- Email: [email protected]
Molecular tissue typing in renal transplantation: A single centre report
Abstracts
S91
3 102
MOLECULAR TISSUE TYPING IN RENAL TRANSPLANTATION: A SINGLE CENTRE REPORT Mahendra N. Mishra, Haresh Mani, Rabindra N. Misra, Vinod K. Saxena, Rajshri Ramsetu, Pathology, INHS Asvini, Mumbai, Maharashtra, India; Urology, INHS Asvini, Mumbai, Maharashtra, India; Nephrology, INHS Asvini, Mumbai, Maharashtra, India Background: The last 5 years in India have witnessed an increase in live unrelated donor (LURD) transplantation and a gradual switch over to DNA based tissue. Material and Methods: At the outset available data of tissue typing for HLA – A, B and DR alleles pertaining to 200 samples by serology and only DR typing by SSP for 70 samples were studied . In a prospective study at our center HLA typing was done by sequence specific primers (SSP) for 60 samples out of which 30 were typed by microlymphocytoxocity also. The patients were followed up for one year. Results: A discrepancy of 16 –35 % was observed between serology and SSP. The donor profile showed female preponderance and over 50 % were LURD. Fifteen patients were suspected to have clinical acute allograft rejection. The one-year graft survival was 87 %. Discussion: SSP was useful in defining alleles at private level (A*68, A*66, A*33-3 cases, A*69) in six patients. The blank alleles in serology which were identified by SSP included A* 36 in 2 related samples was, A* 74 and A* 03 in one sample each. Three alleles that were not identified may have been due to amplification failure or homozygousity. In this study neither the graft neither survival nor the AR episodes showed any correlation with the extent of HLA mismatch. The limitation of this study is the small sample size and non-availability of better matched donors. Conclusions: In our opinion SSP based HLA typing is very well suited for laboratories with workload of up to 500 samples per year.
3 103
WE IMPROVE OUR RESULTS: AFTER A CRITICAL ANALISYS OF THE QUALITY FROM THE EXTRACTED DNA Moramay Z.P. Ikeda, Sandra M. Ferreira, Carolina Kneib, Helena B. Cazarote, Carlos F. Alves, Gisele C. Possebon, Juliana P. Camargo, Ivy N. Vieira, Luciane Kaminski, Maria-Madalena Okoinski, Claudia H. Takimura, Vanessa S. Paladino, Taiana B. Bertozzi, Vania C.F. Bandeira, Cleide Santos, Roberta G. Amorim, Leonardo M. Ferreira, Rodrigo O.S. Ribeiro, Renata von Glehn-Ponsirenas, Cristina Q.C. von Glehn, Immunogenetics Laboratory, Cajuru University Hospital / Health Allinance / PUCPR, Curitiba, Parana, Brazil The purpose of this work was using the HLA-A, B and DRB1 typing results to evaluate the quality of the extracted DNA. The DNA extraction and the HLA typing were done by commercial kits. We used agarose gel (2.5%) stained with EB. Our professional staff analyzed the final results. The retrospective analysis of the DNA samples quality was done by spectrophotometry. The final HLA typing results of the 700 DNA samples had been analyzed: 78% (550) of these samples had satisfactory typing results; 22% (150) of these samples had unsatisfactory results. Among these samples 23% (34) didn’t have DNA amplification and in 77% (116) the unsatisfactory results was caused by: lack of positive bands, lack of the internal control bands, problems with the gel, etc. Conclusion: Our results were improved by: 1) The identification of the DNAs with low concentration and high impurities and their correlation with blood samples badly conditioned. 2) The introduction of a centrifugal machine for the trays. 3) The confection of one “book” with the photos of unsatisfactory results to help the new technicians in the fast detection of the main problems.
S91
3 102
MOLECULAR TISSUE TYPING IN RENAL TRANSPLANTATION: A SINGLE CENTRE REPORT Mahendra N. Mishra, Haresh Mani, Rabindra N. Misra, Vinod K. Saxena, Rajshri Ramsetu, Pathology, INHS Asvini, Mumbai, Maharashtra, India; Urology, INHS Asvini, Mumbai, Maharashtra, India; Nephrology, INHS Asvini, Mumbai, Maharashtra, India Background: The last 5 years in India have witnessed an increase in live unrelated donor (LURD) transplantation and a gradual switch over to DNA based tissue. Material and Methods: At the outset available data of tissue typing for HLA – A, B and DR alleles pertaining to 200 samples by serology and only DR typing by SSP for 70 samples were studied . In a prospective study at our center HLA typing was done by sequence specific primers (SSP) for 60 samples out of which 30 were typed by microlymphocytoxocity also. The patients were followed up for one year. Results: A discrepancy of 16 –35 % was observed between serology and SSP. The donor profile showed female preponderance and over 50 % were LURD. Fifteen patients were suspected to have clinical acute allograft rejection. The one-year graft survival was 87 %. Discussion: SSP was useful in defining alleles at private level (A*68, A*66, A*33-3 cases, A*69) in six patients. The blank alleles in serology which were identified by SSP included A* 36 in 2 related samples was, A* 74 and A* 03 in one sample each. Three alleles that were not identified may have been due to amplification failure or homozygousity. In this study neither the graft neither survival nor the AR episodes showed any correlation with the extent of HLA mismatch. The limitation of this study is the small sample size and non-availability of better matched donors. Conclusions: In our opinion SSP based HLA typing is very well suited for laboratories with workload of up to 500 samples per year.
3 103
WE IMPROVE OUR RESULTS: AFTER A CRITICAL ANALISYS OF THE QUALITY FROM THE EXTRACTED DNA Moramay Z.P. Ikeda, Sandra M. Ferreira, Carolina Kneib, Helena B. Cazarote, Carlos F. Alves, Gisele C. Possebon, Juliana P. Camargo, Ivy N. Vieira, Luciane Kaminski, Maria-Madalena Okoinski, Claudia H. Takimura, Vanessa S. Paladino, Taiana B. Bertozzi, Vania C.F. Bandeira, Cleide Santos, Roberta G. Amorim, Leonardo M. Ferreira, Rodrigo O.S. Ribeiro, Renata von Glehn-Ponsirenas, Cristina Q.C. von Glehn, Immunogenetics Laboratory, Cajuru University Hospital / Health Allinance / PUCPR, Curitiba, Parana, Brazil The purpose of this work was using the HLA-A, B and DRB1 typing results to evaluate the quality of the extracted DNA. The DNA extraction and the HLA typing were done by commercial kits. We used agarose gel (2.5%) stained with EB. Our professional staff analyzed the final results. The retrospective analysis of the DNA samples quality was done by spectrophotometry. The final HLA typing results of the 700 DNA samples had been analyzed: 78% (550) of these samples had satisfactory typing results; 22% (150) of these samples had unsatisfactory results. Among these samples 23% (34) didn’t have DNA amplification and in 77% (116) the unsatisfactory results was caused by: lack of positive bands, lack of the internal control bands, problems with the gel, etc. Conclusion: Our results were improved by: 1) The identification of the DNAs with low concentration and high impurities and their correlation with blood samples badly conditioned. 2) The introduction of a centrifugal machine for the trays. 3) The confection of one “book” with the photos of unsatisfactory results to help the new technicians in the fast detection of the main problems.