9.Optimization of DNA purification nano beads specifically for FFPE samples
Lin XIAO ; Xiaofang LÜ ; Shiyu CAO ; Dan JIANG ; Linyong SUN ; Kun LI ; Feng YE
Chinese Journal of Clinical and Experimental Pathology 2024;40(7):725-730
Purpose Focusing on the characteristics of for-malin fixed paraffin embedded(FFPE)samples,explored nano-magnetic bead nucleic acid extraction solutions with higher qual-ity/yield and continued to improve molecular pathology technolo-gy.Methods Alternative magnetic beads were synthesised in four major categories and 15 sub-categories and we screened to obtain high-quality/yield magnetic beads centred on FFPE samples.Simulated conventional tissues,simulated coarse needle punctures(liver),and simulated fiberoptic bronchoscopy sam-ples(lungs)were sectioned with the same number of serial slices in tubes.The nucleic acids of slices were extracted using the best magnetic beads screened in this study and common com-mercially available kits,and then perform comparison and purifi-cation quality parameters such as total amount and fragment size.The downstream applications of nucleic acids were validated by PCR and Sanger sequencing.Results Screening all homemade nanomagnetic beads centered on the DNA of FFPE samples,the total recoveries of the best performance nanomagnetic beads were obtained to be 58.5%±1.58%,and the total recoveries of five commercially available commercial magnetic beads and three do-mestic kit magnetic beads ranged from 18.68%to 40.71%.The total amount of DNA(ng)extracted from the same amount of tis-sue(serial slices),the nucleic acid yield of this study in simu-lated conventional tissues,simulated coarse needle punctures,and simulated fiberoptic bronchoscopy samples were increased by 39.49%-181.72%compared with those of the commercially a-vailable kits(P<0.05).The total amount of extracted nucleic acid from simulated fiberoptic bronchoscopy tissue sections can be more than 100 ng for 1 slice(4 μm)and more than 400 ng for 5 slices.Conclusion The DNA purification nanomagnetic beads screened with DNA from FFPE samples have a significant enhancement comparing to the existing commercial bead proto-cols,and provide space for quality assurance,automated testing,and program expansion for clinical molecular pathology testing.
10.Clinical and pathological characteristics and prognostic analysis of large B-cell lymphoma with IRF4 rearrangement
Shi HE ; Chengyu WU ; Shuyi LU ; Longfeng KE ; Yating QIU ; Wenwen ZHANG ; Gang CHEN ; Yanping CHEN
Chinese Journal of Clinical and Experimental Pathology 2024;40(7):730-735
Purpose To explore the clinicopathological features,diagnosis and differential diagnosis of large B-cell lym-phoma with IRF4 rearrangement(LBCL-IRF4r).Methods Clinical data of 8 cases of LBCL-IRF4r were collect,hematoxy-lin-eosin and immunohistochemical of EnVision two-step stains,in situ hybridization and FISH was used to study the histology,immunotypes and molecular genetic characteristics.The rele-vant literatures were reviewed.Results Among 8 cases of LBCL-IRF4r,the male to female ratio was 1.67:1,with age range 10-53 years(mean 25.8 years).Five cases occurred in tonsils,2 cases in nasopharynx and 1 cases in inguinal lymph node.Microscopically,the tumors presented with a purely follic-ular,purely diffuse or a combined follicular and diffuse architec-ture.The tumor cells were typical centroblasts and less frequent-ly medium-sized blastic cells with smaller nucleoli,apoptosis and nuclear fragmentation were easily seen.Immunophenotypi-cally,the tumor cells of the eight cases of LBCL-IRF4r diffuse strongly expressed CD20(8/8),PAX5(2/2),CD79a(3/3),BCL6(8/8)and MUM-1(8/8),mostly expressed CD10(7/8),partially expressed BCL2(5/8)and CD5(4/8),and did not express Cyclin D1,CD23 and CD30.The percentage of Ki67 index ranged from 70%to 95%.EBER in situ hybridiza-tion was negative in all cases.IRF4 rearrangements were detec-ted in all cases(8/8).BCL6 rearrangements were detected in one case(1/2).MYC(0/4)and BCL2(0/3)rearrangements were not detected in all cases.Conclusion LBCL-IRF4r is more common in children and adolescents with characteristic IG::IRF4 rearrangement and a good prognosis,which needs to be differentiated from other types of large B-cell lymphoma.

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