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Questions & Answers

General Questions

What is Epi proColon®2.0 CE?

Epi proColon® 2.0 CE is a molecular, colorectal cancer screening test that detects methylated Septin 9 DNA in blood. DNA methylation of the SEPT9 gene is increased in colorectal cancer. Methylated SEPT9 DNA from tumors is shed into the bloodstream and the methylation pattern is detectable in plasma by Real-Time PCR.

Why a blood test?

A blood test is a common and accepted method of testing. A colorectal cancer screening test based on a blood draw may be acceptable to screening-eligible people who have a history of not completing CRC screening. Even after counseling with their healthcare provider, only 1 in 7 people in the EU undergo screening by tests requiring stool sample collection and shipping, or more invasive tests requiring advance preparation and sedation such as colonoscopy and sigmoidoscopy. A blood test has the potential to increase screening participation in the eligible population that is currently not screened.

Who should be screened with Epi proColon®2.0 CE?

Epi proColon® 2.0 CE is indicated for average-risk people, age 50 or older, to provide an additional option for colorectal cancer screening for people who have a history of not completing screening by colonoscopy and stool tests.

Can Epi proColon® 2.0 CE be used to screen or monitor patients at high-risk for colorectal cancer (CRC)?

No. The Epi proColon® 2.0 CE test is a screening test for use in patients considered to be at average-risk for acquiring CRC.

Are there any age restrictions for the test?

The test was clinically studied in a demographically diverse group of people between the ages of 50-85 who were considered to be of average-risk for colorectal cancer. CRC Guidelines generally recommend screening for all people between the ages of 50 and 75.

Since this is a routine CRC screening test, how often should patients be tested?

The interval for use for Epi proColon® 2.0 CE has not been established in clinical evaluations. Programmatic performance has not been studied.

What should a patient do if Epi proColon® 2.0 CE is positive?

A positive test means that methylated Septin 9 DNA has been detected in the plasma sample tested. Methylated Septin 9 has been associated with the occurrence of colorectal cancer. Because Epi proColon® 2.0 CE is not a confirmatory test for the presence of colorectal cancer, people with a positive result should be counseled by their healthcare provider to undergo diagnostic colonoscopy to confirm a positive test.

What should a patient do if Epi proColon® 2.0 CE is negative?

A negative blood test result indicates the absence of methylated Septin 9 DNA in the plasma sample tested. Because a negative test result is not confirmatory for the absence of colorectal cancer, persons should be advised to continue participating in a recommended screening program according to screening guidelines.

Colorectal Cancer & Screening

Why should a person be screened for colorectal cancer?

Colorectal cancer is one of the most preventable and treatable cancers when it is found early. Based on the results of very large clinical trials, screening has been shown to reduce the mortality and incidence of colorectal cancer. Early stages of CRC are usually asymptomatic and therefore, early detection by screening can result in a greatly improved 5-year rate of survival and a more likely cure.

What are the recommended screening methods?

There are a number of national screening guidelines that make recommendations for screening and which tests to use. The most common tests for CRC screening recommended in the guidelines include colonoscopy (every 5-10 years) and high-sensitivity fecal occult blood testing (FIT test, annually or biannually).

What is the best way to get screened with Epi proColon®2.0 CE?

Epi proColon® 2.0 CE can be ordered by your healthcare provider during a routine or annual office visit. Your blood can be drawn at your doctor’s office or the blood collection center of a diagnostic lab. There are no dietary or medication restrictions before having your blood drawn. Before choosing Epi proColon®2.0 CE, you and your healthcare provider should discuss your screening options.

The Epi proColon® 2.0 CE Test – Background

What biomarkers are in the Epi proColon® 2.0 CE test?

Epi proColon® 2.0 CE is a test based on the detection of DNA methylation in a specific sequence of a promoter region of the SEPT9 gene. Epi proColon® 2.0 CE also measures an internal control gene (Actin beta) to ensure that the total DNA amount in the sample is sufficient to interpret the test.

What is DNA methylation?

DNA methylation is a natural biological process where the cytosine base of DNA is modified by the addition of a methyl group. This modification is associated with the regulation of gene expression. Changes in DNA methylation often occur in cancer and can serve as a biomarker for finding cancer.

Can I use serum instead of plasma?

