Objective- To compare the consistency of the results from detecting HIV-1 antibody in the paired urine and serum specimens from drug users by ELISA. Methods: The paired urine and serum specimens from 273 drug users de...Objective- To compare the consistency of the results from detecting HIV-1 antibody in the paired urine and serum specimens from drug users by ELISA. Methods: The paired urine and serum specimens from 273 drug users detained at a detoxification unit were collected, and the HIV-1 antibodies in the specimens of them were screened by urine and serum ELISA kits, respectively. Results: Of 273 serum specimens, 94 ones showed positive reaction and among 94 counterpart urine specimens, 93 ones also appeared positive reaction. Taking the results together,the consistent rate of HIV-1 antibody screened by urine and serum ELISA kits was 99.6%. Conclusion: The urine ELISA kit, which screened HIV-1 antibody of urine showing almost the same results tested by serum ELISA kit, is reliable. It is proposed that urine ELISA be introduced in many fields.展开更多
Photodynamic therapy(PDT)has limited effects in treating metastatic breast cancer.Immune checkpoints can deplete the function of immune cells;however,the expression of immune checkpoints after PDT is unclear.This stud...Photodynamic therapy(PDT)has limited effects in treating metastatic breast cancer.Immune checkpoints can deplete the function of immune cells;however,the expression of immune checkpoints after PDT is unclear.This study investigates whether the limited e±cacy of PDT is due to upregulated immune checkpoints and tries to combine the PDT and immune checkpoint inhibitor to observe the e±cacy.A metastatic breast cancer model was treated by PDT mediated by hematoporphyrin derivatives(HpD-PDT).The anti-tumor effect of HpD-PDT was observed,as well as CD4þT,CD8þT and calreticulin(CRT)by immunohistochemistry and immunofluorescence.Immune checkpoints on T cells were analyzed byflow cytometry after HpD-PDT.When combining PDT with immune checkpoint inhibitors,the antitumor effect and immune effect were assessed.For HpD-PDT at 100 mW/cm2 and 40,60 and 80 J/cm2,primary tumors were suppressed and CD4þT,CD8þT and CRT were elevated;however,distant tumors couldn't be inhibited and survival could not be prolonged.Immune checkpoints on T cells,especially PD1 and LAG-3 after HpD-PDT,were upregulated,which may explain the reason for the limited HpD-PDT effect.After PDT combined with anti-PD1 antibody,but not with anti-LAG-3 antibody,both the primary and distant tumors were signi-cantly inhibited and the survival time was prolonged,additionally,CD4þT,CD8þT,IFN-þCD4þT and TNF-þCD4þT cells were signi-cantly increased compared with HpD-PDT.HpD-PDT could not combat metastatic breast cancer.PD1 and LAG-3 were upregulated after HpD-PDT.Anti-PD1 antibody,but not anti-LAG-3 antibody,could augment the antitumor effect of HpD-PDT for treating metastatic breast cancer.展开更多
BACKGROUND The role of diverse antibodies in mediating peripheral nerve injury in Guillain-Barrésyndrome(GBS)is becoming clearer,but positivity for multiple antibodies in one case is uncommon.To our knowledge,thi...BACKGROUND The role of diverse antibodies in mediating peripheral nerve injury in Guillain-Barrésyndrome(GBS)is becoming clearer,but positivity for multiple antibodies in one case is uncommon.To our knowledge,this is the first case involving GBS with positive anti-sulfatide,anti-GT1a,and anti-GT1b antibodies.CASE SUMMARY A 20-year-old female patient was admitted to the hospital due to weakness of limbs for 5 d,and deterioration of the weakness and muscle aches for 1 d.The patient's limbs were weak,but the tendon reflexes in the part of the limbs were normal.There was no comorbid peripheral nociception or deep sensory dysfunction.She was diagnosed with GBS and was discharged after receiving intravenous human immunoglobulin pulse therapy.CONCLUSION In this article,the clinical manifestations,neurophysiological examination,and auxiliary examination findings of a GBS patient positive for multiple antibodies were analyzed to improve the identification of the disease by clinical physicians at an early stage.展开更多
