Ordering Recommendations

A completed test request form must be included with the specimen. Ensure that the following information is included on the test request form:
  • Clinical Indication
  • ICD-10 Diagnosis Code
  • If Surgical Pathology needs to perform lymphoma protocol, this must be notated on the requisition

Performed

Monday - Friday

Methodology

Polymerase Chain Reaction / Sanger Sequencing

Reported

Routine: 10 days

Synonyms

  • Tumor
  • Tissue
  • FFPE Tissue
  • Cancer
  • Lung Adenocarcinomas
  • Colorectal Carcinoma
  • Colorectal Cancer
  • Pancreatic Carcinoma
  • Pancreatic Cancer
  • KRAS Gene
  • NRAS Gene
  • HRAS Gene
  • RAS Genes
  • c.38G>A
  • p.G13D
  • Anti-Epidermal Growth Factor Receptor Therapy
  • Epidermal Growth Factor Receptor
  • EGFR Therapy
  • Tyrosine Kinase Inhibitor Therapy
  • Clinical Genomics

Performing Lab

Surgical Pathology: Specimen Preparation and Storage of FFPE ONLY
Clinical Genomics: Testing Laboratory

Turnaround Time

After specimen receipt in the Clinical Genomics Lab, results are generally available within 10 days.

Add-on Eligibility

Yes
*If DNA has been extracted previously for other Clinical Genomics tests, this is stored for 6 weeks and may qualify for add-on

Specimen Type

Tumor Tissue

Specimen Preparation

  1. Formalin Fix the specimen by submerging the Tissue in 10% Neutral Buffered Formalin
  2. Send the Formalin-Fixed Tissue to the Surgical Pathology Lab
  3. The Surgical Pathology Lab will paraffin-embed the tissue and prepare a Tissue Block
  4. The Tissue block will be sent to the Clinical Genomics Laboratory for testing

Specimen Volume

Fresh Tissue Size: 1 cm3 (Minimum: 0.5 cm3)
  • Specimen must contain a region with at least 20% tumor
Fixative Volume: 15 times the volume of the tissue

*This refers to specimen volume at collection prior to any processing

Collection Container

*Tissue specimens need to be sent to Surgical Pathology after collection so that the specimen can be processed prior to transport to the Clinical Genomics Laboratory. Do NOT send directly to Clinical Genomics.
 
  1. Formalin Container
  • EXCEPT for cases of suspected lymphoma
 
OR
 
  1. Non-Sterile Liquid-Proof Container
  • Medium to large specimens for routine processing that do not fit in prefilled formalin containers can be can be sent in specimen containers without formalin
  • Surgical Pathology staff will add formalin when the specimen is recieved in the lab

Unacceptable Conditions

  1. Specimens containing < 20% or no tumor tissue
  2. The provider will be contacted regarding receipt of non-validated specimen types
  3. Specimens that have been improperly collected, stored, or transported
  • Specimens that have been preserved in fixatives other than formalin
  • Specimens that have been preserved in heavy metal fixatives (B-4 or B-5)
  • Specimens that have been frozen
  1. Specimens with insufficient volume for testing
  2. Unlabeled or mislabeled specimens

Storage/Transport Temperature

*Specimens should be collected and transported to the Surgical Pathology Lab for FFPE Tissue Block preparation
 
Transport Instructions
Collection Location Specimen Type Transport Temperature Timeframe
ED/
Inpatient		 Fresh Specimen Room Temperature Specimen must be received by the lab within 1 hour of collection
Refrigerated (Ice Packs) Specimen must be received by the lab within 24 hours of collection
Formalin Fixed Specimen Room Temperature Specimen must be received by the lab within 24 hours of collection
Laboratory/
Outpatient/
Off-Site 		 Fresh Specimen Room Temperature Specimen must be received by the lab within 1 hour of collection
Refrigerated (Ice Packs) Specimen must be received by the lab within 24 hours of collection
Formalin Fixed Specimen Room Temperature Specimen must be received by the lab within 24 hours of collection

