Application of a Newly Developed High-Sensitivity HBsAg Chemiluminescent Enzyme Immunoassay for Hepatitis B Patients with HBsAg Seroclearance

Abstract

We modified and automated a highly sensitive chemiluminescent enzyme immunoassay (CLEIA) for surface antigen (HBsAg) detection using a combination of monoclonal antibodies, each for a specific epitope of HBsAg, and by improving an earlier conjugation technique. Of 471 hepatitis B virus (HBV) carriers seen in our hospital between 2009 and 2012, 26 were HBsAg seronegative as determined by the Abbott Architect assay. The Lumipulse HBsAg-HQ assay was used to recheck those 26 patients who demonstrated seroclearance by the Abbott Architect assay. The performance of the Lumipulse HBsAg-HQ assay was compared with that of a quantitative HBsAg detection system (Abbott Architect) and the Roche Cobas TaqMan HBV DNA assay (CTM) (lower limit of detection, 2.1 log copies/ml) using blood serum samples from patients who were determined to be HBsAg seronegative by the Abbott Architect assay. Ten patients had spontaneous HBsAg loss. Of 8 patients treated with nucleotide analogues (NAs), two were HBsAg seronegative after stopping lamivudine therapy and 6 were HBsAg seronegative during entecavir therapy. Eight acute hepatitis B (AH) patients became HBsAg seronegative. Of the 26 patients, 16 were HBsAg positive by the Lumipulse HBsAg-HQ assay but negative by the Abbott Architect assay. The differences between the two assays in terms of detectable HBsAg persisted over the long term in the spontaneous loss group (median, 10 months), the NA-treated group (2.5 months), and the AH group (0.5 months). In 9 patients, the Lumipulse HBsAg-HQ assay detected HBsAg when HBV DNA was negative by the CTM assay. HBsAg was also detected by the Lumipulse HBsAg-HQ assay in 4 patients with an anti-HBs concentration of >10 mIU/ml, 3 of whom had no HBsAg escape mutations. The automatic, highly sensitive HBsAg CLEIA Lumipulse HBsAg-HQ is a convenient and precise assay for HBV monitoring.T oday, Ͼ400 million people worldwide are hepatitis B virus (HBV) carriers (1). We have monitored HBV markers, such as HBV DNA, hepatitis B surface antigen (HBsAg), hepatitis B e antigen (HBeAg), and HB core-related antigen (HBcrAg), in chronic hepatitis B patients. The measurement of HBV DNA levels by a PCR-based method is the state-of-the-art technique for monitoring HBV replication in clinical practice (2). However, it is suboptimal for chronic hepatitis B patients who are medicated with nucleotide analogues (NAs), as those, in many cases, can decrease HBV DNA to below the limit of detection.HBsAg is a secreted envelope protein that is continuously shed into the blood as long as HBV infection persists, irrespective of viral replication. Recent advances in HBsAg quantification (qHBsAg) have opened up new perspectives in the study of HBV; qHBsAg levels are correlated with intrahepatic covalently closed circular (ccc) DNA, which is used as a template for viral transcription and maintains the chronic HBV infection state (3-5). Additionally, a correlation between qHBsAg and HBV DNA has been suggested, with the possibility of a role for qHBsAg as a surrogate marker for viral r...

Clinical characteristics and molecular analysis of hepatitis B virus reactivation in hepatitis B surface antigen-negative patients during or after immunosuppressive or cytotoxic chemotherapy

“…At the time of reactivation, 11 patients which show in Table 4 were measured by Lumipulse HBsAg-HQ assay. This assay used two monoclonal antibodies against the external structural region as determinant ''a'' and the internal epitope as the capture target and this system can overcome limitation such as HBV S gene mutants associated with failed HBsAg detection and difference of HBV subgenotypes [18].…”

“…The quantitative HBsAg detection was used chemiluminescent enzyme immunoassay (Lumipulse HBsAg-HQ assay Fujirebio, Inc. Tokyo, Japan) [18].…”

Evidence of serologic activity in chronic hepatitis B after surface antigen (HBsAg) seroclearance documented by conventional HBsAg assay

“…A recent study employed an innovative and highly sensitive chemiluminescent enzyme immunoassay (CLEIA) for the quantitative detection of HBsAg (prototype) [18]. Using a combination of monoclonal antibodies, targeting both the exposed common determinant “a” of the surface antigen and the epitope embedded inside the lipid bilayer of the viral envelope, this linearized HBsAg assay is able to identify HBsAg mutants that evade detection by current serologic assays [13].…”

Ultradeep Sequencing for Detection of Quasispecies Variants in the Major Hydrophilic Region of Hepatitis B Virus in Indonesian Patients

“…The serum HBsAg titer was quantified in all the subjects using the Lumipulse HBsAg-HQ assay, using a fully automated chemiluminescent enzyme immunoassay system (Lumipulse; Fujirebio, Tokyo, Japan). The sensitivity of the Lumipulse HBsAg-HQ assay is 0.005 IU/ml, and the coefficient of variation in this assay is Ͻ5.9% for samples with a low concentration of HBsAg (25). Antibodies to the hepatitis B surface antigen (anti-HBs) and hepatitis B e antigen (HBeAg) were assayed with chemiluminescence immunoassays (CLIA) (Architect AUSAB and Architect HBeAg, respectively; Abbott Japan).…”