BACKGROUND Miller fisher syndrome(MFS)is a variant of Guillain-Barrésyndrome,an acute immune-mediated peripheral neuropathy that is often secondary to viral infections.Anti-ganglioside antibodies play crucial rol...BACKGROUND Miller fisher syndrome(MFS)is a variant of Guillain-Barrésyndrome,an acute immune-mediated peripheral neuropathy that is often secondary to viral infections.Anti-ganglioside antibodies play crucial roles in the development of MFS.The positive rate of ganglioside antibodies is exceptionally high in MFS patients,particularly for anti-GQ1b antibodies.However,the presence of other ganglioside antibodies does not exclude MFS.CASE SUMMARY We present a 56-year-old female patient who suddenly developed right blepharoptosis and progressively worsening vision in both eyes.There were flu symptoms prior to onset,and a coronavirus disease 2019 test was positive.On physical examination,the patient exhibited bilateral extraocular muscle paralysis,weakened reflexes in both limbs,and impaired coordination.The cerebrospinal fluid examination results showed no obvious abnormalities.Bilateral peroneal nerve F-waves were not extracted.Serum anti-GD1b IgG and anti-GT1a IgG antibodies were positive.The patient received intravenous methylprednisolone(1000 mg/day),with the dosage gradually decreased.Additionally,intravenous high-dose immunoglobulin treatment was administered for 5 days(0.4 g/kg/day)from day 2 to day 6 of hospitalization.The patient’s symptoms improved after treatment with immunoglobulins and hormones.CONCLUSION Positive ganglioside antibodies may be used as supporting evidence for the diagnosis;however,the diagnosis of MFS is more reliant on clinical symptoms.展开更多
Despite enormous efforts to achieve the goal of eliminating mother-to-child transmission of HIV-1, it remains a major challenge for many countries in sub-Saharan Africa, particularly Mali. Our objective is to assess c...Despite enormous efforts to achieve the goal of eliminating mother-to-child transmission of HIV-1, it remains a major challenge for many countries in sub-Saharan Africa, particularly Mali. Our objective is to assess changes in the rate of mother-to-child transmission of HIV-1. We conducted a cross-sectional study between January 1, 2009 to December 31, 2018 (10 years) of early diagnosis activity in newborns and children born to HIV-1-positive mothers at the National Institute for Public Health (INSP). The samples came from health and referral centers in mali. All samples were received at the Laboratory of Molecular Biology at the INSP. Proviral DNA extraction was performed from a blood spot sample with a Roche DNA kit, Cobas AmpliPrep/Cobas TaqMan HIV-1 qualitative Test, V2.0 (Roche Molecular System, Inc, USA) following the company procedures. Molecular diagnosis was performed using the same kits using an algorithm of three identical PCRs. The Epi Info version 7 software was used for data analysis with a significance threshold of 5%. A total of 10,714 samples of infants and children born to HIV-positive mothers were analyzed by PCR. Ninety-six percent of mothers were on ARV prophylaxis (AZT 3TC NVP and AZT NVP) and 60% of newborns received the same ARV prophylaxis. Of these children, 956 tested positive with an overall transmission rate of 8.92%, varying between 7.27% in 2009 and 08.01% in 2018. This rate was relatively low among children receiving prophylaxis at 2.04% and remained high for children who received breastfeeding at 5.62%. However, the transmission rate remains low for those who have benefited from mixed and artificial breastfeeding at 1.58% and 1.27% respectively. A significant proportion of children remained infected by their mothers during pregnancy, childbirth or breastfeeding. This study shows the importance of early diagnosis of HIV in children using molecular technology.展开更多