Storage:
  • Fixed Specimens: Room Temperature
  • Fresh Specimens: Refrigerated

Stability (from collection to initiation)

Stability:

Formalin-Fixed Specimens
  • Room Temperature: 1 week
  • Refrigerated: Not recommended
  • Frozen: Unacceptable

Sterile Container
  • Room Temperature: 3 hours
  • Refrigerated: 24 hours
  • Frozen: Unacceptable

FFPE Tissue Block
  • Room Temperature: Indefinitely
  • Refrigerated: Indefinitely
  • Frozen: Unacceptable

Extracted DNA:
  • Room Temperature: Unacceptable
  • Refrigerated: Unacceptable
  • Frozen: Indefinitely

Laboratory Storage:
  • Tumor Tissue:
    • All tissue specimens are stored at the Surgical Pathology Lab
      • Fixed Specimens: Room Temperature
      • Fresh Specimens: Refrigerated
      • FFPE Tissue Blocks: Room Temperature
  • Extracted DNA: Frozen

Laboratory Retention:
  • FFPE Tissue Blocks: Returned to Surgical Pathology Lab after testing is completed
  • Extracted DNA: 6 weeks

Collection Instructions

Collection Instructions:

Routine Small and Medium Sized Specimens
  1. Collect Tissue specimen and place into a Sterile Container with enough 10% formalin to cover the tissue by at least 2 - 3 inches.
  • Specimens must contain a region with at least 20% tumor
  • Breast Specimens: Immerse in fixative within 1 hour of biopsy or resection procedure for optimum tissue processing. Document the time that the tissue was placed into formalin.
  • Suspicion of Lymphoma: Send FRESH. Do NOT place specimen into formalin.
*Must state that lymphoma protocol is needed on requisition
 
  1. Specimens should be delivered to the Surgical Pathology Lab as soon as possible.  If delivery of a resection specimen to the Surgical Pathology department is delayed (e.g. specimens from remote sites), the tumor should be bisected prior to immersion in fixative.
  • It is important that the surgeon ensure that the identity of the resection margins is retained in the bisected specimen
  • Alternatively, the margins may be separately submitted
  1. Ensure that the cap is tightly secured on the Collection Container
  2. The time of removal of the tissue and the time of immersion of the tissue in fixative should be recorded and submitted to the laboratory

Routine Large Sized Specimens
  1. Collect tissue specimen and place in large leak-proof container
  2. Bring specimen to the Surgical Pathology laboratory immediately, or refrigerate

Specimen Container Labeling Instructions:
  1. Specimen Containers submitted to Surgical Pathology must be labeled with the following patient information:
  • First and Last Name
  • Date of Birth
  • Medical Record Number
  • CSN Number
  • Name of Specimen Site/Source
*If more than one site is sampled, a separate collection container must be submitted for each site
 
  1. Check all labeling of containers prior to leaving the collection site
 
  1. Place the following in a biohazard bag for transport to the Surgical Pathology Lab:
*If more than one site is sampled, all specimen material collected must be packaged and separated according to specimen site
 
  • Specimen Containers
  • Specimen Requisitions

Processing Instructions:
  1. Formalin Fix fresh tissues in 10% Neutral Buffered Formalin
  2. Paraffin-embed the fixed tissue and prepare FFPE Tissue Block
  • Specimens must contain a region with at least 20% tumor
 
  1. Protect specimen from excessive heat and transport one of the following to the Clinical Genomics Laboratory for testing:
  • Tissue Block
    • Tissue block will be returned to the Surgical Pathology Lab after testing
  • Two to four 20 micron sections
  • Two to four unstained tissue sections on slides
    • Place slides into a protective container
     
 
  1. Include the Surgical Pathology report with the specimen

Reference Interval

Not Detected

Interpretive Data

Refer to report for result specific interpretation details.

