HBV-DNA Concentration Research Articles

Performance of the Cobas AmpliPrep/Cobas TaqMan Real-Time PCR Assay for Hepatitis B Virus DNA Quantification

Hepatitis B virus (HBV) DNA quantification is used to establish the prognosis of chronic HBV-related liver disease, to identify those patients who need to be treated, and to monitor the virologic response and resistance to antiviral therapies. Real-time PCR-based assays are gradually replacing other technologies for routine quantification of HBV DNA in clinical practice. The goal of this study was to evaluate the intrinsic characteristics and clinical performance of the real-time PCR Cobas AmpliPrep/Cobas TaqMan (CAP/CTM) platform for HBV DNA quantification. Specificity was satisfactory (95% confidence interval, 98.1 to 100%). Intra-assay coefficients of variation ranged from 0.22% to 2.68%, and interassay coefficients of variation ranged from 1.31% to 4.13%. Quantification was linear over the full dynamic range of quantification of the assay (1.7 to 8.0 log(10) IU/ml) and was not affected by dilution. The assay was accurate regardless of the HBV genotype. Samples containing HBV DNA levels above 4.5 log(10) IU/ml were slightly underestimated relative to another accurate assay based on branched-DNA technology, but this is unlikely to have noteworthy clinical implications. Thus, the CAP/CTM HBV DNA assay is sensitive, specific, and reproducible, and it accurately quantifies HBV DNA levels in patients chronically infected by HBV genotypes A to F. Samples with HBV DNA concentrations above the upper limit of quantification need to be diluted and then retested. Broad use of fully automated real-time PCR assays should improve the management of patients with chronic HBV infection.

What is the clinical significance of the high prevalence of occult hepatitis B in US liver transplant patients with chronic hepatitis C?

What is the clinical significance of the high prevalence of occult hepatitis B in US liver transplant patients with chronic hepatitis C?

Comparison of Versant HBV DNA 3.0 and COBAS AmpliPrep–COBAS TaqMan assays for hepatitis B DNA quantitation: Possible clinical implications

We compared two commercial assays for HBV DNA quantitation, Versant HBV 3.0, System 340 (bDNA; Bayer Diagnostics) and COBAS AmpliPrep–COBAS TaqMan HBV Test (TaqMan; Roche Diagnostics). Analytical sensitivity, calculated on WHO International Standard, predicted 95% detection rate at 11.4 and 520.2 IU/ml for TaqMan and bDNA, respectively. Specificity, established on 50 blood donor samples, was 100% and 84% for TaqMan and bDNA, respectively. When using clinical samples, HBV DNA was detected by TaqMan in 21/55 samples negative to bDNA. Mean values of HBV DNA obtained with bDNA were higher than those obtained with TaqMan (4.09 log 10 ± 1.90 versus 3.39 log 10 ± 2.41, p < 0.001), and 24.4% of samples showed differences in viral load values >0.5 log 10, without association with HBV genotype. There was a good correlation for HBV DNA concentrations measured by the two assays ( r = 0.94; p < 0.001) within the overlapping range, and the distribution of results with respect to relevant clinical threshold recently confirmed (20,000 and 2000 IU/ml) was similar. Approximately 50% of samples with low HBV DNA, appreciated by TaqMan but not by bDNA, were successfully sequenced in pol region, where drug resistance mutations are located.

Clinical Features and Outcome of Chronic Viral Hepatitis with Acute Exacerbation in Patients with Concurrent Infections of Hepatitis B and C Virus

Studies have shown that concurrent infection of hepatitis B virus and hepatitis C virus may be associated with severe forms of chronic liver disease or with rapid progression. However, very little is known about the role and course of concurrent HBV and HCV infection in patients with acute viral hepatitis. This study retrospectively compared the clinical features of 83 patients diagnosed with HBV- or HCV-related chronic hepatitis with acute exacerbation (12 with concurrent HBV and HCV infection, 46 with HBV infection alone, and 25 with HCV infection alone) encountered at Chia-Yi Chang Gung Memorial Hospital, Taiwan, between January 2003 and December 2005. The clinical course of chronic hepatitis with acute exacerbation in patients with concurrent HBV and HCV infection is similar to patients with single HBV infection, and more severe than patients with single HCV infection, evidenced by increased hepatic decompensation (P = 0.05), failure (P = 0.036), and mortality (P = 0.036). Elevated serum HCVRNA-negative percentage in HBVDNA-positive patients and low serum HBVDNA concentrations in HCVRNA-positive patients imply reciprocal interference of HBV and HCV in patients with concurrent HBV and HCV infections during acute-phase hepatitis. In patients with concurrent HBV and HCV infection, the mortality rate for detectable HBVDNA patients seemed higher than that for undetectable HBVDNA patients, although it did not reach statistical significance (P = 0.066). Virus interference existed in chronic hepatitis with acute exacerbation patients with concurrent HBV and HCV infections. Clinical outcome for patients positive for serum HBVDNA was much worse than those negative for serum HBVDNA. When chronic hepatitis with acute exacerbation occurs in patients with concurrent HBV and HCV infection, aggressive management should be investigated and antiviral therapy targeting of HBV infection should be administered early.