The emergence of antiretroviral resistance mutations represents a major threat to the achievement of national and global goals for the elimination of HIV-1 infection. The global strategy in 2019 in Cte d'Ivoire is...The emergence of antiretroviral resistance mutations represents a major threat to the achievement of national and global goals for the elimination of HIV-1 infection. The global strategy in 2019 in Cte d'Ivoire is a new national policy for the management of people living with HIV with the administration of dolutegravir (DTG)-based fixed-dose combination. The aim of our study was to evaluate HIV-1 resistance to antiretrovirals (ARVs) in infected adult subjects in Cte d’Ivoire in the context of a systematic switch to a DTG-based combination. Between February 2022 and October 2023, a cross-sectional survey with random sampling was conducted in 06 services caring for people living with HIV. A total of 139 participants were included in the study. Adults with a viral load ≥ 1000 copies/mL were tested for HIV-1 ARV resistance mutations. Molecular analyses were performed using protocol of ANRS-MIE (National Agency for Research on AIDS and emerging infectious diseases). The interpretation is performed by HIVGRAD (https://www.hiv-grade.de/cms/grade/). The frequencies of HIV-1 resistance to non-nucleotide reverse transcriptase inhibitors (NNRTIs), nucleotide reverse transcriptase inhibitors (NRTIs), integrase inhibitors (IINTs) and protease inhibitors (PIs) were 82%, 73%, 19% and 11% respectively. The main mutations observed in the different classes were K103N (45%), M184V (64%), E157Q (19%) and L10V/M46I/A71V/I54V (6%) respectively. This study reveals the emergence of resistance to DTG-based fixed-dose combinations, favored by high rates of resistance to NRTIs and NNRTIs. This finding underlines the need for enhanced viral load monitoring and HIV-1 genotyping tests to guide the choice of NRTIs for combination therapy. In addition, monitoring for mutations to second-generation NRTIs is essential, given the scale-up of DTG-based regimens currently underway in Cte d’Ivoire.展开更多
Introduction: HIV, the human immunodeficiency virus, is the etiological agent of acquired immunodeficiency syndrome (AIDS). The aim of this study was to assess the evolution of the viral load in patients under treatme...Introduction: HIV, the human immunodeficiency virus, is the etiological agent of acquired immunodeficiency syndrome (AIDS). The aim of this study was to assess the evolution of the viral load in patients under treatment. Methodology: This was a study carried out from July 2017 to June 2022 at the Point G University Hospital laboratory. The determination of the viral load of patients was carried out by PCR on the ABOTT M2000sp/rt platform. Results: A total of 129 patients infected with HIV-1, aged 19 to 72 years with a mean age of 40.05 years ± 10.71;all on antiretroviral chemotherapy. The female gender predominated among our patients. The most common treatment regimen was 2INTI + 1INNTI with 72.9% followed by 2INTI + 1INI with 13.2%. As for the combinations of molecules, the combination TDF + 3TC + EFV and TDF + 3TC + DTG predominated, respectively 65.1% and 13.2%. 89.9% of our patients had undetectable viremia after 12 months of treatment (p < 0.005) with an average viral load which had evolved from 681315.65 copies/ml ± 1616908.484 to M0 at 5742.36 copies /ml ± 35756.883 at M12 (p Conclusion: Generally speaking, antiretroviral treatment had contributed to controlling viral loads, however the therapeutic combination TDF + 3TC + DTG had made it possible to obtain more patients with undetectable viremia instead.展开更多
To prepare HIV-1 Vif and hAPOBEC3G and to produce their antibodies, the full length gene fragment of HIV-1 vif was amplified by PCR from a plasmid of HIV-1 NL4.3 cDNA, and the APOBEC3G gene was obtained by RT-PCR from...To prepare HIV-1 Vif and hAPOBEC3G and to produce their antibodies, the full length gene fragment of HIV-1 vif was amplified by PCR from a plasmid of HIV-1 NL4.3 cDNA, and the APOBEC3G gene was obtained by RT-PCR from the total RNA of H9 cells. The resulting DNA construct was cloned into a prokaryotic expression vector (pET-32a). Recombinant pET-vif and pET-APOBEC3G were expressed respectively in Eserichia coli BL21 (DE3) as an insoluble protein. The vector also contained a six-histidine tag at the C-terminus for convenient purification and detection. To express and purify the HIV-1 Vif and hAPOBEC3G in E.coli cells, the accuracy of inserted gene and specificity of proteins were detected by the two enzyme digestion method, SDS-PAGE, and Western blotting. Rabbits were then immunized by Vif or APOBEC3G protein and serum samples were tested by indirect ELISA to determine the level of antibodies. Immunoenzyme and immunofluorescence assays were performed to identify the specificity of polyclonal antibodies. The titer of the anti-Vif antibodies was 1:204800, and that of the anti-APOBEC3G antibodies was 1:102400. Thus the antibodies could detect the antigen expression in the cells, demonstrating that fusion proteins with high purity and their corresponding polyclonal antibodies with high titer and specificity were achieved.展开更多