KRAS Mutation Not Detected: The absence of KRAS exon 2 (i.e. codons 12 and 13), exon 3 (i.e. codons 59 and 61), or exon 4 (i.e. codons 117 and 146) mutations in the tumor specimen suggests that the patient may be a candidate for such targeted anti-EGFR therapy.
  • This assay is specific for mutations in and around codons 12, 13, 59, 61,117, and 146 of the KRAS gene. Mutations occurring at sites outside of these regions within the KRAS gene or within other genes will not be detected by this assay.
  • A negative test does not exclude the presence of a variant as the limit of detection of this assay is approximately 20% variant KRAS sequence in a background of wild-type sequence.
  • Not all patients with tumors harboring wild-type KRAS sequences will respond to EGFR-targeted therapies.

KRAS Mutation Detected: In patients with advanced or metastatic colorectal cancer, current literature suggests that tumors with a mutation in exons 2, 3, or 4 of the KRAS gene are essentially insensitive to anti-EGFR therapies such as cetuximab or panitumumab.  The presence of KRAS exon 2 (i.e. codons 12 and 13), exon 3 (i.e. codons 59 and 61), or exon 4 (i.e. codons 117 and 146) mutations in the tumor specimen suggests that the patient may not be a candidate for such targeted anti-EGFR therapy and is not recommended to receive treatment with cetuximab or panitumumab according to NCCN guidelines.
  • Point mutations in the KRAS gene which result in amino acid substitutions and produce a constitutively active form of KRAS are observed in a variety of human malignancies, including colorectal carcinoma, pancreatic carcinoma, and lung adenocarcinoma
  • The presence of mutations in KRAS exon 2 (codons 12, 13), exon 3 (codons 59, 61), or exon 4 (codons 117, 146) has been shown to be predictive of lack of response to anti-epidermal growth factor receptor (EGFR) therapy in metastatic colorectal carcinoma
  • Mutations within these codons represent approximately 99% of the KRAS mutations observed in colorectal cancer as listed in the Catalogue of Somatic Mutations in Cancer (COSMIC) database.
Ordering

Ordering Recommendations

A completed test request form must be included with the specimen. Ensure that the following information is included on the test request form:
  • Clinical Indication
  • ICD-10 Diagnosis Code
  • If Surgical Pathology needs to perform lymphoma protocol, this must be notated on the requisition

Performed

Monday - Friday

Methodology

Polymerase Chain Reaction / Sanger Sequencing

Reported

Routine: 10 days

Synonyms

  • Tumor
  • Tissue
  • FFPE Tissue
  • Cancer
  • Lung Adenocarcinomas
  • Colorectal Carcinoma
  • Colorectal Cancer
  • Pancreatic Carcinoma
  • Pancreatic Cancer
  • KRAS Gene
  • NRAS Gene
  • HRAS Gene
  • RAS Genes
  • c.38G>A
  • p.G13D
  • Anti-Epidermal Growth Factor Receptor Therapy
  • Epidermal Growth Factor Receptor
  • EGFR Therapy
  • Tyrosine Kinase Inhibitor Therapy
  • Clinical Genomics

Performing Lab

Surgical Pathology: Specimen Preparation and Storage of FFPE ONLY
Clinical Genomics: Testing Laboratory

Turnaround Time

After specimen receipt in the Clinical Genomics Lab, results are generally available within 10 days.

Add-on Eligibility

Yes
*If DNA has been extracted previously for other Clinical Genomics tests, this is stored for 6 weeks and may qualify for add-on
Collection

Specimen Type

Tumor Tissue

Specimen Preparation

  1. Formalin Fix the specimen by submerging the Tissue in 10% Neutral Buffered Formalin
  2. Send the Formalin-Fixed Tissue to the Surgical Pathology Lab
  3. The Surgical Pathology Lab will paraffin-embed the tissue and prepare a Tissue Block
  4. The Tissue block will be sent to the Clinical Genomics Laboratory for testing

Specimen Volume

Fresh Tissue Size: 1 cm3 (Minimum: 0.5 cm3)
  • Specimen must contain a region with at least 20% tumor
Fixative Volume: 15 times the volume of the tissue

*This refers to specimen volume at collection prior to any processing

Collection Container

*Tissue specimens need to be sent to Surgical Pathology after collection so that the specimen can be processed prior to transport to the Clinical Genomics Laboratory. Do NOT send directly to Clinical Genomics.
 
