Performance verification and comparison of TianLong automatic hypersensitive hepatitis B virus DNA quantification system with Roche CAP/CTM system.

AIM: To investigate and compare the analytical and clinical performance of TianLong automatic hypersensitive hepatitis B virus (HBV) DNA quantification system and Roche CAP/CTM system. METHODS: Two hundred blood samples for HBV DNA testing, HBV-DNA negative samples and high-titer HBV-DNA mixture samples were collected and prepared. National standard materials for serum HBV and a worldwide HBV DNA panel were employed for performance verification. The analytical performance, such as limit of detection, limit of quantification, accuracy, precision, reproducibility, linearity, genotype coverage and cross-contamination, was determined using the TianLong automatic hypersensitive HBV DNA quantification system (TL system). Correlation and Bland-Altman plot analyses were carried out to compare the clinical performance of the TL system assay and the CAP/CTM system. RESULTS: The detection limit of the TL system was 10 IU/mL, and its limit of quantification was 30 IU/mL. The differences between the expected and tested concentrations of the national standards were less than ± 0.4 Log10 IU/mL, which showed high accuracy of the system. Results of the precision, reproducibility and linearity tests showed that the multiple test coefficient of variation (CV) of the same sample was less than 5% for 102-106 IU/mL; and for 30-108 IU/mL, the linear correlation coefficient r2 = 0.99. The TL system detected HBV DNA (A-H) genotypes and there was no cross-contamination during the "checkerboard" test. When compared with the CAP/CTM assay, the two assays showed 100% consistency in both negative and positive sample results (15 negative samples and 185 positive samples). No statistical differences between the two assays in the HBV DNA quantification values were observed (P > 0.05). Correlation analysis indicated a significant correlation between the two assays, r2 = 0.9774. The Bland-Altman plot analysis showed that 98.9% of the positive data were within the 95% acceptable range, and the maximum difference was -0.49. CONCLUSION: The TL system has good analytical performance, and exhibits good agreement with the CAP/CTM system in clinical performance.

Autoantibodies in Chinese patients with chronic hepatitis B: prevalence and clinical associations.

AIM: To investigate the prevalence of autoantibodies and their associations with clinical features in Chinese patients with chronic hepatitis B (CHB). METHODS: A total of 325 Chinese patients with CHB were enrolled in this retrospective, hospital-based study. Patients with chronic hepatitis C (CHC), autoimmune hepatitis (AIH), or primary biliary cirrhosis (PBC) were included, with healthy donors acting as controls. A panel of autoantibodies that serologically define AIH and PBC was tested by indirect immunofluorescence assay and line immunoassay. The AIH-related autoantibody profile included homogeneous anti-nuclear antibodies (ANA-H), smooth-muscle antibodies, anti-liver kidney microsome type 1, anti-liver cytosolic antigen type 1, and anti-soluble liver antigen/liver pancreas; the PBC-related antibodies were characterized by ANA-nuclear dots/membranous rim-like, anti-mitochondrial antibodies-M2 (AMA-M2), anti-BPO (recombinant antigen targeted by AMA-M2), anti-Sp100, anti-promyelocytic leukemia protein (anti-PML), and anti-gp210. The dichotomization of clustering was used to unequivocally designate the AIH or PBC profiles for each case. Anti-Ro52 antibodies were also tested. RESULTS: The prevalence of any autoantibody in CHB amounted to 58.2%, which was similar to the 66.2% prevalence in CHC, significantly higher than the 6.7% in the healthy controls (P < 0.001), and lower than the 100% found in AIH and PBC (P = 0.004 and P < 0.001, respectively). There were more anti-PML and anti-gp210 antibodies among the CHB patients than the CHC patients (11.1% vs 0%, P = 0.003; 12.6% vs 0%, P < 0.001, respectively). The prevalence and titer of AMA, anti-BPO, anti-PML, and anti-gp210 were higher in PBC than in those with CHB. Among the CHB patients, the prevalence of ANA, especially ANA-H, was significantly lower in patients with compensated and decompensated cirrhosis compared with patients without cirrhosis. Thirty-eight cases of hepatocellular carcinoma (HCC) in CHB showed a significant difference compared with non-HCC patients in the prevalence of anti-PML (0% vs 12.5%, P = 0.013). Dichotomization of the autoantibodies revealed that the PBC profile was more prevalent in patients with CHB than in those with CHC, and that it was strongly correlated with both compensated and decompensated cirrhosis. In contrast, the prevalence of the AIH profile was significantly higher in non-cirrhosis patients with CHB than in those with compensated cirrhosis (18.5% vs 8.2%, P = 0.039). Moreover, the AIH profile was also closely associated with hepatitis B e-antigen positivity. CONCLUSION: ANA-H could be an indicator of early-stage CHB. Dichotomizing the autoantibody profiles revealed that the PBC profile is strongly associated with cirrhosis in CHB.

