COPAN eNAT with FLOQSwabs: A Novel Collection Kit for ...
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COPAN eNAT with FLOQSwabs: A Novel Collection Kit for Genetic Testing Zhijing Huang, PhD, Safedin Sajo Beqaj PhD, HCLD, CC, (ABB) Patients Choice Laboratories, Indianapolis, USA MATERIALS AND METHODS Ø Buccal swab samples were co-collected using COPAN FLOQSwabs TM with 1ml eNAT TM medium and the Epicentre Catch-All TM Sample Collection Swab (Fig 1). Both swabs were placed next to each other and collected at the same time. The FLOQSwabs TM can be broken at the red breakpoint and can be secured onto the eNAT TM medium cap. Ø Copan recommends that FLOQSwabs TM are always used alone for buccal sample collection. Ø Genomic DNA were extracted in parallel from samples collected with both collection devices using Thermo Fisher MagMAX™ DNA Multi-Sample Ultra Kit. About 600ul eNAT TM medium left while the entire Epicentre swab was used. Ø gDNA from both samples was tested on the current comprehensive panel at Patients Choice Laboratories with Applied Biosystems 7900HT Fast Real-Time PCR System (Fig. 2) and the results were compared : Ø Pharmacogenetics: CYP2D6, CYP2C9, CYP2C19, CYP3A4, CYP3A5, COMT, OPRM1, SLCO1B1, ANKK1 Ø Coagulation Factors: FV, FII, MTHFR mutation Ø APOE genotyping INTRODUCTION Buccal swab has been a convenient collection device for clinical diagnosis. Current practice involves a dry swab that collects DNA from the cells on the inside of a person’s cheek. Although effective, dry swabs sometimes cannot yield sufficient nucleic acid and require re-extraction or recollection. Dry swabs also collect microbial debris which propagate during storage and interfere with assay performance. The goal of the molecular diagnostic community is to find a collection device that can give quality nucleic acid with high yield, and require minimal steps for downstream molecular applications. The New collection system Human DNA-free FLOQSwabs™ and eNAT TM medium from COPAN are specifically designed for genetic testing might be a solution. The material and design of FLOQSwabs TM allows to recover more cells and efficiently release samples into liquid medium. The eNAT TM medium is capable of preserving the DNA while inhibiting bacterial growth. Moreover, rapid release of cells into the eNAT TM medium enables immediate preservation of patient samples, allowing repeats and additional tests performed with the original sample and facilitating automated tests. Therefore this collection system aims to conveniently acquire a high yield of patient sample with minimal contamination. RESULTS Procedures for preservation and transportation Buccal swab samples were co-collected using COPAN FLOQSwabs TM with 1ml eNAT TM medium and the Epicentre Catch-All Sample Collection Swab. Both swabs were placed next to each other and collected at the same time. After collection, FLOQSwab TM was broken and preserved in the eNAT TM medium tube immediately. Epicentre Catch-All samples needs to be refrigerated if not processed immediately while FLOQSwabs TM with eNAT TM medium can be stored at room temperature without compromising sample quality. During extraction, a 400mL portion of eNAT TM medium was transferred directly for process while the Epicenter swab needed to be cut and transferred into a new tube for pretreatment. More than half of the eNAT TM medium including the FLOQSwab TM were left for future applications. SNP genotyping assays Genomic DNA were extracted using Thermo Fisher MagMAX™ DNA Multi-Sample Ultra Kit. Customized PGx panels were run using 7900HT Fast Real-Time PCR System on the extracted gDNA. Data were analyzed with TaqMan® Genotyper Software. Results generated with eNAT TM -extracted DNA showed good amplification and confident calls (Fig 3). We compared the genotyping results with Epicenter swab-extracted DNA and eNAT TM -extracted DNA, all 47 assays of 17 genes showed 100% agreement (Table 1) . Fig 3. Screenshot of TaqMan® Genotyper Software analyzing genotyping data from both sample types. Table 1. Agreement of all 47 assays of 17 genes in the PGx panel from both sample types. Epicentre Catch-All TM CONCLUSIONS COPAN FLOQSwabs TM with 1ml eNAT TM medium kit is a very convenient device for buccal sample collection, storage and transportation for nucleic acids preservation. Direct lysis in the eNAT TM medium tube inactivates bacterial proliferation, reduces sample processing steps and facilitates automation. The gDNA yielded from a 400ul portion of eNAT TM medium is pure and adequate for genetic testing. Moreover, the leftover eNAT TM medium can be stored for additional and repeat confirmatory testing if needed, avoiding sample recollection. Fig 1. Picture of FLOQSwabs TM with 1ml eNAT TM medium and the Epicentre Catch-All TM Sample Collection Swab Fig 2. 7900HT Fast Real-Time PCR System FLOQSwabs™ and eNAT™ Gene ANKK1 APOE COMT CYP1A2 CYP2B6 CYP2C19 CYP2C9 CYP2D6 CYP3A4 Agreement 100% 100% 100% 100% 100% 100% 100% 100% 100% Gene CYP3A5 FII FV GRIK4 MTHFR OPRM1 SLCO1B1 VKORC1 Agreement 100% 100% 100% 100% 100% 100% 100% 100%
Transcript of COPAN eNAT with FLOQSwabs: A Novel Collection Kit for ...
