Test Catalog

Test Id : BRMLH

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MLH1 Hypermethylation and BRAF Mutation Analysis, Tumor

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Overview

Test Catalog

Useful For
Suggests clinical disorders or settings where the test may be helpful

An adjunct to MSI / Microsatellite Instability (MSI), Tumor and IHC / Mismatch Repair (MMR) Protein Immunohistochemistry Only, Tumor testing, when colon tumor demonstrates microsatellite instability (MSI-H) and loss of MLH1 protein expression, to help distinguish a somatic versus germline event prior to performing expensive germline testing

 

An adjunct to negative MLH1 germline testing in cases where colon tumor from the same patient demonstrates MSI-H and loss of MLH1 protein expression

Genetics Test Information
Provides information that may help with selection of the correct genetic test or proper submission of the test request

If this test is ordered in conjunction with the MLH1 immunostain (IHC / Mismatch Repair [MMR] Protein Immunohistochemistry Only, Tumor) and MSI (MSI / Microsatellite Instability [MSI], Tumor), this test will only be performed when clinically indicated.

Profile Information
A profile is a group of laboratory tests that are ordered and performed together under a single Mayo Test ID. Profile information lists the test performed, inclusive of the test fee, when a profile is ordered and includes reporting names and individual availability.

Test Id Reporting Name Available Separately Always Performed
PBMLH MLH1 Hypermethylation/BRAF Mutation No Yes

Reflex Tests
Lists tests that may or may not be performed, at an additional charge, depending on the result and interpretation of the initial tests.

Test Id Reporting Name Available Separately Always Performed
BMLHH MLH1 Hypermethylation Analysis Yes, (order ML1HM) No
BBRAF BRAF Analysis Yes, (order BRAFD) No

Additional Tests
Lists tests that are always performed, at an additional charge, with the initial tests.

Test Id Reporting Name Available Separately Always Performed
SLIRV Slide Review in MG No, (Bill Only) Yes

Testing Algorithm
Delineates situations when tests are added to the initial order. This includes reflex and additional tests.

When this test is ordered, BRAF analysis and MLH1 hypermethylation analysis will always be performed. The exception would be if the tissue origin is an endometrial tumor; in those cases, only the MLH1 hypermethylation analysis component will be performed.

 

When this test is ordered, slide review will always be performed at an additional charge.

 

See Lynch Syndrome Testing Algorithm

Method Name
A short description of the method used to perform the test

Methylation-Specific Polymerase Chain Reaction (PCR) and Digital Droplet Polymerase Chain Reaction (ddPCR)

NY State Available
Indicates the status of NY State approval and if the test is orderable for NY State clients.

Yes

Reporting Name
Lists a shorter or abbreviated version of the Published Name for a test

MLH1 Hypermethylation/BRAF Mutation

Aliases
Lists additional common names for a test, as an aid in searching

BRAF Mutation

BRAF V600E

Hypermethylation

MLH1 Hypermethylation

Promoter Hypermethylation

MLBRF

Lynch syndrome

Testing Algorithm
Delineates situations when tests are added to the initial order. This includes reflex and additional tests.

When this test is ordered, BRAF analysis and MLH1 hypermethylation analysis will always be performed. The exception would be if the tissue origin is an endometrial tumor; in those cases, only the MLH1 hypermethylation analysis component will be performed.

 

When this test is ordered, slide review will always be performed at an additional charge.

 

See Lynch Syndrome Testing Algorithm

Specimen Type
Describes the specimen type validated for testing

Varies

Ordering Guidance

This test is not recommended as a first-tier screening for Lynch syndrome. Order TMSI / Microsatellite Instability, Tumor and IHC / Mismatch Repair (MMR) Protein Immunohistochemistry Only.

 

This test will only be performed on colon tumors demonstrating loss of MLH1 protein expression.

Necessary Information

Pathology report must accompany specimen in order for testing to be performed.

Specimen Required
Defines the optimal specimen required to perform the test and the preferred volume to complete testing

Specimen Type: Tissue block or slide

Collection Instructions:

1. Submit formalin-fixed, paraffin-embedded tissue block (preferred) or 1 slide stained with hematoxylin and eosin and 10 unstained, nonbaked slides (5 micron-thick sections) of the tumor tissue.