[Dynamic analysis of the lymphocyte subsets in HCV children with different genotypes during treatment].

OBJECTIVE: To discuss the changes of lymphocyte subsets in HCV children with different genotypes during treatment with pegylated interferon alfa-2b and ribavirin. METHODS: The genotype of 45 HCV infected children were identified by real time PCR. The lymphocyte subsets were dynamically detected by BD FACSCalibur flow cytometer with four color MultiTEST IMK Kit during the treatment. RESULTS: For the children with 1b genotype, after 24 weeks, the CD4+ T cells were higher than pre-treatment (P < 0.05). For the children with 2a genotype, after 12 weeks and after 24 weeks, the CD3+ T cells and CD4+ T cells significantly increased while the NK cells decreased than pre-treatment (P < 0.05). CONCLUSIONS: The lymphocyte subsets of HCV children with 2a genotype were different from 1b genotype during trentment with pegylated interferon alfa-2b and ribavirin.

[Cloning and expressing of tissue inhibitor of metalloproteinases I gene fragment and preparation of monoclonal antibodies against the recombinant protein].

OBJECTIVE: To prepare the monoclonal antibody (mAb) against tissue inhibitor of metalloproteinases I (TIMP-I) fusion protein. METHODS: TIMP-I gene was amplified from fibrotic human liver tissue by RT-PCR, then ligated with pQE31 to form recombinant plasmid pQE-TIMP-I and transformed into E. coli BL21. The protein induced by IPTG was purified by 6 x His-tag and used to immunize the BALB/c mice. The specific monoclonal antibodies (mAbs) were prepared by the cell fusion technique. Western Blot were used to detect specificity of mAbs. RESULTS: The prokaryotic plasmid expressing the recombinant protein was constructed, and the TIMP-I recombinant protein was expressed and purified. Four hybridoma cell lines that secreted anti-TIMP-I mAbs were obtained. 3 of 4 mAbs were the IgG1 subtype. Western Blot indicated the mAbs showed specific combination with TIMP-I protein. CONCLUSION: The TIMP-I recombinant protein is highly purified and has strong antigenicity. The anti- TIMP-I mAbs were prepared successfully.

[Establishment of a purificatory method for alpha-fetoprotein variant by affinity adsorption].

OBJECTIVE: To establish a purificatory method of alpha-fetoprotein variant (AFP-L3) based on microspincolumn with lens culinaris agglutinin (LCA). METHODS: LCA was isolated by ammonium sulfate precipitation method from lens culinaris. AFP-L3 affinity adsorption microspincolumns which were made from LCA coupled with activated Sepharose 4B were prepared. By adding into the centrifuge column, serum was absorbed and eluted to purify AFP-L3. The results of purified AFP-L3 detection of 10 cases AFP positive sera by electro-chemiluminescence immunoassay were compared with traditional crossed affinity immunoelectrophoresis. RESULTS: 8 of 10 cases AFP-L3 concentration were greater than 5 ng/ml in purified sera. Six cases show positive reaction in affinity immune cross electrophoresis experiment. CONCLUSION: Successfully established purification method of AFP-L3 by affinity absorption based on microspincolumn. The method was more conducive to clinical laboratory applications due to its high sensitive and easy operation.

[The investigantion of comparative experiments for different clinical laboratory instruments].

OBJECTIVE: To estimate the consistency of two VITROS 3600 chemiluminescent analyzers according to the requirement of ISO15189. METHODS: Verification tests were made for precision and accuracy of anti-HCV in two instruments. While 40 serum samples including Anti-HCV negative (10 cases) , positive (10 cases) , and weakly positive (20 cases). and the test results were statistical analised. RESULTS: Two instruments negative and positive control samples intra-batch precision and coefficients of variation were 5% , 4% and 7. 14% , 7. 23% , inter-batch precision and coefficients of variation were 9. 47% , 7. 7% and 8.04%, 7. 6%, are less than requirement CV (15%) by ISO15189. The accuracy of two instrument were 100% , The test results of the control samples showed no significant difference (P < 0. 05). The correlation analysis of the test results of clinical samples R2 =0. 9984, with good consistency. CONCLUSION: Test results of two Vitros 3600 has good consistency and comparability.