COPAN eNAT with FLOQSwabs: A Novel Collection Kit for Genetic Testing
Zhijing Huang, PhD, Safedin Sajo Beqaj PhD, HCLD, CC, (ABB)Patients Choice Laboratories, Indianapolis, USA
MATERIALS AND METHODSØ Buccal swab samples were co-collected using COPAN
FLOQSwabsTM with 1ml eNATTM medium and the EpicentreCatch-AllTM Sample Collection Swab (Fig 1). Both swabs were placed next to each other and collected at the same time. The FLOQSwabsTM can be broken at the red breakpoint and can be secured onto the eNATTM medium cap.
Ø Copan recommends that FLOQSwabsTM are always used alone for buccal sample collection.
Ø Genomic DNA were extracted in parallel from samples collected with both collection devices using Thermo Fisher MagMAX™ DNA Multi-Sample Ultra Kit. About 600ul eNATTM medium left while the entire Epicentre swab was used.
Ø gDNA from both samples was tested on the current comprehensive panel at Patients Choice Laboratories with Applied Biosystems 7900HT Fast Real-Time PCR System (Fig. 2) and the results were compared :
Ø Pharmacogenetics: CYP2D6, CYP2C9, CYP2C19, CYP3A4, CYP3A5, COMT, OPRM1, SLCO1B1, ANKK1
Ø Coagulation Factors: FV, FII, MTHFR mutationØ APOE genotyping
INTRODUCTIONBuccal swab has been a convenient collection device for clinical diagnosis. Current practice involves a dry swab that collects DNA from the cells on the inside of a person’s cheek. Although effective, dry swabs sometimes cannot yield sufficient nucleic acid and require re-extraction or recollection. Dry swabs also collect microbial debris which propagate during storage and interfere with assay performance. The goal of the molecular diagnostic community is to find a collection device that can give quality nucleic acid with high yield, and require minimal steps for downstream molecular applications. The New collection system Human DNA-free FLOQSwabs™ and eNATTM medium from COPAN are specifically designed for genetic testing might be a solution. The material and design of FLOQSwabsTM allows to recover more cells and efficiently release samples into liquid medium. The eNATTM medium is capable of preserving the DNA while inhibiting bacterial growth. Moreover, rapid release of cells into the eNATTM medium enables immediate preservation of patient samples, allowing repeats and additional tests performed with the original sample and facilitating automated tests. Therefore this collection system aims to conveniently acquire a high yield of patient sample with minimal contamination.
RESULTS Procedures for preservation and transportationBuccal swab samples were co-collected using COPAN FLOQSwabsTM with 1ml eNATTM medium and the EpicentreCatch-All Sample Collection Swab. Both swabs were placed next to each other and collected at the same time. After collection, FLOQSwabTM was broken and preserved in the eNATTM medium tube immediately. Epicentre Catch-All samples needs to be refrigerated if not processed immediately while FLOQSwabsTM with eNATTM medium can be stored at room temperature without compromising sample quality. During extraction, a 400mL portion of eNATTM medium was transferred directly for process while the Epicenter swab needed to be cut and transferred into a new tube for pretreatment. More than half of the eNATTM medium including the FLOQSwabTM were left for future applications.
SNP genotyping assaysGenomic DNA were extracted using Thermo Fisher MagMAX™ DNA Multi-Sample Ultra Kit. Customized PGx panels were run using 7900HT Fast Real-Time PCR System on the extracted gDNA. Data were analyzed with TaqMan® Genotyper Software. Results generated with eNATTM-extracted DNA showed good amplification and confident calls (Fig 3). We compared the genotyping results with Epicenter swab-extracted DNA and eNATTM-extracted DNA, all 47 assays of 17 genes showed 100% agreement (Table 1) .
Fig 3. Screenshot of TaqMan® Genotyper Software analyzing genotyping data from both sample types.
Table 1. Agreement of all 47 assays of 17 genes in the PGx panel from both sample types.
Epicentre Catch-AllTM
CONCLUSIONSCOPAN FLOQSwabsTM with 1ml eNATTM medium kit is a very convenient device for buccal sample collection, storage and transportation for nucleic acids preservation. Direct lysis in the eNATTM medium tube inactivates bacterial proliferation, reduces sample processing steps and facilitates automation. The gDNAyielded from a 400ul portion of eNATTM medium is pure and adequate for genetic testing. Moreover, the leftover eNATTM
medium can be stored for additional and repeat confirmatory testing if needed, avoiding sample recollection.
Fig 1. Picture of FLOQSwabsTM with 1ml eNATTM medium and the Epicentre Catch-AllTM
Sample Collection Swab
Fig 2. 7900HT Fast Real-Time PCR System
FLOQSwabs™ and eNAT™
Gene ANKK1 APOE COMT CYP1A2 CYP2B6 CYP2C19 CYP2C9 CYP2D6 CYP3A4
Agreement 100% 100% 100% 100% 100% 100% 100% 100% 100%
Gene CYP3A5 FII FV GRIK4 MTHFR OPRM1 SLCO1B1 VKORC1
Agreement 100% 100% 100% 100% 100% 100% 100% 100%