The cytokine repertoire of ADP/ATP carrier-specific humoral immune responses and the cytokine-dependent anti-ADP/ATP carrier antibody IgG subclasses were examined in a cohort of ADP/ATP carrier-immunized BALB/c mice t...The cytokine repertoire of ADP/ATP carrier-specific humoral immune responses and the cytokine-dependent anti-ADP/ATP carrier antibody IgG subclasses were examined in a cohort of ADP/ATP carrier-immunized BALB/c mice treated with anti-CD4 monoclonal antibody. Eighteen male BALB/c mice （6–8 weeks old） were randomized into 3 groups： dilated cardiomyopathy （DCM） group, DCM-tolerance （Tol） group and control group. The mice in DCM group were immunized with the peptides derived from human ADP/ATP carrier protein for 6 months and mice in the control group were sham-immunized, while the mice in DCM-Tol group were immunized with ADP/ATP carrier protein and anti-CD4 McAb simultaneously. Serum autoantibody against ADP/ATP carrier and IgG subclasses were measured by ELISA, intracellular cytokines IFN-γ and IL-4 of Th cells were moni- tored with flow cytometry, and splenic T cell cytokines IFN-γ, IL-2, IL-4 and IL-6 were detected by using real-time fluorescent quantitative PCR. The results showed that the autoantibody against ADP/ATP carrier was found in all mice in DCM group, and the antibody level, serum IgG1 and IgG2a subclasses, cytokines in T cells and Th cells were all elevated in DCM group, as compared with those in control group （P〈0.01）. On the other hand, in DCM-Tol group, the autoantibody level and contents of all the cytokines were significantly different from those in DCM group （P〈0.01）, and were close to those in control group. And the levels of IgG1, IgG2a, IgG2b and IgG3 were influenced, to varying degrees, by anti-CD4 McAb as compared with those in DCM group. All these four types of IgG subclasses were substantially decreased in DCM-Tol group as compared with DCM group. It is concluded that the treatment with anti-CD4 McAb could prevent the activation of T cells, reverse the abnormal secretion of cytokines and the imbalance between Th1/Th2 cell subsets and abnormal production of autoantibody against ADP/ATP carrier, and eventually avoid myocardial injuries.展开更多
Simpli RED HIV-1 antibody test was used to detect 17,378 samples from people of entering coutry. 10 samples were positive. Using Western Blot as the cofirmatory test, 8/10 was positive. The advantages of this test was...Simpli RED HIV-1 antibody test was used to detect 17,378 samples from people of entering coutry. 10 samples were positive. Using Western Blot as the cofirmatory test, 8/10 was positive. The advantages of this test was very sensitive, quick and easy. 20 ul whole blood could be used as sample and got the results in two minutes.展开更多
文摘Objective- To compare the consistency of the results from detecting HIV-1 antibody in the paired urine and serum specimens from drug users by ELISA. Methods: The paired urine and serum specimens from 273 drug users detained at a detoxification unit were collected, and the HIV-1 antibodies in the specimens of them were screened by urine and serum ELISA kits, respectively. Results: Of 273 serum specimens, 94 ones showed positive reaction and among 94 counterpart urine specimens, 93 ones also appeared positive reaction. Taking the results together,the consistent rate of HIV-1 antibody screened by urine and serum ELISA kits was 99.6%. Conclusion: The urine ELISA kit, which screened HIV-1 antibody of urine showing almost the same results tested by serum ELISA kit, is reliable. It is proposed that urine ELISA be introduced in many fields.
基金supported by the National Key Research and Development Program of China[2018YFB0407200]National Natural Science Foundation of China[61975239]Medical and Health Technology Innovation Project of the Chinese Academy of Medical Sciences[2019-I2M-5061].
