Background Recombinant erythropoietin(rEPO)is commonly used in therapy but may be abused in sports to enhance endurance.In doping analysis,rEPO can be detected in human urine or blood samples at picogram(pg)levels bas...Background Recombinant erythropoietin(rEPO)is commonly used in therapy but may be abused in sports to enhance endurance.In doping analysis,rEPO can be detected in human urine or blood samples at picogram(pg)levels based on its slightly higher molecular weight(MW)than that of endogenous EPO using western blotting(WB).However,a type of variant erythropoietin(VAR-EPO)encoded by the EPO c.577del variant has a similar MW to rEPO,and these 2 molecules cannot be distinguished using conventional analytical methods.A fit-for-purpose method needs to be developed immediately.Methods In this study,we introduced a reverse–normal immunopurification technique for sample pretreatment to remove VAR-EPO from samples to eliminate its interference with rEPO detection.Firstly,a rabbit monoclonal antibody(mAb)that can specifically recognize trace amounts of VAR-EPO with high affinity was generated.Then,using this antibody to enrich VAR-EPO,we developed reverse–normal immunopurification coupled with WB on the purpose of analyzing rEPO in urine and serum samples.Next,the method was fully validated and evaluated using blank samples,spiked samples and rEPO excreted samples.Finally,the identification criteria of rEPO was established.Results A specific anti-VAR mAb with high affinity was developed.Using it,we developed the doping analytical method for rEPO.Our method effectively detects and removes VAR-EPO,enabling accurate rEPO detection.Conclusion A method has already been applied for rEPO confirmation in routine doping analyses.展开更多
Drug sensitivity testing to establish resistance to TB drugs takes many months to arrive at. Public health physicians have difficulties with such an approach due to long wait periods and cannot use it to establish com...Drug sensitivity testing to establish resistance to TB drugs takes many months to arrive at. Public health physicians have difficulties with such an approach due to long wait periods and cannot use it to establish community wide prevalence as a way to understand where resistance may be emerging faster and to limit its spread. The objective of this study was to use the dot-blot hybridization technique in the detection of resistance to rifamycin (RIF) and streptomycin (SM) in South- Western Cameroon and to compare the technique with the routine culture and drug susceptibility testing for detecting resistance in a resource poor country, Cameroon. A hospital-based study was conducted at the Regional hospitals of Buea and Limbe and Tiko Central Clinic. Tuberculosis (TB) patients aged 15 to 50 (mean age: 30.50 ± 8.33 standard deviation) were recruited for the study between December 2006 and April 2007. Cultures from 59 patients were tested for rifampicin and streptomycin sensitivity by the modified proportion method and mutational analysis for rpoB codon 516 and rrs codon 513 was performed by the dot-blot hybridization technique. Of the 59 sputum samples collected (36 were males and 23 were females) came from Buea 19 (32.2%), Limbe 20 (33.9%) and Tiko 20 (33.9%) towns respectively. Amplification for the gene showed that there was (59) 100% amplification with primers used for rpoB genes and 43 (72.9%) amplification with primers used for the rrs gene. Mutational analysis demonstrated that resistance to RIF was common in females (52.1%) than males (41.7%) while 6% of the samples were indeterminate. 12 (20.3%) samples showed phenotypic and genotypic resistance to RIF compared to 34 samples (58.1%) for SM. Phenotypic resistance and genotypic susceptibility were found in 5 (8.5%) RIF and 3 (4.7%) SM compared to phenotypic susceptibility and genotypic resistance that were found in 2 (3.5%) RIF and 3(4.7%) SM. Double mutation on rpoB and rrs genes occurred in 8 (13.6%) DNA samples. Resistance to RIF and SM due to mutations on the rpoB and rrs genes respectively in the SW region was found to be high and comparable to the drug susceptibility testing by 92%, (95% CI: 75.7 - 99.1). The Dot-blot technique will be useful in rapidly assessing the effectiveness of national TB control programs in limiting the spread of resistance strains in Cameroon.展开更多
Objective To probe into the pathogenic mechanisms of paraneoplastic limbic encephali- tis( PL E) in patients with small cell lung carcinoma( SCLC) .Methods The indirect im- munoperoxidase method and Western blotanalys...Objective To probe into the pathogenic mechanisms of paraneoplastic limbic encephali- tis( PL E) in patients with small cell lung carcinoma( SCLC) .Methods The indirect im- munoperoxidase method and Western blotanalysis were used for detecting anti- Hu antibodies in1 6 PLE patients associated with SCL C.Autopsy and pathological study were performed on two cases.Results Eight patients( 5 0 % ) had anti- Hu antibodies( anti- Hu+) whereas eight patients ( 5 0 % ) no detectable antineuronal antibodies ( anti- Hu- ) .The clinical and laboratory features of PLE and time to diagnosis of SCLC were similar in the anti- Hu+and anti- Hu- groups.Involvement of other areas of the nervous system in seven( 87.5 % ) patients of the anti- Hu+group but in only one ( 1 2 .5 ) % of the anti- Hu- group ( P=0 .0 1 2 ) . Conclusions The presence of this characteristic neurological disorder strongly suggested that with an ac- companying SCLC,existence of anti- Hu autoantibody was in favour of an autoimmune mech- anism participating in PLE.展开更多