2. Sections should contain both tumor and normal tissue.

Special Instructions
Library of PDFs including pertinent information and forms related to the test

Forms

1. Molecular Genetics: Inherited Cancer Syndromes Patient Information (T519)

2. If not ordering electronically, complete, print, and send an Oncology Test Request (T729) with the specimen.

Reject Due To
Identifies specimen types and conditions that may cause the specimen to be rejected

Specimens that have been decalcified (all methods)
Specimens that have not been formalin-fixed, paraffin-embedded
Extracted DNA 		Reject

Specimen Stability Information
Provides a description of the temperatures required to transport a specimen to the performing laboratory, alternate acceptable temperatures are also included

Specimen Type Temperature Time Special Container
Varies Ambient (preferred)
Frozen
Refrigerated

Useful For
Suggests clinical disorders or settings where the test may be helpful

An adjunct to MSI / Microsatellite Instability (MSI), Tumor and IHC / Mismatch Repair (MMR) Protein Immunohistochemistry Only, Tumor testing, when colon tumor demonstrates microsatellite instability (MSI-H) and loss of MLH1 protein expression, to help distinguish a somatic versus germline event prior to performing expensive germline testing

 

An adjunct to negative MLH1 germline testing in cases where colon tumor from the same patient demonstrates MSI-H and loss of MLH1 protein expression

Genetics Test Information
Provides information that may help with selection of the correct genetic test or proper submission of the test request

If this test is ordered in conjunction with the MLH1 immunostain (IHC / Mismatch Repair [MMR] Protein Immunohistochemistry Only, Tumor) and MSI (MSI / Microsatellite Instability [MSI], Tumor), this test will only be performed when clinically indicated.

Testing Algorithm
Delineates situations when tests are added to the initial order. This includes reflex and additional tests.

When this test is ordered, BRAF analysis and MLH1 hypermethylation analysis will always be performed. The exception would be if the tissue origin is an endometrial tumor; in those cases, only the MLH1 hypermethylation analysis component will be performed.

 

When this test is ordered, slide review will always be performed at an additional charge.

 

See Lynch Syndrome Testing Algorithm

Clinical Information
Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test

Lynch syndrome is an inherited cancer syndrome caused by a germline pathogenic variant in one of several genes involved in DNA mismatch repair (MMR), including MLH1, MSH2, MSH6, and PMS2. There are several laboratory-based strategies that help establish the diagnosis of Lynch syndrome, including testing tumor tissue for the presence of microsatellite instability (MSI-H) and loss of protein expression for any one of the MMR proteins by immunohistochemistry (IHC). It is important to note, however, that the MSI-H tumor phenotype is not restricted to inherited cancer cases; approximately 20% of sporadic colon cancers are MSI-H. Thus, MSI-H does not distinguish between a somatic (sporadic) and a germline (inherited) etiology, nor does it identify which gene is involved. Although IHC analysis is helpful in identifying the responsible gene, it also does not distinguish between somatic and germline defects.

 

Defective MMR in sporadic colon cancer is most often due to an abnormality in MLH1, and the most common cause of gene inactivation is promoter hypermethylation (epigenetic silencing). A specific alteration in the BRAF gene (V600E) has been shown to be present in approximately 70% of tumors with hypermethylation of the MLH1 promoter. Importantly, the V600E alteration is rarely identified in cases with germline MLH1 pathogenic variants. Thus, direct assessment of MLH1 promoter methylation status and testing for the BRAF V600E alteration can be used to help distinguish between germline etiologyand epigenetic/somatic inactivation of MLH1. Tumors that have the BRAF V600E alteration and demonstrate MLH1 promoter hypermethylation are almost certainly sporadic, whereas tumors that show neither are most often caused by an inherited (germline) pathogenic variant.

 

Although testing for the BRAF V600E alteration and MLH1 promoter hypermethylation are best interpreted together, they are also available separately to accommodate various clinical situations and tumor types. These tests can provide helpful diagnostic information when evaluating an individual suspected of having Lynch syndrome, especially when testing is performed in conjunction with MSI / Microsatellite Instability (MSI), Tumor and IHC / Mismatch Repair (MMR) Protein Immunohistochemistry Only, Tumor. It should be noted that these tests are not genetic tests, but rather stratify the risk of having an inherited cancer predisposition and identify patients who might benefit from subsequent genetic testing.

 

See Lynch Syndrome Testing Algorithm

 

Reference Values
Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.

An interpretive report will be provided.

Interpretation
Provides information to assist in interpretation of the test results

An interpretive report will be provided.

Cautions
Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances

Testing tumors other than colon (in the evaluation of Lynch syndrome) for BRAF and MLH1 hypermethylation has not been fully evaluated; therefore, other specimens are not accepted.

 

Colon cancer is relatively common, and it is possible for a sporadic colon cancer to occur in a Lynch syndrome family. Therefore, evaluation of other family members should still be considered in cases with MLH1 promoter hypermethylation and absence of the BRAF V600E alteration if there is high clinical suspicion of Lynch syndrome.