Candidates for therapy: HBV

Hepatitis B may cause liver damage ranging from mild chronic hepatitis to severe active hepatitis, cirrhosis and hepatocellular carcinoma. HIV and HBV co-infection is more likely to lead to lower rates of HBeAg seroconversion, and higher HBV DNA concentrations. Immune restitution may lead to more severe hepatitis. The timing of acquisition of HBV versus HIV will have a bearing on considerations of treatment. Patients may have acquired HIV super-infection of chronic hepatitis B, HBV super-infection of HIV; alternatively, reactivation of hepatitis B may occur in a HIV positive patient, or the patient may be co-infected at diagnosis. The patient may be naïve or experienced or have resistant (HBV) at the time of superinfection. The risk of death is higher in patients with co-infection compared to those with HBV alone. The goals of therapy for hepatitis B are to prevent progression of the disease. If HBV replication can be suppressed, the accompanying reduction in histological activity lessens the risk of progression. Patients may request treatment to reduce infectivity, and this is relevant in co-infected patients. HBV has little effect on HIV or the effect of treatment on HIV; however, HIV, and HIV treatment profoundly affects the natural history of HBV. Therefore, it is usually important to target treatment of HBV to alter the outcome and take into account the impact of HBV treatment on HIV. Special concepts of treatment are applicable in HIV and HBV co-infected patients.

Hepatitis B virus core and core‐related antigen quantitation in Chinese patients with chronic genotype B and C hepatitis B virus infection

Hepatitis B virus (HBV) core-related antigen (HBcrAg) and HBV core antigen (HBcAg) assays were developed for the measurement of serum HBV load. The aim of this study was to assess the clinical utility of these assays in Chinese patients with chronic genotype B and C HBV infection. One hundred and ninety-three chronic hepatitis B patients were enrolled. Serum HBcrAg and HBcAg were measured by chemiluminescence enzyme immunoassay, and HBV-DNA was measured by using a sensitive polymerase chain reaction assay. The data were analyzed in patients with HBV genotype B (HBV/B) and genotype C (HBV/C). The HBcrAg/HBcAg ratio was calculated and compared between patients with and without hepatitis B e antigen (HBeAg). The concentrations of HBcrAg and HBcAg showed significant positive correlation with the HBV-DNA concentration in both HBV/B (r = 0.79, P < 0.001, and r = 0.77, P < 0.001, respectively) and HBV/C (r = 0.87, P < 0.001, and r = 0.90, P < 0.001, respectively). The cut-off for a positive HBcAg corresponded to approximately 4.5 log copies/mL, and that for a positive HBcrAg result corresponded to 3-4 log copies/mL. The HBcrAg/HBcAg ratio was higher in patients with HBeAg than in those without HBeAg. The HBcrAg assay and HBcAg assay are clinically useful in viral quantitation of HBV/B and HBV/C. A combination of these assays would be a valuable tool for analyzing the clinical status of HBV infection.

A clinical prediction model predicted absence of significant fibrosis in chronic hepatitis B

Hui AY, Chan HL, Wong VW, et al . Identification of chronic hepatitis B patients without significant liver fibrosis by a simple noninvasive predictive model. Am J Gastroenterol 2005;100:616–23. [OpenUrl][1][CrossRef][2][PubMed][3][Web...