  1. Formalin Container
  • EXCEPT for cases of suspected lymphoma
 
OR
 
  1. Non-Sterile Liquid-Proof Container
  • Medium to large specimens for routine processing that do not fit in prefilled formalin containers can be can be sent in specimen containers without formalin
  • Surgical Pathology staff will add formalin when the specimen is recieved in the lab

Unacceptable Conditions

  1. Specimens containing < 20% or no tumor tissue
  2. The provider will be contacted regarding receipt of non-validated specimen types
  3. Specimens that have been improperly collected, stored, or transported
  • Specimens that have been preserved in fixatives other than formalin
  • Specimens that have been preserved in heavy metal fixatives (B-4 or B-5)
  • Specimens that have been frozen
  1. Specimens with insufficient volume for testing
  2. Unlabeled or mislabeled specimens

Storage/Transport Temperature

*Specimens should be collected and transported to the Surgical Pathology Lab for FFPE Tissue Block preparation
 
Transport Instructions
Collection Location Specimen Type Transport Temperature Timeframe
ED/
Inpatient		 Fresh Specimen Room Temperature Specimen must be received by the lab within 1 hour of collection
Refrigerated (Ice Packs) Specimen must be received by the lab within 24 hours of collection
Formalin Fixed Specimen Room Temperature Specimen must be received by the lab within 24 hours of collection
Laboratory/
Outpatient/
Off-Site 		 Fresh Specimen Room Temperature Specimen must be received by the lab within 1 hour of collection
Refrigerated (Ice Packs) Specimen must be received by the lab within 24 hours of collection
Formalin Fixed Specimen Room Temperature Specimen must be received by the lab within 24 hours of collection

Storage:
  • Fixed Specimens: Room Temperature
  • Fresh Specimens: Refrigerated

Stability (from collection to initiation)

Stability:

Formalin-Fixed Specimens
  • Room Temperature: 1 week
  • Refrigerated: Not recommended
  • Frozen: Unacceptable

Sterile Container
  • Room Temperature: 3 hours
  • Refrigerated: 24 hours
  • Frozen: Unacceptable

FFPE Tissue Block
  • Room Temperature: Indefinitely
  • Refrigerated: Indefinitely
  • Frozen: Unacceptable

Extracted DNA:
  • Room Temperature: Unacceptable
  • Refrigerated: Unacceptable
  • Frozen: Indefinitely

Laboratory Storage:
  • Tumor Tissue:
    • All tissue specimens are stored at the Surgical Pathology Lab
      • Fixed Specimens: Room Temperature
      • Fresh Specimens: Refrigerated
      • FFPE Tissue Blocks: Room Temperature
  • Extracted DNA: Frozen

Laboratory Retention:
  • FFPE Tissue Blocks: Returned to Surgical Pathology Lab after testing is completed
  • Extracted DNA: 6 weeks

Collection Instructions

Collection Instructions:

Routine Small and Medium Sized Specimens
  1. Collect Tissue specimen and place into a Sterile Container with enough 10% formalin to cover the tissue by at least 2 - 3 inches.
  • Specimens must contain a region with at least 20% tumor
  • Breast Specimens: Immerse in fixative within 1 hour of biopsy or resection procedure for optimum tissue processing. Document the time that the tissue was placed into formalin.
  • Suspicion of Lymphoma: Send FRESH. Do NOT place specimen into formalin.
*Must state that lymphoma protocol is needed on requisition
 
