteins. (B) Correlation between HCV RNA levels in 22 liver-serum sample pairs, expressed in terms of international units per ml of
serum sample and international units per microgram of total liver RNA.
Liver HCV RNA and Core Antigen Level Correlation
February 2012 Volume 50 Number 2jcm.asm.org 467
beassociatedwithHCVparticles.Theremaindermaythereforebe Download full-text
associated with HCV RNA-free structures (6).
In liver tissue, we demonstrated that 1 pg of total HCV Ag is
can suppose that around two-thirds of the core Ag in liver should
be associated with positive-strand RNA (5, 14). The measured
levels of core Ag suggest overproduction of the latter by infected
hepatocytes and the subsequent release into the blood as RNA-
to-core Ag ratios in serum and liver were not equivalent suggests
the presence of RNA-free core structures secreted by the infected
case for HBsAg, the excess of HCV Ag could function as a decoy
for the immune system. However, larger studies including HCV-
infected individuals with broader clinical status would be needed
to support these data.
related in both the liver and the serum of chronically infected,
treatment-naïve patients. This result demonstrates that serum
liver. On the basis of this good correlation, the newly developed,
automated HCV Ag assay could be used as an alternative to HCV
We are grateful to A. Baron for her expert technical assistance and to C.
This work was funded by the Picardy Regional Council, the Jules
Verne University of Picardy, the Fonds National de la Recherche Scienti-
fique, and the Fonds Emile Defay.
1. Alsaleh K, et al. 2010. Identification of basic amino acids at the
N-terminal end of the core protein that are crucial for hepatitis C virus
infectivity. J. Virol. 84:12515–12528.
2. Bouvier-Alias M, et al. 2002. Clinical utility of total HCV core antigen
quantification: a new indirect marker of HCV replication. Hepatology
3. Castelain S, et al. 2004. TaqMan amplification system with an internal
positive control for HCV RNA quantitation. J. Clin. Virol. 31:227–234.
4. Chevaliez S. 2011. Virological tools to diagnose and monitor hepatitis C
virus infection. Clin. Microbiol. Infect. 17:116–121.
5. Goueslain L, et al. 2010. Identification of GBF1 as a cellular factor re-
quired for hepatitis C virus RNA replication. J. Virol. 84:773–787.
6. Maillard P, et al. 2001. Nonenveloped nucleocapsids of hepatitis C virus
in the serum of infected patients. J. Virol. 75:8240–8250.
7. Matsuura K, et al. 2009. Abbott RealTime hepatitis C virus (HCV) and
Roche Cobas AmpliPrep/Cobas TaqMan HCV assays for prediction of
sustained virological response to pegylated interferon and ribavirin in
chronic hepatitis C patients. J. Clin. Microbiol. 47:385–389.
8. Mederacke I, et al. 2009. Performance and clinical utility of a novel fully
automated quantitative HCV-core antigen assay. J. Clin. Virol. 46:210–
9. Miedouge M, et al. 2010. Analytical evaluation of HCV core antigen and
interest for HCV screening in haemodialysis patients. J. Clin. Virol. 48:
10. Morota K, et al. 2009. A new sensitive and automated chemiluminescent
microparticle immunoassay for quantitative determination of hepatitis C
virus core antigen. J. Virol. Methods 157:8–14.
11. Moscato GA, et al. 2011. Quantitative determination of hepatitis C core
antigen in therapy monitoring for chronic hepatitis C. Intervirology 54:
12. Park Y, Lee JH, Kim BS, Kim DY, Han KH, Kim HS. 2010. New
automated hepatitis C virus (HCV) core antigen assay as an alternative to
real-time PCR for HCV RNA quantification. J. Clin. Microbiol. 48:2253–
13. Pawlotsky JM, et al. 2000. Standardization of hepatitis C virus RNA
quantification. Hepatology 32:654–659.
14. Pawlotsky JM, Chevaliez S, McHutchison JG. 2007. The hepatitis C virus
life cycle as a target for new antiviral therapies. Gastroenterology 132:
15. Pyne MT, Konnick EQ, Phansalkar A, Hillyard DR. 2009. Evaluation of
the Abbott investigational use only RealTime hepatitis C virus (HCV)
assay. J. Clin. Microbiol. 47:2872–2878.
16. Romero-Gomez M, et al. 2001. Intrahepatic hepatitis C virus replication
is increased in patients with regular alcohol consumption. Dig. Liver Dis.
17. Ross RS, et al. 2010. Analytical performance characteristics and clinical
utility of a novel assay for total hepatitis C virus core antigen quantifica-
tion. J. Clin. Microbiol. 48:1161–1168.
18. Schuttler CG, et al. 2004. Variable ratio of hepatitis C virus RNA to viral
core antigen in patient sera. J. Clin. Microbiol. 42:1977–1981.
19. Takahashi M, Saito H, Higashimoto M, Atsukawa K, Ishii H. 2005.
Benefit of hepatitis C virus core antigen assay in prediction of therapeutic
response to interferon and ribavirin combination therapy. J. Clin. Micro-
20. Tedeschi R, et al. 2003. Quantification of hepatitis C virus (HCV) in liver
specimens and sera from patients with human immunodeficiency virus
coinfection by using the Versant HCV RNA 3.0 (branched DNA-based)
DNA assay. J. Clin. Microbiol. 41:3046–3050.
21. Vona G, et al. 2004. Intrahepatic hepatitis C virus RNA quantification in
microdissected hepatocytes. J. Hepatol. 40:682–688.
22. White PA, et al. 2002. Quantification of hepatitis C virus in human liver
Descamps et al.
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