Significance of hepatitis B virus DNA clearance and early prediction of hepatocellular carcinogenesis in patients with cirrhosis undergoing interferon therapy: Long‐term follow up of a pilot study

Because the anti-carcinogenic effect and mechanism of interferon (IFN) in patients with hepatitis B virus (HBV)-related cirrhosis have not been elucidated, quantitative analysis of HBV-DNA concentration was carried out sequentially. Of 60 consecutive patients with cirrhosis who began IFN therapy between 1986 and 1990, 57 patients were completely observed for the appearance of hepatocellular carcinoma (HCC). All patients underwent intermittent administration of IFN for a median period of 18 months. HBV-DNA was quantified using transcription mediated amplification and hybridization protection assay. A HBV-DNA count <3.7 log-genome equivalent (LGE)/mL (equivalent to 10(3.7) or 5000 copies/mL) was considered to be a negative value. Of 25 patients who had HBV-DNA loss after IFN therapy, nine lost HBV-DNA during the therapy and 16 lost HBV-DNA after cessation of the therapy. The other nine patients showed a transient loss of HBV-DNA, and the remaining 23 retained persistently positive HBV-DNA during and after therapy. Although HCC developed in two (8.0%) of the 25 patients with HBV-DNA loss, carcinogenesis was found in 11 (34.4%) of 32 patients without HBV-DNA loss (Fisher's exact test, P = 0.026). In the two exceptional patients, HCC was detected at 1.2 and 3.6 years after loss of HBV-DNA, respectively. When the HBV-DNA concentration decreased by 2 LGE/mL (decrease to 1/100) at 6 months after initiation of interferon, HBV-DNA became negative eventually in 15 (60.0%) of 25 patients. A significant decrease or loss of serum HBV-DNA prevents development of HCC, and sequential analysis of HBV-DNA could be very useful in both the prediction and the early detection of small HCC.

Interferon-α plus lamivudinevslamivudine reduces breakthroughs, but does not affect sustained response in HBeAg negative chronic hepatitis B

The HBeAg negative form of chronic hepatitis B (CHB) predominates in the Mediterranean area and has a rising frequency in Europe and North America. At present there are three approved therapies for patients with CHB: interferon-alpha (IFN-alpha), lamivudine and adefovir dipivoxil. Unfortunately, none of these drugs are effective in achieving a sustained response in patients with HBeAg negative CHB. Therefore, new therapeutic approaches have been examined. Our aim was to investigate the efficacy of combination treatment of IFN-alpha and lamivudine compared to lamivudine monotherapy, after 24 mo of administration in HBeAg-negative hepatitis B patients. Fifty consecutive patients were randomly assigned to receive IFN-alpha-2b (5 MU thrice per week, n = 24) plus lamivudine (100 mg daily) or lamivudine only (n = 26) for 24 mo. Patients were followed up for further 6 mo. The primary outcome was the proportion with sustained virological response (undetectable serum HBV DNA concentrations) and or sustained biochemical response (transaminase levels within normal range) at 30 mo (6 mo after the end of therapy). Secondary end-points were timed from initial virological (biochemical) response to VBR (BBR, respectively) and the emergence of YMDD mutants across the two arms. Five of twenty-four (21%) patients in the combination arm vs 3/26 (12%) in the lamivudine arm had sustained response (i.e., normal serum transaminase levels and undetectable HBV DNA by PCR assay) 6 mo after treatment discontinuation. A reduction in the emergence of YMDD mutants and in the development of virological breakthroughs was observed in patients receiving combination treatment (10% vs 46%, P = 0.01 and 14% vs 46%, P = 0.03, respectively). Time from initial virologic response to virologic breakthrough (VBR) was greater among initial responders receiving combination treatment compared to those receiving lamivudine (22.9 mo vs 15.9 mo, respectively; P = 0.005). Our results demonstrate that IFN-alpha plus lamivudine combination therapy did not increase the sustained response, compared to lamivudine. However, combination therapy reduces the likelihood of VBR due to YMDD mutants and prolongs the time period until the breakthrough development.

Hepatitis B Virus-Related Hepatocellular Carcinogenesis and Its Prevention

To elucidate the influence of serum hepatitis B virus (HBV) load on hepatocellular carcinogenesis in cirrhotic patients, HBV-DNA was sequentially measured. In a nested, case-control study using 96 patients without antiviral therapy, high HBV-DNA (≧10^3.7 copies/ml) in the last 3 years was significantly associated with carcinogenesis (a patient group without hepatocellular carcinoma (HCC) development; 0/48 vs. a patient group with eventual HCC development; 22/48, p < 0.0001). No patient with a continuously low HBV-DNA for the last 3 years developed HCC. Persistence of high HBV-DNA concentration suggested an increased risk of carcinogenesis. In a retrospective cohort study using 57 patients with interferon therapy, HCC developed in 2 (8.0%) of the 25 patients with HBV-DNA loss, while carcinogenesis was found in 11 (34.4%) of 32 patients without HBV-DNA loss (Fisher’s exact test, p = 0.026). A significant decrease or loss of serum HBV-DNA stops HCC development, and its sequential analysis could be very useful both in the prediction and early detection of small HCC.

