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AmoyDx Multi-Gene Mutations Detection Kit

Manufactured by Amoy Diagnostics
Sourced in China

The AmoyDx Multi-Gene Mutations Detection Kit is a laboratory diagnostic product. It is designed to detect multiple gene mutations simultaneously. The kit utilizes advanced molecular techniques to identify specific genetic alterations. It provides a comprehensive analysis of selected genes relevant to various clinical conditions.

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6 protocols using AmoyDx Multi-Gene Mutations Detection Kit

Specimens were preserved in formalin and were transferred for pathological diagnosis and genomic testing. The pathological diagnosis was evaluated after hematoxylin & eosin and immunohistochemistry staining. The DNA/RNA was extracted from formalin-fixed, paraffin-embedded specimens via an FFPE DNA Extraction Kit (Amoy Diagnostics). The genomic testing of all specimens was performed in an AmoyDx Multi-Gene Mutations Detection Kit (Amoy Diagnostics), which harbored DNA-based mutation and RNA-based fusion detection real-time PCR assays, and can detect the mutational status of EGFR, KRAS, BRAF, NRAS, HER2, PIK3CA, ALK, ROS1, MET, and RET simultaneously [22 (link)].
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Oncogenic alterations of EGFR, ALK, ROS1, RET, KRAS, BRAF, HER2, NRAS, and PIK3CA were detected by fluorescence quantitative PCR in tumor samples using Stratagene Mx3000P™ and AmoyDx Multi-Gene Mutations Detection Kit (Amoy Diagnostics, Xiamen, China). Fusions of ALK, ROS1 and RET genes were identified by mRNA-based methods using specific primers and fluorescent probes in a one-step RT-PCR [13 (link)] and point mutations in EGFR, KRAS, BRAF, NRAS, HER2, and PIK3CA genes were identified using the ADx-amplification refractory mutation system (ARMS) as previously described [14 (link)]. Only the canonical oncogenic mutations such as EGFR L858R/19Del/T790M, KRAS G12C/A/V/R, BRAF V600E, NRAS G12C/R/V/A, PIK3CA E545K, HER2 20ins etc. were included in the current study. This sensitive ARMs-qPCR assay can detect mutations present in as low as 1% of cell population [15 (link)]. The results were confirmed according to the manufacturer’s guidelines.
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The AmoyDx Multigene Mutations Detection Kit (Amoy Diagnostics, Xiamen, People's Republic of China) was performed for all positive samples, according to the manufacturer's instructions. This RNA-based assay is designed to detect six different RET fusion variants (i.e., CCDC6[1]-RET[12], NCOA4[6]-RET[12], KIF5B[15]-RET[12], KIF5B[16]-RET[12], KIF5B[22]-RET[12] and KIF5B[23]-RET[12]) on a Cobas z 480 (user-defined function channel) instrument.
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DNA and RNA were required for next molecular analyses for every sample. The concentration of DNA should be greater than 2 ng/μl, the concentration of RNA should be between 10 and 500 ng/μl. And the OD260/OD280 of DNA and RNA should be between 1.8 and 2.1. The molecular analysis of all samples was carried out by using AmoyDx® Multi‐Gene Mutations Detection Kit (Amoy Diagnostics) according to the manufacturer's instructions. This kit contains DNA‐based mutation detection real‐time PCR assays and RNA‐based fusion detection real‐time PCR assays, which can detect 118 hotspot mutations/fusions in genes of EGFR, KRAS, BRAF, NRAS, HER2, PIK3CA, ALK, ROS1, and RET in a single test8 (Supplementary Method and Table S1).
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Three formalin-fixed paraffin-embedded (FFPE) specimens were needed for the test which were sectioned into thickness of 5-μm and contained at least 30% tumor cells in specimens. The molecular analysis was carried out by AmoyDx® Multi-Gene Mutations Detection Kit (Amoy Diagnostics, Xiamen, China), which is a real-time PCR assay for qualitative detection of 118 hotspot mutations/fusions in EGFR, KRAS, BRAF, NRAS, HER2, PIK3CA, ALK, ROS1 and RET genes (Details are described in Table S1). This kit contains RNA gene fusion and DNA gene mutation detection systems. The RNA gene fusion detection system uses specific primers and fluorescent probes which are designed for both side genes of the fusion, combining a one-step RT-PCR procedure to detect ALK, ROS1 and RET gene fusions based on tumor message RNA. The DNA gene mutation detection system uses ADx-ARMS technology, which comprises specific primers and fluorescent probes to detect gene mutations. All experiments were performed according to the manufacturer’s instructions.
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The concentration of cf‐DNA and cf‐RNA should be greater than 2 and 10 ng/μl, respectively. According to the manufacturer's instructions, the genomic testing of all samples was carried out using an AmoyDx Multi‐Gene Mutations Detection Kit (Amoy Diagnostics). The DNA‐based mutation detection and RNA‐based fusion detection real‐time PCR assays were contained in the kit, which can detect 118 hotspot mutations/fusions in genes of EGFR, KRAS, BRAF, NRAS, HER2, PIK3CA, ALK, ROS1, MET, and RET simultaneously.
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