文摘Photodynamic therapy(PDT)has limited effects in treating metastatic breast cancer.Immune checkpoints can deplete the function of immune cells;however,the expression of immune checkpoints after PDT is unclear.This study investigates whether the limited e±cacy of PDT is due to upregulated immune checkpoints and tries to combine the PDT and immune checkpoint inhibitor to observe the e±cacy.A metastatic breast cancer model was treated by PDT mediated by hematoporphyrin derivatives(HpD-PDT).The anti-tumor effect of HpD-PDT was observed,as well as CD4þT,CD8þT and calreticulin(CRT)by immunohistochemistry and immunofluorescence.Immune checkpoints on T cells were analyzed byflow cytometry after HpD-PDT.When combining PDT with immune checkpoint inhibitors,the antitumor effect and immune effect were assessed.For HpD-PDT at 100 mW/cm2 and 40,60 and 80 J/cm2,primary tumors were suppressed and CD4þT,CD8þT and CRT were elevated;however,distant tumors couldn't be inhibited and survival could not be prolonged.Immune checkpoints on T cells,especially PD1 and LAG-3 after HpD-PDT,were upregulated,which may explain the reason for the limited HpD-PDT effect.After PDT combined with anti-PD1 antibody,but not with anti-LAG-3 antibody,both the primary and distant tumors were signi-cantly inhibited and the survival time was prolonged,additionally,CD4þT,CD8þT,IFN-þCD4þT and TNF-þCD4þT cells were signi-cantly increased compared with HpD-PDT.HpD-PDT could not combat metastatic breast cancer.PD1 and LAG-3 were upregulated after HpD-PDT.Anti-PD1 antibody,but not anti-LAG-3 antibody,could augment the antitumor effect of HpD-PDT for treating metastatic breast cancer.
基金The Scientific and Technological Innovation Talent Team Project of Zunyi City,No.[2022]2Guizhou Maotai Hospital Research and Talent Cultivation Funding Project,No.MTYK 2022-06.
文摘BACKGROUND The role of diverse antibodies in mediating peripheral nerve injury in Guillain-Barrésyndrome(GBS)is becoming clearer,but positivity for multiple antibodies in one case is uncommon.To our knowledge,this is the first case involving GBS with positive anti-sulfatide,anti-GT1a,and anti-GT1b antibodies.CASE SUMMARY A 20-year-old female patient was admitted to the hospital due to weakness of limbs for 5 d,and deterioration of the weakness and muscle aches for 1 d.The patient's limbs were weak,but the tendon reflexes in the part of the limbs were normal.There was no comorbid peripheral nociception or deep sensory dysfunction.She was diagnosed with GBS and was discharged after receiving intravenous human immunoglobulin pulse therapy.CONCLUSION In this article,the clinical manifestations,neurophysiological examination,and auxiliary examination findings of a GBS patient positive for multiple antibodies were analyzed to improve the identification of the disease by clinical physicians at an early stage.
文摘BACKGROUND Miller fisher syndrome(MFS)is a variant of Guillain-Barrésyndrome,an acute immune-mediated peripheral neuropathy that is often secondary to viral infections.Anti-ganglioside antibodies play crucial roles in the development of MFS.The positive rate of ganglioside antibodies is exceptionally high in MFS patients,particularly for anti-GQ1b antibodies.However,the presence of other ganglioside antibodies does not exclude MFS.CASE SUMMARY We present a 56-year-old female patient who suddenly developed right blepharoptosis and progressively worsening vision in both eyes.There were flu symptoms prior to onset,and a coronavirus disease 2019 test was positive.On physical examination,the patient exhibited bilateral extraocular muscle paralysis,weakened reflexes in both limbs,and impaired coordination.The cerebrospinal fluid examination results showed no obvious abnormalities.Bilateral peroneal nerve F-waves were not extracted.Serum anti-GD1b IgG and anti-GT1a IgG antibodies were positive.The patient received intravenous methylprednisolone(1000 mg/day),with the dosage gradually decreased.Additionally,intravenous high-dose immunoglobulin treatment was administered for 5 days(0.4 g/kg/day)from day 2 to day 6 of hospitalization.The patient’s symptoms improved after treatment with immunoglobulins and hormones.CONCLUSION Positive ganglioside antibodies may be used as supporting evidence for the diagnosis;however,the diagnosis of MFS is more reliant on clinical symptoms.