Significance of detecting HBV- DNA by the fluorescence quantitative PCR

To study the correlativity between HBV-DNA and the markers of hepatitis B virus infection and different clinical types of hepatitis B. The fluorescence quantitation (FQ) of HBV-DNA of 105 patients with hepatitis B was performed by PCR, and the correlativity between the fluorescence quantitation of HBV-DNA and the markers of hepatitis B virus and different clinical types of hepatitis B was analyzed. Ninety-seven percent of the patients were found HBsAg(+), HBeAg(+), HBcAb(+); 75% were HBsAg(+), HBeAb(+), HBcAb(+); 60% were HBsAg(+), HBcAb(+); 40% were HBsAg(+); in HBsAb(+), HBeAb(+), HBcAb(+) (or both HBsAb and HBcAb were positive) group the HBV DNA was undetectable. The analysis indicated that there was a significant difference among different groups (P less than 0.05).HBV-DNA was detected in 72.2% in acute hepatitis B group, in 75% of chronic hepatitis B group, and in 70% of cases of liver cirrhosis with hepatitis B group. The analysis indicated that there was no significant difference among the different clinical types of hepatitis (P greater than 0.05). The levels of viral replication were not correlated with different clinical types of hepatitis B; the concentration of HBV-DNA in serum was related to hepatitis B antigen.

Reactivation of resolved hepatitis B virus infection after allogeneic haematopoietic stem cell transplantation.

Hepatitis B virus (HBV) reactivation after allogeneic haematopoietic stem cell transplantation (allo-HSCT) is well known in HBsAg-positive carriers but has only occasionally been reported in patients with resolved HBV infection. We investigated six allo-HSCT recipients with pretransplant anti-HBs and anti-HBc antibodies for serologic markers of HBV infection and for the presence of HBV-DNA in serum. Reverse seroconversion, that is, reappearance of HBsAg after a gradual loss of anti-HBs, but no severe liver damage was observed in three patients at 14, 22 and 12 months after HSCT, respectively. There was an increase in HBV-DNA concentration prior to reverse seroconversion. One patient was repeatedly HBV-DNA positive (10(2)-10(3) copies/ml) without reverse seroconversion. Sequencing of the HBsAg and precore region derived from the four HBV-DNA-positive patients showed no relevant mutations. In conclusion, this study demonstrated a high risk (50%) of reverse seroconversion in allo-HSCT recipients with resolved HBV infection. A highly sensitive HBV-DNA assay (TaqMan-PCR) allowed early identification of the individual patients at risk.

Quantifying anti-HBV effect of targeted ribonuclease by real-time fluorescent PCR.

To quantify the inhibition of HBV replication by targeted ribonuclease by using real-time fluorescent PCR. Targeted ribonuclease was introduced into 2.2.15 cells by liposome-mediated transfection or HIV-TAT mediated protein transduction. Forty-eight hours after the transfection and 24 h after the transduction, supernatants of 2.2.15 cells were collected and HBV DNA in the supernatants was quantified by real-time fluorescent PCR with a commercial kit. HBV DNA concentrations in the supernatants of 2.2.15 cells transfected or transducted with targeted ribonuclease were 4.9+/-2.4 x 10(8) copies/L and 8.3+/-4.0 x 10(8) copies/L, respectively. Compared with controls, transfection or transduction of targeted ribonuclease reduced HBV DNA concentration in the supernatants of 2.2.15 cells by 90.4% and 90.1%, respectively (P<0.05). Targeted ribonuclease can inhibit HBV replication in 2.2.15 cells.