Granulocyte-colony stimulating factor therapy improves survival in patients with hepatitis B virus-associated acute-on-chronic liver failure.

AIM: To evaluate the safety and efficacy of granulocyte-colony stimulating factor (G-CSF) therapy in patients with hepatitis B virus (HBV)-associated acute-on-chronic liver failure (ACLF). METHODS: Fifty-five patients with HBV-associated ACLF were randomized into two groups: the treatment group and the control group. Twenty-seven patients in the treatment group received G-CSF (5 µg/kg per day, six doses) treatment plus standard therapy, and 28 patients in the control group received standard therapy only. The peripheral CD34(+) cell count was measured consecutively by flow cytometry. Circulating white blood cell count, biochemical parameters, and other clinical data of these patients were recorded and analyzed. All patients were followed up for a period of 3 mo to evaluate the changes in liver function and survival rate. RESULTS: The peripheral neutrophil and CD34(+) cell counts in the G-CSF group increased on day 3 from the onset of therapy, continued to rise on day 7, and remained elevated on day 15 compared to those of the control group. Child-Turcotte-Pugh score of patients in the treatment group was improved on day 30 from the onset of G-CSF therapy, compared to that in the controls (P = 0.041). Model for End-Stage of Liver Disease score of patients in the treatment group was improved on day 7 (P = 0.004) and remained high on day 30 from the onset of G-CSF therapy (P < 0.001) compared to that in controls. After 3 mo of follow-up observation, the survival rate in the treatment group (48.1%) was significantly higher than that in the control group (21.4%) (P = 0.0181). CONCLUSION: G-CSF therapy promoted CD34(+) cell mobilization in patients with HBV-associated ACLF, and improved the liver function and the survival rate of these patients.

[Cloning and expressing of golgi protein73 gene fragment and preparation of monoclonal antibodies against the recombinant protein].

OBJECTIVE: To clone and express human Golgi glycoprotein73 protein, and prepare the monoclonal antibody (mAb) against the protein. METHODS: GP73 gene was amplified from HepG2 cells by RT-PCR, then ligated with pQE31 to form recombinant plasmid pQE-GP73 and transformed into E. coli BL21. The protein induced by IPTG was purified by 6 x His-tag and used to immunize the BALB/c mice. The specific monoclonal antibodies (mAbs) were prepared by the cell fusion technique. Western Blot was used to detect specificity of mAbs. RESULTS: The prokaryotic plasmid expressing the recombinant protein was constructed, and the GP73 recombinant protein was expressed and purified. Five hybridoma cell lines that secreted anti-GP73 mAbs were obtained. 2 of 5 mAbs were the IgG1 subtype. Western Blot indicated the mAbs showed specific combination with GP73 protein. CONCLUSION: The GP73 recombinant protein is highly purified and has strong antigenicity. The anti-GP73 mAbs were prepared successfully.

[Distribution of genotypes in ESBLs producing E. coli strains isolated from posthepatitic cirrhosis' patients with bloodstream infection].

OBJECTIVE: To study the genotype distribution of extended-spectrum beta-lactamases (ESBLs) in ESBLs-producing Escherichia coli (E. coli) isolates from posthepatitic cirrhosis' patients with bloodstream infection. METHODS: E. coli were isolated in bloodstream from patients with posthepatitic cirrhosis between January and December in 2011. The strains were identified by VITEK-II. The antibiol susceptibility tests were performed with K-B method. beta-lactamases genes were detected multi-PCR, PCR, sequence and blast. RESULTS: A total of 79 non-duplicate clinical isolates of E coli were consecutively collected from liver cirrhosis' patients with bloodstream infection. There were 20 isolates produced TEM-1 type beta-lactamases and 1 isolate produced SHV-1 typebeta-lactamases. 40 clinical isolates were detected to produce CTX-M type ESBLs, there were 20 CTX-M-1 group and 26 CTX-M-9 group, including 6 stains habouring both CTX-M-1 and CTX-M-9 group. Eight CTX-M genotypes were confirmed by sequencing of the PCR products, including CTX-M-3, CTX-M-14, CTX-M-15, CTX-M-24, CTX-M-28, CTX-M-31, CTX-M-65 and CTX-M-79. CONCLUSION: CTX-M genotype ESBLs was the most popular extended-spectrum beta-lactamases in E. coli isolated from liver cirrhosis' patients with bloodstream infection. The CTX-M-14 is the dominant epidemic type.