文摘Despite enormous efforts to achieve the goal of eliminating mother-to-child transmission of HIV-1, it remains a major challenge for many countries in sub-Saharan Africa, particularly Mali. Our objective is to assess changes in the rate of mother-to-child transmission of HIV-1. We conducted a cross-sectional study between January 1, 2009 to December 31, 2018 (10 years) of early diagnosis activity in newborns and children born to HIV-1-positive mothers at the National Institute for Public Health (INSP). The samples came from health and referral centers in mali. All samples were received at the Laboratory of Molecular Biology at the INSP. Proviral DNA extraction was performed from a blood spot sample with a Roche DNA kit, Cobas AmpliPrep/Cobas TaqMan HIV-1 qualitative Test, V2.0 (Roche Molecular System, Inc, USA) following the company procedures. Molecular diagnosis was performed using the same kits using an algorithm of three identical PCRs. The Epi Info version 7 software was used for data analysis with a significance threshold of 5%. A total of 10,714 samples of infants and children born to HIV-positive mothers were analyzed by PCR. Ninety-six percent of mothers were on ARV prophylaxis (AZT 3TC NVP and AZT NVP) and 60% of newborns received the same ARV prophylaxis. Of these children, 956 tested positive with an overall transmission rate of 8.92%, varying between 7.27% in 2009 and 08.01% in 2018. This rate was relatively low among children receiving prophylaxis at 2.04% and remained high for children who received breastfeeding at 5.62%. However, the transmission rate remains low for those who have benefited from mixed and artificial breastfeeding at 1.58% and 1.27% respectively. A significant proportion of children remained infected by their mothers during pregnancy, childbirth or breastfeeding. This study shows the importance of early diagnosis of HIV in children using molecular technology.
文摘The emergence of antiretroviral resistance mutations represents a major threat to the achievement of national and global goals for the elimination of HIV-1 infection. The global strategy in 2019 in Cte d'Ivoire is a new national policy for the management of people living with HIV with the administration of dolutegravir (DTG)-based fixed-dose combination. The aim of our study was to evaluate HIV-1 resistance to antiretrovirals (ARVs) in infected adult subjects in Cte d’Ivoire in the context of a systematic switch to a DTG-based combination. Between February 2022 and October 2023, a cross-sectional survey with random sampling was conducted in 06 services caring for people living with HIV. A total of 139 participants were included in the study. Adults with a viral load ≥ 1000 copies/mL were tested for HIV-1 ARV resistance mutations. Molecular analyses were performed using protocol of ANRS-MIE (National Agency for Research on AIDS and emerging infectious diseases). The interpretation is performed by HIVGRAD (https://www.hiv-grade.de/cms/grade/). The frequencies of HIV-1 resistance to non-nucleotide reverse transcriptase inhibitors (NNRTIs), nucleotide reverse transcriptase inhibitors (NRTIs), integrase inhibitors (IINTs) and protease inhibitors (PIs) were 82%, 73%, 19% and 11% respectively. The main mutations observed in the different classes were K103N (45%), M184V (64%), E157Q (19%) and L10V/M46I/A71V/I54V (6%) respectively. This study reveals the emergence of resistance to DTG-based fixed-dose combinations, favored by high rates of resistance to NRTIs and NNRTIs. This finding underlines the need for enhanced viral load monitoring and HIV-1 genotyping tests to guide the choice of NRTIs for combination therapy. In addition, monitoring for mutations to second-generation NRTIs is essential, given the scale-up of DTG-based regimens currently underway in Cte d’Ivoire.
文摘Introduction: HIV, the human immunodeficiency virus, is the etiological agent of acquired immunodeficiency syndrome (AIDS). The aim of this study was to assess the evolution of the viral load in patients under treatment. Methodology: This was a study carried out from July 2017 to June 2022 at the Point G University Hospital laboratory. The determination of the viral load of patients was carried out by PCR on the ABOTT M2000sp/rt platform. Results: A total of 129 patients infected with HIV-1, aged 19 to 72 years with a mean age of 40.05 years ± 10.71;all on antiretroviral chemotherapy. The female gender predominated among our patients. The most common treatment regimen was 2INTI + 1INNTI with 72.9% followed by 2INTI + 1INI with 13.2%. As for the combinations of molecules, the combination TDF + 3TC + EFV and TDF + 3TC + DTG predominated, respectively 65.1% and 13.2%. 89.9% of our patients had undetectable viremia after 12 months of treatment (p < 0.005) with an average viral load which had evolved from 681315.65 copies/ml ± 1616908.484 to M0 at 5742.36 copies /ml ± 35756.883 at M12 (p Conclusion: Generally speaking, antiretroviral treatment had contributed to controlling viral loads, however the therapeutic combination TDF + 3TC + DTG had made it possible to obtain more patients with undetectable viremia instead.