核酸増幅検査導入後のＨＢＶウインドウ期の血小板製剤による輸血後肝炎

We experienced a case of HBV infection transmitted through platelet concentrate donated during the window period. Nucleic acid amplification test(NAT)has been performed since February 2000 using 50-pooled samples. The donor was found to be positive by individual nested PCR using stored samples in a retrospective study. The HBV DNA concentration was 2,800 copies ml in the blood. The patient was a 3-year-old boy with acute myeloid leukemia and was negative for HBsAg at notification. Although we injected him with anti-HBs IgG, he became positive for HBsAg 6 months after the infected blood transfusion. He subsequently developed acute hepatitis and progressed to protracted viral infection. HBV nucleotide sequences of the patient and donor were identical. He received allogeneic peripheral blood stem cell transplantation from his HLA-identical mother 11 months after the infected blood transfusion. As the mother was negative for anti-HBs antibody, immunization with HBV vaccination was performed before stem cell harvest. Although the incidence of transfusion-transmitted HBV infection has decreased through the introduction of NAT, this case indicates that NAT using 50-pooled samples may not be effective for the detection of very early-stage HBV infection.

Consistently Low Hepatitis B Virus DNA Saves Patients from Hepatocellular Carcinogenesis in HBV-Related Cirrhosis

Background: To elucidate the influence of serum hepatitis B virus (HBV) load on hepatocellular carcinogenesis in cirrhotic patients, HBV-DNA was sequentially measured. Patients and Methods: Among 160 consecutive patients with HBV-related cirrhosis who received no anti-viral therapy, serial assay of HBV-DNA concentration was available in 146 (91.3%): 48 developed hepatocellular carcinoma (HCC) and 98 did not during a median of 11.7 years. HBV-DNA of 10^3.7 copies/ml or less was considered as low. Results: Of the 48 cases with eventual HCC development (group A), 9 showed intermittently high HBV-DNA, and the other 39 persistently high values. Among 48 age- and sex-matched control patients (group B) selected from the 98 HCC-free patients, 9 had continuously low HBV-DNA, 13 showed a settled down course of HBV-DNA, 9 intermittently high, and the remaining 17 patients demonstrated continuously high HBV-DNA. High HBV-DNA in the last 3 years was significantly associated with carcinogenesis (group A; 0/48 vs. group B; 22/48, p < 0.0001). No patient with a continuously low HBV-DNA for the last 3 years developed HCC. Conclusion: Persistence of high HBV-DNA concentration suggested an increased risk of carcinogenesis. Sequential analysis of HBV-DNA is important in the assessment of the carcinogenesis risk.

Retrospektive Kohortenstudie zur Lamivudintherapie bei Patienten mit chronischer Hepatitis B

Lamivudine, a nucleoside analogue with specific antiviral activity against the hepatitis B virus, is now available as an alternative therapeutic option to standard interferon-alpha treatment in chronic hepatitis B. Larger studies with lamivudine treatment in chronic hepatitis B were mainly performed in North America and Asia. Data on treatment responses in European patients are sparse. Therefore, we evaluated the efficacy and safety of lamivudine therapy in Central European patients and compared the data with the results from international trials. In this retrospective, multicenter, cohort study, 95 patients with chronic hepatitis B (median age: 40.4 years, male: 87 patients, female: 8 patients, HBeAg positive: 47 patients, anti-HBe positive: 48 patients), who were treated with lamivudine (100-300 mg/d per os) between 1997 to 1999, were enrolled. During lamivudine treatment a virologic response with HBeAg to anti-HBe seroconversion was achieved in 22/47 (47%) of the HBeAg positive patients. Pretreatment ALT levels (> threefold the upper limit of normal; p = 0.03) and HBV-DNA serum concentration (< or = 100 pg/ml; p = 0.08) were identified as positive predictors for virologic responses. The virologic response was sustained in six of nine patients who had a follow-up period (median 26 weeks). In anti-HBe positive patients a virologic response with undectable HBV-DNA levels was achieved in 35/48 (73%) patients during lamivudine treatment. Side effects during lamivudine therapy were generally mild and reversible. In this retrospective cohort study virologic end-of-treatment responses to lamivudine monotherapy in patients with chronic hepatitis B were comparable with yet reported international trials. Thus, lamivudine represents a cost-effective and well tolerable option in addition to IFN-alpha in the treatment of chronic hepatitis B.

Second-generation Hybrid capture test and Amplicor monitor test generate highly correlated hepatitis B virus DNA levels

The performance of the Digene Hybrid Capture II HBV DNA Test HC II and the Roche Cobas Amplicor Monitor Test (Cobas-HBV) was evaluated on 252 serum samples. One hundred and seventy-three samples were HBV DNA positive and 75 HBV DNA negative by both assays. Four samples were HBV DNA positive by Cobas-HBV only. Linear regression analysis showed that the HBV DNA concentrations obtained from both assays were significantly related ( n=173, r=0.976, P<0.0001). The results of the study show that Hybrid capture II and Cobas-HBV could be used equally in the management for patients with chronic HBV infection.

