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Background: A number of international research groups have developed DNA quantitation assays in order to investigate the role of mitochondrial DNA depletion in anti-retroviral therapy-induced toxicities. Objectives: A collaborative study was undertaken to evaluate intra-assay precision and between laboratory concordance of measurements of mitochondrial DNA quantity, as a component of a comprehensive quality assurance project. Study design: Four laboratories were asked to measure and report mitochondrial DNA and nuclear DNA genome copy number, as well as mitochondrial DNA copy number/cell, for 17 coded aliquots of DNA derived from serial dilutions of pooled DNA from a lymphoblastoid cell line. Samples included masked replicates and five standards. All samples had similar mitochondrial DNA/nuclear DNA ratios. Precision within laboratories was assessed by determining the coefficient of variation of replicates. Concordance between laboratories was assessed by determining the average coefficient of variation of the mean replicate values for each sample. The effect of standardising the assay for these three measurements was also assessed for laboratories A, B and C. Results: Measurements of mitochondrial DNA and nuclear DNA content for replicate samples varied by an average of less than 6% (based on log10 values, 72% non-logged values), and measurements of mitochondrial DNA/cell for replicates varied by less than 12% (based on log10 values, 32% non-logged values), with no improvement of precision after standardisation. Standardisation did significantly improve the concordance of results for measurements of mitochondrial DNA content and mitochondrial DNA/cell. Non-standardised measurements of mitochondrial DNA content for the same sample set varied by 19% between laboratories (based on log10 values, 96% non-logged values), and after standardisation results varied by less than 3% (based on log10 values, 54% non-logged values). There was no significant improvement for concor

Background: A number of international research groups have developed DNA quantitation assays in order to investigate the role of mitochondrial DNA depletion in anti-retroviral therapy-induced toxicities. Objectives: A collaborative study was undertaken to evaluate intra-assay precision and between laboratory concordance of measurements of mitochondrial DNA quantity, as a component of a comprehensive quality assurance project. Study design: Four laboratories were asked to measure and report mitochondrial DNA and nuclear DNA genome copy number, as well as mitochondrial DNA copy number/cell, for 17 coded aliquots of DNA derived from serial dilutions of pooled DNA from a lymphoblastoid cell line. Samples included masked replicates and five standards. All samples had similar mitochondrial DNA/nuclear DNA ratios. Precision within laboratories was assessed by determining the coefficient of variation of replicates. Concordance between laboratories was assessed by determining the average coefficient of variation of the mean replicate values for each sample. The effect of standardising the assay for these three measurements was also assessed for laboratories A, B and C. Results: Measurements of mitochondrial DNA and nuclear DNA content for replicate samples varied by an average of less than 6% (based on log10 values, 72% non-logged values), and measurements of mitochondrial DNA/cell for replicates varied by less than 12% (based on log10 values, 32% non-logged values), with no improvement of precision after standardisation. Standardisation did significantly improve the concordance of results for measurements of mitochondrial DNA content and mitochondrial DNA/cell. Non-standardised measurements of mitochondrial DNA content for the same sample set varied by 19% between laboratories (based on log10 values, 96% non-logged values), and after standardisation results varied by less than 3% (based on log10 values, 54% non-logged values). There was no significant improvement for concor

Background The use of abacavir a potent HIV—1 nucleosideanalogue reverse—transcriptase inhibitor is complicated by a potentially life-threatening hypersensitivity syndrome in about 5% of cases. Genetic factors influencing the immune response to abacavir might confer susceptibility. We aimed to find associations between MHC alleles and abacavir hypersensitivity in HIV—1-positive individuals treated with abacavir. Methods MHC region typing was done in the first 200 Western Australian HIV Cohort Study participants exposed to abacavir. Definite abacavir hypersensitivity was identified in 18 cases, and was excluded in 167 individuals with more than 6 weeks' exposure to the drug (abacavir tolerant). 15 individuals experienced some symptoms but did not meet criteria for abacavir hypersensitivity. p values were corrected for comparisons of multiple HLA alleles (pc) by multiplication of the raw p value by the estimated number of HLA alleles present within the loci examined. Findings HLA-B*5701 was present in 14 (78%) of the 18 patients with abacavir hypersensitivity, and in four (2%) of the 167 abacavir tolerant patients (odds ratio 117 [95% CI 29—481], pc<0·0001), and the HLA-DR7 and HLA-DQ3 combination was found in 13 (72%) of hypersensitive and five (3%) of tolerant patients (73 [20—268], pc<0·0001). HLA-B*5701, HLA-DR7, and HLA-DQ3 were present in combination in 13 (72%) hypersensitive patients and none of the tolerant patients (822 [43—15 675], pc<0·0001). Other MHC markers also present on the 57·1 ancestral haplotype to which the three markers above belong confirmed the presence of haplotype-specific linkage disequilibrium, and mapped potential susceptibility loci to a region bounded by C4A6 and HLA-C. Within the entire abacavir-exposed cohort (n=200), presence of HLA-B*5701, HLA-DR7, and HLA-DQ3 had a positive predictive value for hypersensitivity of 100%, and a negative predictive value of 97%. Interpretation Genetic susceptibility to abacavir hypersensitivity is car

Background The use of abacavir a potent HIV—1 nucleosideanalogue reverse—transcriptase inhibitor is complicated by a potentially life-threatening hypersensitivity syndrome in about 5% of cases. Genetic factors influencing the immune response to abacavir might confer susceptibility. We aimed to find associations between MHC alleles and abacavir hypersensitivity in HIV—1-positive individuals treated with abacavir. Methods MHC region typing was done in the first 200 Western Australian HIV Cohort Study participants exposed to abacavir. Definite abacavir hypersensitivity was identified in 18 cases, and was excluded in 167 individuals with more than 6 weeks' exposure to the drug (abacavir tolerant). 15 individuals experienced some symptoms but did not meet criteria for abacavir hypersensitivity. p values were corrected for comparisons of multiple HLA alleles (pc) by multiplication of the raw p value by the estimated number of HLA alleles present within the loci examined. Findings HLA-B*5701 was present in 14 (78%) of the 18 patients with abacavir hypersensitivity, and in four (2%) of the 167 abacavir tolerant patients (odds ratio 117 [95% CI 29—481], pc<0·0001), and the HLA-DR7 and HLA-DQ3 combination was found in 13 (72%) of hypersensitive and five (3%) of tolerant patients (73 [20—268], pc<0·0001). HLA-B*5701, HLA-DR7, and HLA-DQ3 were present in combination in 13 (72%) hypersensitive patients and none of the tolerant patients (822 [43—15 675], pc<0·0001). Other MHC markers also present on the 57·1 ancestral haplotype to which the three markers above belong confirmed the presence of haplotype-specific linkage disequilibrium, and mapped potential susceptibility loci to a region bounded by C4A6 and HLA-C. Within the entire abacavir-exposed cohort (n=200), presence of HLA-B*5701, HLA-DR7, and HLA-DQ3 had a positive predictive value for hypersensitivity of 100%, and a negative predictive value of 97%. Interpretation Genetic susceptibility to abacavir hypersensitivity is car