基金National Natural Science Foundation ofChina(30400368)The Natural Science foundation ofBeijing(5072003)Beijing Natural Science foundationProgram and Scientific Research Key Program of BeijingMunicipal commission of Education(KZ20051005001).
文摘To prepare HIV-1 Vif and hAPOBEC3G and to produce their antibodies, the full length gene fragment of HIV-1 vif was amplified by PCR from a plasmid of HIV-1 NL4.3 cDNA, and the APOBEC3G gene was obtained by RT-PCR from the total RNA of H9 cells. The resulting DNA construct was cloned into a prokaryotic expression vector (pET-32a). Recombinant pET-vif and pET-APOBEC3G were expressed respectively in Eserichia coli BL21 (DE3) as an insoluble protein. The vector also contained a six-histidine tag at the C-terminus for convenient purification and detection. To express and purify the HIV-1 Vif and hAPOBEC3G in E.coli cells, the accuracy of inserted gene and specificity of proteins were detected by the two enzyme digestion method, SDS-PAGE, and Western blotting. Rabbits were then immunized by Vif or APOBEC3G protein and serum samples were tested by indirect ELISA to determine the level of antibodies. Immunoenzyme and immunofluorescence assays were performed to identify the specificity of polyclonal antibodies. The titer of the anti-Vif antibodies was 1:204800, and that of the anti-APOBEC3G antibodies was 1:102400. Thus the antibodies could detect the antigen expression in the cells, demonstrating that fusion proteins with high purity and their corresponding polyclonal antibodies with high titer and specificity were achieved.
基金the National Natural Science Foundation of China (No. 30000070)
文摘The cytokine repertoire of ADP/ATP carrier-specific humoral immune responses and the cytokine-dependent anti-ADP/ATP carrier antibody IgG subclasses were examined in a cohort of ADP/ATP carrier-immunized BALB/c mice treated with anti-CD4 monoclonal antibody. Eighteen male BALB/c mice （6–8 weeks old） were randomized into 3 groups： dilated cardiomyopathy （DCM） group, DCM-tolerance （Tol） group and control group. The mice in DCM group were immunized with the peptides derived from human ADP/ATP carrier protein for 6 months and mice in the control group were sham-immunized, while the mice in DCM-Tol group were immunized with ADP/ATP carrier protein and anti-CD4 McAb simultaneously. Serum autoantibody against ADP/ATP carrier and IgG subclasses were measured by ELISA, intracellular cytokines IFN-γ and IL-4 of Th cells were moni- tored with flow cytometry, and splenic T cell cytokines IFN-γ, IL-2, IL-4 and IL-6 were detected by using real-time fluorescent quantitative PCR. The results showed that the autoantibody against ADP/ATP carrier was found in all mice in DCM group, and the antibody level, serum IgG1 and IgG2a subclasses, cytokines in T cells and Th cells were all elevated in DCM group, as compared with those in control group （P〈0.01）. On the other hand, in DCM-Tol group, the autoantibody level and contents of all the cytokines were significantly different from those in DCM group （P〈0.01）, and were close to those in control group. And the levels of IgG1, IgG2a, IgG2b and IgG3 were influenced, to varying degrees, by anti-CD4 McAb as compared with those in DCM group. All these four types of IgG subclasses were substantially decreased in DCM-Tol group as compared with DCM group. It is concluded that the treatment with anti-CD4 McAb could prevent the activation of T cells, reverse the abnormal secretion of cytokines and the imbalance between Th1/Th2 cell subsets and abnormal production of autoantibody against ADP/ATP carrier, and eventually avoid myocardial injuries.
文摘Simpli RED HIV-1 antibody test was used to detect 17,378 samples from people of entering coutry. 10 samples were positive. Using Western Blot as the cofirmatory test, 8/10 was positive. The advantages of this test was very sensitive, quick and easy. 20 ul whole blood could be used as sample and got the results in two minutes.