Clinical Reference
Recommendations for in-depth reading of a clinical nature

1. Cunningham JM, Kim CY, Christensen ER, et al: The frequency of hereditary defective mismatch repair in a prospective series of unselected colorectal carcinomas. Am J Hum Genet. 2001;69:780-790

2. Wang L, Cunningham JM, Winters JL, et al: BRAF mutations in colon cancer are not likely attributable to defective DNA mismatch repair. Cancer Res. 2003;63:5209-5212

3. Domingo E, Laiho P, Ollikainen M, et al: BRAF screening as a low-cost effective strategy for simplifying HNPCC genetic testing. J Med Genet. 2004;41:664-668

4. Bettstetter M, Dechant S, Ruemmele P, et al: Distinction of hereditary nonpolyposis colorectal cancer and sporadic microsatellite-unstable colorectal cancer through quantification of MLH1 methylation by real-time PCR. Clin Cancer Res. 2007;13:3221-3228

5. Gupta S, Provenzale D, Llor X, et al: NCCN Guidelines Insights: Genetic/familial high-risk assessment: colorectal, version 2.2019. J Natl Compr Canc Netw. 2019;17(9):1032-1041

Method Description
Describes how the test is performed and provides a method-specific reference

A methylation-specific polymerase chain reaction (PCR)-based assay is used to test tumor DNA for the presence of hypermethylation of the MLH1 promoter, based on a modification of the method described by Grady et al (Grady WM, Rajput A, Lutterbaugh JD, Markowitz S: Detection of aberrantly methylated hMLH1 promoter DNA in the serum of patients with microsatellite unstable colon cancer. Cancer Res 2001;61:900), and digital droplet PCR (ddPCR) is used to test for the presence of the V600E alteration within the BRAF gene.(Unpublished Mayo method)

 

PDF Report
Indicates whether the report includes an additional document with charts, images or other enriched information

No

Day(s) Performed
Outlines the days the test is performed. This field reflects the day that the sample must be in the testing laboratory to begin the testing process and includes any specimen preparation and processing time before the test is performed. Some tests are listed as continuously performed, which means that assays are performed multiple times during the day.

Varies

Report Available
The interval of time (receipt of sample at Mayo Clinic Laboratories to results available) taking into account standard setup days and weekends. The first day is the time that it typically takes for a result to be available. The last day is the time it might take, accounting for any necessary repeated testing.

7 to 14 days

Specimen Retention Time
Outlines the length of time after testing that a specimen is kept in the laboratory before it is discarded

Extracted DNA: 3 months

Performing Laboratory Location
Indicates the location of the laboratory that performs the test

Rochester

Fees
Several factors determine the fee charged to perform a test. Contact your U.S. or International Regional Manager for information about establishing a fee schedule or to learn more about resources to optimize test selection.

  • Authorized users can sign in to Test Prices for detailed fee information.
  • Clients without access to Test Prices can contact Customer Service 24 hours a day, seven days a week.
  • Prospective clients should contact their account representative. For assistance, contact Customer Service.

Test Classification
Provides information regarding the medical device classification for laboratory test kits and reagents. Tests may be classified as cleared or approved by the US Food and Drug Administration (FDA) and used per manufacturer instructions, or as products that do not undergo full FDA review and approval, and are then labeled as an Analyte Specific Reagent (ASR) product.

This test was developed and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. It has not been cleared or approved by the US Food and Drug Administration.

CPT Code Information
Provides guidance in determining the appropriate Current Procedural Terminology (CPT) code(s) information for each test or profile. The listed CPT codes reflect Mayo Clinic Laboratories interpretation of CPT coding requirements. It is the responsibility of each laboratory to determine correct CPT codes to use for billing.

CPT codes are provided by the performing laboratory.

Slide Review

88381-Microdissection, manual

 

81210-BRAF (v-raf murine sarcoma viral oncogene homolog B1) (eg, colon cancer), gene analysis, V600E variant, if appropriate

81288-MLH1 promoter methylation analysis, if appropriate

LOINC® Information
Provides guidance in determining the Logical Observation Identifiers Names and Codes (LOINC) values for the order and results codes of this test. LOINC values are provided by the performing laboratory.

Test Id Test Order Name Order LOINC Value
BRMLH MLH1 Hypermethylation/BRAF Mutation 97761-1
Result Id Test Result Name Result LOINC Value
Applies only to results expressed in units of measure originally reported by the performing laboratory. These values do not apply to results that are converted to other units of measure.
53223 Result Summary 50397-9
53224 Result 82939-0
53225 Interpretation 69047-9
53226 Specimen 31208-2
53227 Source 31208-2
53228 Tissue ID 80398-1
54921 BRAF Analysis No LOINC Needed
54440 MLH1 Hypermethylation Analysis No LOINC Needed
53229 Released By 18771-6
55139 Method 85069-3
621822 Disclaimer 62364-5

Test Setup Resources

Setup Files
Test setup information contains test file definition details to support order and result interfacing between Mayo Clinic Laboratories and your Laboratory Information System.

Excel | Pdf

Sample Reports
Normal and Abnormal sample reports are provided as references for report appearance.

Normal Reports | Abnormal Reports

SI Sample Reports
International System (SI) of Unit reports are provided for a limited number of tests. These reports are intended for international account use and are only available through MayoLINK accounts that have been defined to receive them.

SI Normal Reports | SI Abnormal Reports

Test Update Resources

Change Type Effective Date
File Definition - Result ID 2024-10-31