基金WADA and Beijing Sport University for funding under grant numbers 22B06XZ and 2022YB011
文摘Background Recombinant erythropoietin(rEPO)is commonly used in therapy but may be abused in sports to enhance endurance.In doping analysis,rEPO can be detected in human urine or blood samples at picogram(pg)levels based on its slightly higher molecular weight(MW)than that of endogenous EPO using western blotting(WB).However,a type of variant erythropoietin(VAR-EPO)encoded by the EPO c.577del variant has a similar MW to rEPO,and these 2 molecules cannot be distinguished using conventional analytical methods.A fit-for-purpose method needs to be developed immediately.Methods In this study,we introduced a reverse–normal immunopurification technique for sample pretreatment to remove VAR-EPO from samples to eliminate its interference with rEPO detection.Firstly,a rabbit monoclonal antibody(mAb)that can specifically recognize trace amounts of VAR-EPO with high affinity was generated.Then,using this antibody to enrich VAR-EPO,we developed reverse–normal immunopurification coupled with WB on the purpose of analyzing rEPO in urine and serum samples.Next,the method was fully validated and evaluated using blank samples,spiked samples and rEPO excreted samples.Finally,the identification criteria of rEPO was established.Results A specific anti-VAR mAb with high affinity was developed.Using it,we developed the doping analytical method for rEPO.Our method effectively detects and removes VAR-EPO,enabling accurate rEPO detection.Conclusion A method has already been applied for rEPO confirmation in routine doping analyses.
文摘Drug sensitivity testing to establish resistance to TB drugs takes many months to arrive at. Public health physicians have difficulties with such an approach due to long wait periods and cannot use it to establish community wide prevalence as a way to understand where resistance may be emerging faster and to limit its spread. The objective of this study was to use the dot-blot hybridization technique in the detection of resistance to rifamycin (RIF) and streptomycin (SM) in South- Western Cameroon and to compare the technique with the routine culture and drug susceptibility testing for detecting resistance in a resource poor country, Cameroon. A hospital-based study was conducted at the Regional hospitals of Buea and Limbe and Tiko Central Clinic. Tuberculosis (TB) patients aged 15 to 50 (mean age: 30.50 ± 8.33 standard deviation) were recruited for the study between December 2006 and April 2007. Cultures from 59 patients were tested for rifampicin and streptomycin sensitivity by the modified proportion method and mutational analysis for rpoB codon 516 and rrs codon 513 was performed by the dot-blot hybridization technique. Of the 59 sputum samples collected (36 were males and 23 were females) came from Buea 19 (32.2%), Limbe 20 (33.9%) and Tiko 20 (33.9%) towns respectively. Amplification for the gene showed that there was (59) 100% amplification with primers used for rpoB genes and 43 (72.9%) amplification with primers used for the rrs gene. Mutational analysis demonstrated that resistance to RIF was common in females (52.1%) than males (41.7%) while 6% of the samples were indeterminate. 12 (20.3%) samples showed phenotypic and genotypic resistance to RIF compared to 34 samples (58.1%) for SM. Phenotypic resistance and genotypic susceptibility were found in 5 (8.5%) RIF and 3 (4.7%) SM compared to phenotypic susceptibility and genotypic resistance that were found in 2 (3.5%) RIF and 3(4.7%) SM. Double mutation on rpoB and rrs genes occurred in 8 (13.6%) DNA samples. Resistance to RIF and SM due to mutations on the rpoB and rrs genes respectively in the SW region was found to be high and comparable to the drug susceptibility testing by 92%, (95% CI: 75.7 - 99.1). The Dot-blot technique will be useful in rapidly assessing the effectiveness of national TB control programs in limiting the spread of resistance strains in Cameroon.
文摘Objective To probe into the pathogenic mechanisms of paraneoplastic limbic encephali- tis( PL E) in patients with small cell lung carcinoma( SCLC) .Methods The indirect im- munoperoxidase method and Western blotanalysis were used for detecting anti- Hu antibodies in1 6 PLE patients associated with SCL C.Autopsy and pathological study were performed on two cases.Results Eight patients( 5 0 % ) had anti- Hu antibodies( anti- Hu+) whereas eight patients ( 5 0 % ) no detectable antineuronal antibodies ( anti- Hu- ) .The clinical and laboratory features of PLE and time to diagnosis of SCLC were similar in the anti- Hu+and anti- Hu- groups.Involvement of other areas of the nervous system in seven( 87.5 % ) patients of the anti- Hu+group but in only one ( 1 2 .5 ) % of the anti- Hu- group ( P=0 .0 1 2 ) . Conclusions The presence of this characteristic neurological disorder strongly suggested that with an ac- companying SCLC,existence of anti- Hu autoantibody was in favour of an autoimmune mech- anism participating in PLE.