[Analysis of hemagglutination inhibition antibody level in patients with influenza A H1N1].

OBJECTIVE: To understand the hemagglutination inhibition antibody level in patients with influenza A H1N1. METHODS: Sera from 28 patients with influenza A H1N1 at different time points after illness onset were collected and measured by hemagglutination inhibition assay. RESULTS: The serum hemagglutination inhibition antibody titers at 1, 5, 15, 22, 37, 49 and 58 days after illness onset were 5.36, 9.39, 39.02, 57.99, 137.92, 55.19 and 57.99 respectively. The top geometric mean titer of hemagglutination inhibition antibody was 148.55. The antibody seroconversion rate and seroprotection rate were occurred in 96.4% (27/28) of patients. CONCLUSION: The patients with influenza A H1N1 have effective immune response.

[Establishment of confirmatory test for suspicious hepatitis B surface antigen positive samples].

OBJECTIVE: Establish a confirmatory test based on ELISA, and use to verify the authenticity of HBsAg weak positive samples, pick and get rid of the false result, and avoid the mistake diagnosis. METHOD: The particles (reagent A) coated by streptavidin and biotinylated HBsAb (reagent B) were mixed in different proportions, then neutralized with serum whose the COI of HBsAg > 20 by ELISA in order to identify the activity of HBsAb in confirmatory reagent. 30 pieces of HBsAg weak positive serum neutralized with the confirmatory reagent, the serum were considered to be positive if rate of decline of HBsAg COI > 50%. The results were compared to Roche confirmatory Kit. RESULT: Confirmatory reagent was able to neutralized with HBsAg. 24 of 30 pieces of HBsAg weak positive samples were judged to be positive, while 6 poeces were negative. The ELISA comfirm method is fully consistent with Roche confirmatory Kit. CONCLUSION: The ELISA confirmatory test for suspicious HBsAg positive samples is a simple, accurate and low cost initial validation method, After further clinical trials, should be widely applied.

[Comparison of two different methods to detect HIV antibodies].

OBJECTIVE: Evaluated the chemiluminescence and enzyme-linked immunosorbent assay (ELISA) to detect HIV antibodies, and compared the results, to provide a reference for the selection and clinical application of HIV screening. METHODS: 3000 cases of our hospital patients were measured by enzyme-linked immunosorbent assay and chemiluminescence immunoassay, using comfirmming experimental results as gold standards. Comparing sensitivity, specificity and other Indicators. RESULTS: In the diagnosis of HIV infection, enzyme-linked immunosorbent assay and chemiluminescence immunoassay had no significant difference. The positive rate of enzyme-linked immunosorbent assay was 0.93%, while the sensitivity and specificity were 89.66%, 99.93%, the positive rate of chemiluminescence immunoassay was 1.03%, while the sensitivity and specificity were 100%, 99.93%, respectively. CONCLUSION: Both methods are suitable for screening of HIV, having high specificity, and chemiluminescence has greater sensitivity than ELISA.

[The early diagnosis value of EV 71 IgM class antibodies in the hand, foot and mouth disease].

OBJECTIVE: Assessment of detection of IgM antibodies for human enterovirus 71 (EV 71) in early diagnosis for the hand, foot and mouth disease (HFMD). METHOD: The sera and throat swabs from 38 patients which were clinical diagnosis as HFMD, were continuous daily collected in our hospital in 2010. These specimens were detected by EV 71 IgM antibodies assay, real time RT-PCR methods for EV 71 and Enterovirus. RESULTS: Among 38 HFMD patients, the cumulative positive rates of EV 71 IgM antibodies were: 60.5% on day 1, 71.1% on day 2, 81.5% in the first 3-4 days, 92.1% on day 5, 92.1% on day 6, and the positive rate of nucleic acid detected by the real time RT-PCR for EV 71 and Enterovirus were 60.5%, 73.6%. CONCLUSION: The positive rate of EV 71 IgM antibodies in the hand, foot and mouth disease just can occur on day 1, and reach to peak on day 5, which can be used as one of indicators of early diagnosis of hand, foot and mouth disease.

