b'Gel & Capillary | IVD Assays FLT3 Mutation Assay 2.0Gel and Capillary Assays andAssays FLT3 Mutation Assay 2.0Gel and Capillary Assays andAssaysABI Fluorescence DetectionABI Fluorescence Detection ReagentsFLT3 Mutation Assay 2.0 - Controls Concentration Units in Assay Units in MegaKitFLT3 ITD Positive Control 50 g/mL 1 x 100 L tube 5 x 100 L tubesABI Fluorescence Detection FLT3 D835 Positive Control 50 g/mL 1 x 100 L tube 5 x 100 L tubesFLT3 Negative Control 50 g/mL 1 x 100 L tube 5 x 100 L tubesMaster Mixes Target Units in Assay Units in MegaKitFLT3 ITD Master Mix6FAM & HEX FLT3 ITD 1 x 1500 L tube 10 x 1500 L tubesAssay Description Table 1. FLT3 ITD Percent Agreement with 454 Sequencing FLT3 D835 Master Mix6FAM FLT3 TKD 1 x 1500 L tube 10 x 1500 L tubesThe LeukoStrat FLT3 Mutation Assay 2.0 is an in vitro diagnosticPercent Agreement Discordance # Concordance # *95% LLproduct intended for PCR-based detection of FLT3 activating mutations in patients with acute myelogenous leukemia (AML). Negative PA 100% 0 119 96.9% Capillary Electrophoresis Detection (ABI)Specifically, the FLT3 Mutation Assay 2.0 can be used to: Positive PA 98.0% 4 200 95.1% Differential fluorescence detection, such as ABI fluorescence detection, is commonly used to resolve different-sized amplicon products using Identify internal tandem duplications (ITD) in the FLT3 gene * 95% of results would be expected to agree with sequencing at a rate greater thana capillary electrophoresis instrument. Primers can be conjugated with different fluorescent dyes (fluorophores), so that they produce different or equal to the lower limit (LL). emission spectra upon excitation by a laser in the capillary electrophoresis instrument. In this manner, different fluorescent dyes can correspond Identify tyrosine kinase domain (TKD) mutations in the FLT3 gene to different targeted regions. This detection system results in high sensitivity, single nucleotide resolution, differential product detection, and relative quantification. In addition, differential detection allows accurate, reproducible and objective interpretation of primer-specific products. Inter-assay Summary and Explanation of the TestTable 2. FLT3 TKD Percent Agreement with 454 Sequencing and intra-assay reproducibility in size determination using capillary electrophoresis is approximately 1 to 4 nucleotides.AML in general has a poor prognosis. 1,2Many studies in AML haveThe data shown was generated using the master mixes indicated. Amplified products were run on an ABI 3500xL instrument.shown that the presence of FLT3 (fms related tyrosine kinase 3)Percent Agreement Discordance # Concordance # *95% LLactivating mutations portends a poor prognosis making it an attractive target for treatment. For this reason, FLT3 mutation testing is required toNegative PA 100% 0 137 96.9%stratify disease and determine appropriate treatment options.Positive PA 100% 0 240 98.5%Using this assay, DNA is amplified via PCR with fluorophore labeled* 95% of results would be expected to agree with sequencing at a rate greater than primers, TKD amplicon is enzymatically digested, and FLT3 mutationsor equal to the lower limit (LL).are detected via capillary electrophoresis. This test kit includes 2 PCR master mixes, along with positive and negative controls for mutant detection. Each master mix (FLT3 ITD Master Mix and FLT3 D835 Master Mix) contains a fluorophore-labeled PCR primer set for the respectiveReferencesdetection of internal tandem duplication or TKD mutation. 1. Murphy KM et al., A Clinical PCR/Capillary Electrophoresis Assay for the Detection of Internal Tandem Duplication and Point Mutation of Performance Characteristicsthe FLT3 Gene. J. Mol. Diag. 5:96-102 (2003).This assay can reliably detect mutations comprising more than 5% of2. Yamamoto, Y., et al., Activating mutation of D835 within the the total cell population. Also, as demonstrated herein, the LeukoStratactivation loop of FLT3 in human hematologic malignancies. Blood, FLT3 Mutation Assay 2.0 detects FLT3 ITD and TKD mutations with97(8):2434-9 (2001).excellent concordance to NGS methodologies (Table 1, Table 2).3.Acute Myeloid Leukemia, Clinical Practice Guidelines in Oncology, National Comprehensive Cancer Network (v.2.2014)Ordering InformationCatalog # Products Quantity9-412-0091 LeukoStrat FLT3 Mutation Assay 2.0ABI Fluorescence Detection 33 reactions9-412-0101 LeukoStrat FLT3 Mutation Assay 2.0 MegaKitABI Fluorescence Detection 330 reactionsThese are in vitro diagnostic products, and are not available for sale or use within North America.84Invivoscribe 2021|85'