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Human Methylated NDRG4/SEPT9/SFRP2/BMP3/SDC2 Gene Detection Kit (Fluorescence PCR)

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Human Methylated NDRG4/SEPT9/SFRP2/BMP3/SDC2 Gene Detection Kit (Fluorescence PCR)

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Introduction

[Product Name]

Human Methylated NDRG4/SEPT9/SFRP2/BMP3/SDC2 Gene Detection Kit (Fluorescence PCR)
[Packaging Size]
24 tests/kit, 48 tests/kit

[Intended Use]

The kit is intended for in vitro qualitative detection of the methylated NDRG4/SEPT9/SFRP2/BMP3/SDC2 genes in intestinal exfoliated cells in human stool samples.
This kit is suitable for the auxiliary diagnosis of colonoscopy patients recommended by clinicians, and cannot be used as the basis for early diagnosis or diagnosis of tumors. The test results are for clinical reference only. For specific clinical applications, clinicians must make judgments based on the actual situation of the case. In adults, more than 108 intestinal epithelial cells fall off the intestinal wall every day, and are excreted with the feces through large bowel peristalsis. Due to the tumor cells are more likely to fall off the intestinal tract of the abnormal proliferation, the stool of intestinal tumor patients contains many diseased cells and abnormal cell components, which is the material basis for stable stool detection [1]. Studies have found that methylation modification of gene promoters is an early event in tumorigenesis, and genetic material obtained from stool samples of colorectal cancer patients can reflect the presence of cancer in the intestine earlier [2].

NDRG4, also known as SMAP-8 and BDM1, is one of the four members of the NDRG gene family (NDRG1-4), which has been shown to be associated with cell proliferation, differentiation, development and stress [3]. It is verified that NDRG4 methylation is a potential biomarker for non-invasive detection of colorectal cancer in stool samples [4].

SEPT9 is a member of the Septin gene family, which consists of at least 13 genes that encode a conserved GTPase domain that can bind cytoskeleton-related proteins, and is associated with cell division and tumorigenesis. Studies have found that the methylated Septin9 gene content is characteristically increased in stool samples from patients with colorectal cancer [5].

Secreted frizzled-related proteins (sFRPs) are soluble proteins that are a class of Wnt pathway antagonists due to their high structural homology to the frizzled (Fz) receptor for Wnt signaling [6]. Inactivation of the SFRP gene results in uncontrolled activation of Wnt signaling associated with colorectal cancer [7]. Currently, SFRP2 methylation 

in stool can be used as a non-invasive biomarker for the diagnosis of colorectal cancer [8,9].

BMP3 is a member of the TGF-B superfamily and thus plays an important role in

embryonic development by inducing and shaping early bone formation. BMP3 is

hypermethylated in colorectal cancer and can be used as an important tumor marker [10].

SDC2 is a cell surface heparan sulfate proteoglycan involved in the regulation of many physiological and pathological processes. Physical processed include cell proliferation, differentiation, adhesion, cytoskeletal organization, migration, wound healing, cell-matrix communication, angiogenesis; Pathological processes include inflammation and cancer [11]. The methylation level of SDC2 gene in colorectal cancer tissues was significantly higher than that in normal tissues [12].

[Test Principle]

This kit mainly includes two steps which are extraction and transformation step and fluorescent PCR step. The extraction and transformation step is to extract the genomic DNA of intestinal exfoliated cells from human stool samples by magnetic bead capture. Then, the unmethylated DNA is transformed with sulfite while methylated DNA is not transformed by sulfite. The fluorescent PCR step is to detect the conserved sequences of methylated NDRG4 gene (VIC/HEX channel), SEPT9 gene (ROX channel) and ACTB gene (CY5 channel) in one reaction well and detect the conserved sequences of methylated SFRP2 gene (VIC/HEX channel), BMP3 gene (ROX channel), SDC2 gene (FAM channel) and ACTB gene (CY5 channel) by methods of multiplex fluorescent PCR.

The ACTB gene is used as an internal control gene to evaluate whether the sample DNA is sufficient and whether it is qualified.

[Storage Conditions and Shelf-life]

The kit should be stored below -18°C protected from light. The shelf life is 9 months(please use within the shelf life). The number of repeated freezing and thawing should not be more than 4 cycles. It can be stably stored for 5 days below -18°C and protected from light during transportation.

See the packaging label for the production date, batch number and expiration date.

[Applicable Instruments]

QuantStudio®5 Real-Time PCR Systems, SLAN-96P Real-Time PCR Systems(Shanghai Hongshi Medical Technology Co., Ltd.), LightCycler®480 Real-Time PCR system, LineGene 9600 Plus Real-Time PCR Detection System (FQD-96A,Bioer technology), MA-6000 Real-Time Quantitative Thermal Cycler (Suzhou Molarray Co., Ltd.)

[Requirements for Samples]

1.Sample Collection
1.1 Stool Sample
Use a stool collector to collect stool samples from suspected infected patients in a disposable sterile container and close the tube cap tightly. Sample amount requirements:
Solid sample: The collection volume is about the size of a small ball with a diameter of about 2cm.
Liquid samples: The collection volume is about 5mL.
2.Storage
Test samples should be stored at 2-8°C for no more than 7 days, and should be stored below -18°C for no more than 4 months, and could be stored below -70°C for long-term. The samples should be repeated freezing and thawing.

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