Limitations and improvements of the quantiplex branched-DNA assay in Hepatitis B virus-infected patients receiving lamivudine

The branched DNA (bDNA) assay for hepatitis B virus (Chiron Corporation Emerville, USA) was investigated by application to HBV-infected patients in Taiwan, where the B and C genotypes of hepatitis B virus are most prevalent. The study group included sera with hepatitis B surface antigen (HBsAg) and e antigen (HBeAg); Group 1 ( n=70) without treatment; Group 2 ( n=28) lamivudine treatment less than 3 months; Group 3 ( n=73) lamivudine treatment 3–12 months; Group 4 ( n=45) HBeAg-negative sera after 1 year treatment with lamivudine; control group ( n=36) HBsAg-negative sera. Comparison of identical-sample results showed a significantly higher coefficient of variation for low-level HBV DNA (<100 MEq/ml) than for high-level (≥100 MEq/ml), indicating increasing assay inaccuracy uncertainty as the sample HBV DNA concentration decreased. It is thus concluded that low-titered sera should receive special careful pipetting and processing. It was also found that using the relative luminescence of the negative control plus two standard deviations (S.D.) as a new cutoff could promote sensitivity (97.1→97.1%, 89.3→100%, 76.7→84.9%, and 17.8→22.2% in Groups 1–4, respectively) and specificity (94.4→97.2%). In summary, the bDNA HBV assay showed only moderate assay performance for samples with low HBV DNA levels. This problem can be improved partially by choosing a new cutoff value based on the relative luminescence of the negative controls in the kit.

Extended lamivudine treatment in patients with chronic hepatitis B enhances hepatitis B e antigen seroconversion rates: Results after 3 years of therapy

A study in Chinese patients with chronic hepatitis B showed that treatment with lamivudine for 1 year significantly improves liver histology and enhances hepatitis B e antigen (HBeAg) seroconversion compared with placebo. Fifty-eight patients from this 1-year study have received long-term treatment with lamivudine 100 mg; the outcome of 3 years of lamivudine is reported here. Before treatment, all patients had detectable HBeAg. HBeAg seroconversion (HBeAg-negative, anti-HBe–positive), hepatitis B virus (HBV)-DNA suppression, alanine transaminase (ALT) normalization, emergence of YMDD variant HBV, liver histology, and long-term safety were assessed. After 3 years of continuous treatment with lamivudine 100 mg daily, 40% (23 of 58) of patients achieved HBeAg seroconversion. In patients with baseline serum ALT >2× upper limit of normal (ULN), the rate of HBeAg seroconversion was 65% (17 of 26). Median serum HBV-DNA concentrations were below the level of detection, and median ALT concentrations were within the normal range throughout 3 years of treatment. YMDD variant HBV emerged in 33 of 58 (57%) patients during the 3 years, of whom 9 (27%) achieved HBeAg seroconversion (6 after emergence of YMDD variant HBV). ALT levels and histologic scores after emergence of YMDD variant HBV did not show major deterioration. Lamivudine was well tolerated during 3 years of therapy. In conclusion, these data in Chinese patients with chronic hepatitis B show enhanced seroconversion rates with extended lamivudine treatment. Up to two thirds of patients with moderately elevated pretreatment ALT achieved HBeAg seroconversion after 3 years of therapy. (HEPATOLOGY 2001;33:1527-1532.)

Severe hepatitis treated with an artificial liver support system

We designed an artificial liver support system (ALSS) including plasma exchange, charcoal hemoperfusion, plasma bilirubin absorption, charcoal plasma perfusion, hemofiltration and hemodialysis. We chose different methods or their combinations according to the patient's conditions. We investigated the effect of ALSS in 154 patients with severe hepatitis, 72 of whom survived. All data were analyzed by SPSS. The effectiveness of ALSS treatment was compared at different stages (i.e. early, middle and end stages). After each ALSS treatment, the liver function of these patients greatly improved, serum endotoxin and HBV-DNA concentrations were significantly decreased, and the serum concentration of aromatic amino acids (AAA) such as methionine decreased while BCAA/AAA ratio increased. Patients treated with ALSS in the early or middle stages of disease had much higher survival rates than patients in the end stage of disease. Thus, we concluded that ALSS is a reliable therapy for advanced liver diseases and treatment in early or middle stages is appropriate.