[Analysis for post-transplant changes of peripheral lymphocyte subsets in liver transplant receptors with HBV infection].

AIM: To investigate the characters and changes of peripheral lymphocyte subsets and cytokines in liver transplant receptors with HBV infection in short phases after liver transplantation, and to provide evidences for monitoring post-transplant immune condition of liver transplant receptors. METHODS: Peripheral lymphocyte subsets and cytokine levels in pre- and post- transplant 12 h, 3 d, 10 d, 30 d, 60 d of 20 cases of patients with HBV-associated severe hepatic diseases were investigated and analyzed, and were compared respectively with those of 22 cases of healthy adults as control (HC) with flow cytometry (FCM) and ELISA. RESULTS: The patients' accounts of peripheral lymphocyte subsets before liver transplantation were lower than those of HC significantly, but the accounts decreased significantly after transplantation 12 h. Three days later, the accounts of lymphocyte subsets increased significantly. The percentages of CD3, CD4, CD8 and NK cells got to stable stage from post-transplantation 10 d, and the absolute accounts of post-transplantation 60 d were higher than those of pre-transplantation, but were still lower than those of HC; The IFN-γ and IL-10 levels of post-transplantation 12 h increased several times and decreased after 3 days. The IL-10 levels in post-transplantation 60 d were still higher than those of HC. CONCLUSION: The absolute accounts of peripheral lymphocyte subsets increased to stable levels from post-transplantation 10 d, but were still lower than those of HC; Post-transplant immune condition was to Th2 polarization.

[The verification of a new DNA double-detection diagnostic kit for identifying novel influenza A (H1N1)].

OBJECTIVE: To verify a new kit of "universal and novel influenza A (H1N1) virus nucleic acid double-detection methods (PCR-fluorescence probe)". METHODS: 150 cases of throat swab specimens were collected consecutively. After RNA was extracted, the specimens were detected by the verified kit. At the same time, the same specimens were detected by Real-time PCR diagnostic kit from Beijing CDC as the control. The data were analysed by the Kappa in agreement and by McNemar chi2 in difference test. RESULTS: The consistency rate of the verified kit and the Beijing CDC kit was universal primer M 97.33%, H1N1 98.67% respectively. The Kappa test and McNemar chi2 test showed that two methods had a higher consistency. Compared to the CDC kit, the "false negative rate" and "false-positive rate"of double-check kit were lower. CONCLUSION: The kit of "universal and novel influenza A (H1N1) virus nucleic acid double-detection methods (PCR-fluorescence probe)" from Shanghai Kehua Bio-Engineering Co., Ltd can be used to detect influenza A and novel influenza A (H1N1).

[Quantitative analyses of intrahepatic HBV cccDNA and serum HBsAg in 54 patients with chronic hepatitis B].

OBJECTIVE: To quantitatively detect intrahepatic HBV covalently closed circular DNA (cccDNA) and serum HBsAg; and to analyze the relationship between the two parameters and with serum HBV DNA level. METHODS: Intrahepatic cccDNA (copies/cell) was quantitated by plasmid-safe ATP-dependent Danes (PSAD) digestion in combination with rolling circle amplification and gap-spanning selective real-time PCR assay using formalin fixed paraffin-embedded liver biopsy samples. HBsAg was measured by chemiluminescence's reagent manufactured by Abbott Company using sera sampled at time-point of liver biopsy. RESULTS: Intrahepatic cccDNA level was positively correlated with serum HBsAg level (r = 0.459, P < 0.001), but not correlated with serum HBV DNA level. Serum HBsAg level was positively correlated with serum HBV DNA level (r = 0.328, P = 0.015), and reversely correlated with HBV replicative efficiency defined as the ratio of serum HBV DNA to cccDNA (r = -0.373, P = 0.005). CONCLUSION: In patients with chronic hepatitis B, intrahepatic cccDNA level is correlated with serum HBsAg level. The two parameters combined with serum HBV DNA may comprehensively reflect HBV replication activity and help evaluation of antiviral therapeutic efficacy.

[Establishment of confirmatory test for HBsAb in serum of coexistence of hepatitis B surface antigen and antibodies to HBsAg].