  1. Specimens should be delivered to the Surgical Pathology Lab as soon as possible.  If delivery of a resection specimen to the Surgical Pathology department is delayed (e.g. specimens from remote sites), the tumor should be bisected prior to immersion in fixative.
  • It is important that the surgeon ensure that the identity of the resection margins is retained in the bisected specimen
  • Alternatively, the margins may be separately submitted
  1. Ensure that the cap is tightly secured on the Collection Container
  2. The time of removal of the tissue and the time of immersion of the tissue in fixative should be recorded and submitted to the laboratory

Routine Large Sized Specimens
  1. Collect tissue specimen and place in large leak-proof container
  2. Bring specimen to the Surgical Pathology laboratory immediately, or refrigerate

Specimen Container Labeling Instructions:
  1. Specimen Containers submitted to Surgical Pathology must be labeled with the following patient information:
  • First and Last Name
  • Date of Birth
  • Medical Record Number
  • CSN Number
  • Name of Specimen Site/Source
*If more than one site is sampled, a separate collection container must be submitted for each site
 
  1. Check all labeling of containers prior to leaving the collection site
 
  1. Place the following in a biohazard bag for transport to the Surgical Pathology Lab:
*If more than one site is sampled, all specimen material collected must be packaged and separated according to specimen site
 
  • Specimen Containers
  • Specimen Requisitions

Processing Instructions:
  1. Formalin Fix fresh tissues in 10% Neutral Buffered Formalin
  2. Paraffin-embed the fixed tissue and prepare FFPE Tissue Block
  • Specimens must contain a region with at least 20% tumor
 
  1. Protect specimen from excessive heat and transport one of the following to the Clinical Genomics Laboratory for testing:
  • Tissue Block
    • Tissue block will be returned to the Surgical Pathology Lab after testing
  • Two to four 20 micron sections
  • Two to four unstained tissue sections on slides
    • Place slides into a protective container
     
 
  1. Include the Surgical Pathology report with the specimen
Result Interpretation

Reference Interval

Not Detected

Interpretive Data

Refer to report for result specific interpretation details.

KRAS Mutation Not Detected: The absence of KRAS exon 2 (i.e. codons 12 and 13), exon 3 (i.e. codons 59 and 61), or exon 4 (i.e. codons 117 and 146) mutations in the tumor specimen suggests that the patient may be a candidate for such targeted anti-EGFR therapy.
  • This assay is specific for mutations in and around codons 12, 13, 59, 61,117, and 146 of the KRAS gene. Mutations occurring at sites outside of these regions within the KRAS gene or within other genes will not be detected by this assay.
  • A negative test does not exclude the presence of a variant as the limit of detection of this assay is approximately 20% variant KRAS sequence in a background of wild-type sequence.
  • Not all patients with tumors harboring wild-type KRAS sequences will respond to EGFR-targeted therapies.

KRAS Mutation Detected: In patients with advanced or metastatic colorectal cancer, current literature suggests that tumors with a mutation in exons 2, 3, or 4 of the KRAS gene are essentially insensitive to anti-EGFR therapies such as cetuximab or panitumumab.  The presence of KRAS exon 2 (i.e. codons 12 and 13), exon 3 (i.e. codons 59 and 61), or exon 4 (i.e. codons 117 and 146) mutations in the tumor specimen suggests that the patient may not be a candidate for such targeted anti-EGFR therapy and is not recommended to receive treatment with cetuximab or panitumumab according to NCCN guidelines.
  • Point mutations in the KRAS gene which result in amino acid substitutions and produce a constitutively active form of KRAS are observed in a variety of human malignancies, including colorectal carcinoma, pancreatic carcinoma, and lung adenocarcinoma
  • The presence of mutations in KRAS exon 2 (codons 12, 13), exon 3 (codons 59, 61), or exon 4 (codons 117, 146) has been shown to be predictive of lack of response to anti-epidermal growth factor receptor (EGFR) therapy in metastatic colorectal carcinoma
  • Mutations within these codons represent approximately 99% of the KRAS mutations observed in colorectal cancer as listed in the Catalogue of Somatic Mutations in Cancer (COSMIC) database.

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