Epi proColon® 2.0CE was developed using plasma obtained from either BD Vacutainer® K2EDTA or Sarstedt S-Monovette® 9 ml K3E or Sarstedt S-Monovette® 8.5 ml CPDA blood collection tubes with a minimum of 3.5 mL plasma needed to perform the test. While there are many publications describing DNA in serum, serum has not been validated for use with this test.

What is cell-free circulating DNA?

Cell-free circulating DNA is the fraction of DNA that occurs in blood plasma (bloodstream).

Blood Collecting and Processing

What are the blood tube options for collecting blood specimens?

Blood should be collected in either a BD Vacutainer® K2EDTA or a Sarstedt S-Monovette® 9 ml K3E or a Sarstedt S-Monovette® 8.5 ml CPDA blood collection tube. Other manufacturer’s tubes have not been validated for use with Epi proColon® 2.0 CE.

How much blood should be collected?

Epi proColon® 2.0 CE requires a minimum of 3.5 mL of cell-free plasma. To obtain this amount of plasma, we recommend that you collect a full 10 mL tube of blood—ensure that the evacuated tube is allowed to complete the fill volume. Some blood draw stations collect a second blood collection tube as a backup measure.

Note: Do not freeze whole blood samples.

Are there specific plasma preparation requirements?

Plasma preparation should be performed within 4 hours after drawing blood. Whole blood collected in blood collection tubes may be stored at 2°- 30°C prior to plasma preparation.

How should the plasma be stored and transported?

The plasma sample may be stored refrigerated, at 2°C to 8°C for up to 72 hours, or frozen, at -15°C to -25°C for up to 14 days. Transport and shipment of refrigerated or frozen plasma should follow the same temperature and time requirements as stated above.

Epi proColon® 2.0 CE Test Procedure

The test kit is designed for 30 plasma samples and 2 controls (performed in triplicate). What if I have less than 30 patient samples to test?

The components of the Epi proColon® 2.0 CE Plasma Quick Kit and PCR Kit are provided in aliquots to allow multiple runs containing a smaller number of patient samples. The Epi proColon® 2.0 CE Plasma Quick Kit and PCR Kit provide sufficient reagents for 4 runs. The Epi proColon® 2.0 CE Control Kit provides sufficient Positive and Negative control solution for 6 runs.

Is there an automated version of the test?

Epi proColon® 2.0 CE has been validated for use as a manual test protocol. A laboratory may choose to validate an automated protocol for this test; however, the test performance characteristics would not be covered by the manufacturer’s stated metrics and performance specifications.

Can the laboratory substitute test components?

No. The test has been approved for use with only Epi proColon® 2.0 CE labeled kit components, reagents, and materials that are specified in the Instructions for Use (IFU 0009).

Can the laboratory substitute equipment?

Epi proColon® 2.0 CE has been validated for use only with the Applied BioSystems 7500 Fast Dx and the Roche LightCycler 480 (I+II) as specified in the Instructions for Use (IFU 0009).

Can the test be performed on other PCR instruments or with other software?

The test has been validated for use on the AB 7500 Fast DX with SDS v1.4 or v1.4.1 software, 21 CFR Part 11 and the Roche LigtCycler 480 (I+II) with LightCycler 480 Software 1.5.x. The use of any other instrument and software requires additional validation.

Can the magnetic beads be over-dried?

Yes. In Step 9.13 of IFU0009, do not increase drying time or temperature as over-drying of the magnetic beads might reduce bisDNA recovery. If over-drying occurs, the 10 minutes drying time may need to be reduced.

Following the extraction step from plasma, if the magnetic beads appear to remain in solution or magnetic capture appears to be inefficient, what should be the next steps?

The tube should be incubated at 56°C for up to 10 min to improve capture efficiency. Following incubation at 56°C, place the tube into the DynaMag™-15 magnetic racks for another 5 to 10 min and continue processing following the Instructions for Use (IFU 0009).

Are there any special considerations when working with magnetic beads?

A homogeneous suspension of the Epi proColon® 2.0 CE Magnetic Beads is essential for proper performance. Deviations from the specified amount of beads may lead to false results. To ensure correct magnetic bead concentration, the bottle should be mixed thoroughly just before pipetting. There should be no visible sediment at the bottom of the bottle. Ensure a homogeneous suspension between the pipetting steps.

What results are reported to the health care provider?

The test is a qualitative test and provides three reportable result outcomes: Negative; Positive; Invalid. Test runs obtaining invalid results should not be reported to a healthcare provider. A new blood sample should be obtained and the test repeated. If invalid results are obtained a second time, please call Support.

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