OBJECTIVE: Establish a kind of confirmation method based on ELISA, and use to verify authenticity of HBsAb + in HBsAg + HBsAb + serum, pick and get rid of the false masculine gender the result, and avoid the mistake diagnosis. METHOD: Collect 60 pieces of serum whose thick degree of HBsAg at 1000 COI above tested by ECLIA as confirm serum, mixed the confirm serum of different dilution with HBsAb positive serum to screen and verify best thick degree of HBsAg. Collected 40 pieces of HBsAg + HBsAb + serum, ELISA tested the descend rate of HBsAb COI after neutralized with confirm serum in order to confirm authenticity of HBsAb + in pieces of HBsAg + HBsAb + serum. RESULT: When thick degree of HBsAg is 2000 COI, the performance of neutralization to HBsAb is best. The ELISA confirmatory test is fully consistent with the ECLIA method with true positive of 37 pieces of HBsAg + HBsAb + serum while false-positive of 3 pieces of serum. CONCLUSION: The ELISA confirm method is a simple, accurate and low cost initial validation method.

[Evaluation of the Helicobacter pylori stool antigen test].

OBJECTIVE: To evaluate the clinical value of an enzyme-linked immunosorbent assay by the Helicobacter pylori stool antigen (HpSA) test for the detection of H. pylori infection. METHODS: 328 patients were measured upper gastrointestinal endoscopic examination which as gold standards and HpSA test in the meantime, comparing accuracy, sensitivity, specificity and other Indicators. RESULTS: In the diagnosis of Hp infection, HpSA test had no significant difference comparing with gold standards and had a P value of over 0.5. The sensitivity of HpSA test was 94.6%, while the specificity, veracity, expected positive value, and expected negative value were 96.9%, 96.3%, 89.7% and 98.4%, respectively. CONCLUSION: The H. pylori stool antigen test is a simple, non-invasive method for accurate diagnosis of H. pylori infection.

[Clinical and laboratory characteristics of a new influenza A (HIN1) epidemic].

OBJECTIVE: To analysis the clinical and laboratory characteristics of Patients infected with new influenza A (HIN1) virus. METHODS: All cases with new influenza A (H1N1) confirmed on polymerase chain reaction assay on throat swabs. There were included in a prospective evaluation of clinical characteristics, laboratory results, treatment and overcome of new influenza A (H1N1). RESULTS: There were 35 patients in the epidemic. Clinical illness developed within a mean of 1.7 days. Fever occurred in 97.1%, sore throat 65.7% cough 51.4%, headache 28.6%, and myalgia 31.4%. All patients were treated with oseltamivir lasted 5 days. The mean duration of viral shedding was 4.5 days. All were cured and left hospital after day 7. CONCLUSION: It was infected by new influenza A (H1N1) typically in this epidemic.

[Changes and analysis of peripheral white blood cells and lymphocyte subsets for patients with pandemic influenza A virus (H1N1) infection].

OBJECTIVE: To investigate the characters and changes of peripheral white blood cells and lymphocyte subsets of patients with pandemic influenza A virus (H1N1) infection and to provide evidences for diagnosis, treatment and prognosis of influenza A (H1N1) infection. METHODS: Peripheral white blood cell parameters and the percentages of lymphocyte subsets in acute and recovery phases of 59 cases of influenza A virus (H1N1) infectious patients (42 mild cases and 17 severe cases) were investigated and analyzed, and compared respectively with those of 43 cases of healthy adults as control (HC) and 24 cases of general influenza A virus (no-H1N1) infectious using whole blood cell analysis and flow cytometry. RESULTS: Peripheral white blood cell counts of mild cases decreased greatly but those of severe cases did not decrease significantly; the neutrophils of severe cases increased significantly in acute phase; similar to general influenza A virus (no-H1N1) infectious, the peripheral lymphocytes, CD3, CD4, CD8 and B cells of all patients with influenza A virus (H1N1) infection decreased greatly in acute phase and quickly recovered in recovery phase; NK and NKT cells absolute counts of severe cases decreased significantly in acute phase, and the decreasing extent of which were more than 20%. CONCLUSION: There were similar characteristics of change in peripheral white blood cells and lymphocyte subsets between patients with pandemic influenza A virus (H1N1) infection and general pandemic A virus (No-H1N1); the great decrease of NK and NKT cells absolute counts may suggest the severe tendency of diseases.