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Order Code GNHMB Hemophilia B, F9 Gene, Next-Generation Sequencing, Varies


Ordering Guidance


For male patients, this test should only be considered if clinical and family history, initial coagulation screens, and/or initial activity tests indicate a diagnosis of hemophilia B. For female patients, this test should only be considered if there is a confirmed diagnosis of hemophilia B in a family member or the patient has abnormally low factor IX (FIX) activity.

 

This test does not measure FIX activity levels. For assessment of FIX activity, F_9 / Coagulation Factor IX Activity Assay, Plasma.

 

For individuals with bleeding symptoms and no known personal or family history of hemophilia B, consider ALBLD / Bleeding Diathesis Profile, Limited, Plasma or the specific factor assays.

 

If genetic testing for hereditary bleeding disorders using a larger panel is desired, both a 6-gene focused bleeding panel and a 25-gene comprehensive bleeding panel are available. For more information see GNBLF / Bleeding Disorders, Focused Gene Panel, Next-Generation Sequencing, Varies or GNBLC / Bleeding Disorders, Comprehensive Gene Panel, Next-Generation Sequencing, Varies.

 

Testing for the F9 gene as part of a customized panel is available. For more information see CGPH / Custom Gene Panel, Hereditary, Next-Generation Sequencing, Varies.

 

Targeted testing for familial variants (also called site-specific or known variants testing) is available for the F9 gene. See FMTT / Familial Variant, Targeted Testing, Varies. To obtain more information about this testing option, call 800-533-1710.



Additional Testing Requirements


All prenatal specimens must be accompanied by a maternal blood specimen; order MATCC / Maternal Cell Contamination, Molecular Analysis, Varies on the maternal specimen as this must be a different order number than the prenatal specimen.



Shipping Instructions


Specimen preferred to arrive within 96 hours of collection.



Necessary Information


Hemophilia B Patient Information is required. Testing may proceed without the patient information; however, the information aids in providing a more thorough interpretation. Ordering providers are strongly encouraged to fill out the form and send with the specimen.



Specimen Required


Patient Preparation: A previous bone marrow transplant from an allogenic donor will interfere with testing. For instructions for testing patients who have received a bone marrow transplant, call 800-533-1710.

 

Submit only 1 of the following specimens:

 

Specimen Type: Whole blood

Container/Tube:

Preferred: Lavender top (EDTA)

Acceptable: Yellow top (ACD)

Specimen Volume: 3 mL

Collection Instructions:

1. Invert several times to mix blood.

2. Send whole blood specimen in original tube. Do not aliquot.

Specimen Stability Information: Ambient (preferred) 4 days/Refrigerated

 

Prenatal Specimens

Due to its complexity, consultation with the laboratory is required for all prenatal testing; call 800-533-1710 to speak to a genetic counselor. 

 

Specimen Type: Amniotic fluid

Container/Tube: Amniotic fluid container

Specimen Volume: 20 mL

Specimen Stability Information: Refrigerated (preferred)/Ambient

Additional information:

1. A separate culture charge will be assessed under CULAF / Culture for Genetic Testing, Amniotic Fluid.

2. All prenatal specimens must be accompanied by a maternal blood specimen; order MATCC / Maternal Cell Contamination, Molecular Analysis, Varies on the maternal specimen.

 

Specimen Type: Chorionic villi

Container/Tube: 15-mL tube containing 15 mL of transport media

Specimen Volume: 20 mg

Specimen Stability Information: Refrigerated

Additional Information: 1. A separate culture charge will be assessed under CULFB / Fibroblast Culture for Biochemical or Molecular Testing.

2. All prenatal specimens must be accompanied by a maternal blood specimen; order MATCC / Maternal Cell Contamination, Molecular Analysis, Varies on the maternal specimen.

 

Acceptable:

Specimen Type: Confluent cultured cells

Container/Tube: T-25 flask

Specimen Volume: 2 Flasks

Collection Instructions: Submit confluent cultured cells from another laboratory.

Specimen Stability Information: Ambient (preferred)/Refrigerated

Additional Information:

All prenatal specimens must be accompanied by a maternal blood specimen; order MATCC / Maternal Cell Contamination, Molecular Analysis, Varies on the maternal specimen.


Forms

1. Hemophilia B Patient Information (T518) is required.

2. New York Clients-Informed consent is required. Document on the request form or electronic order that a copy is on file. The following documents are available:

-Informed Consent for Genetic Testing (T576)

-Informed Consent for Genetic Testing (Spanish) (T826)

3. If not ordering electronically, complete, print, and send an Coagulation Test Request (T753) with the specimen.

Useful For

Confirming a clinical diagnosis of hemophilia B in affected male patients with the identification of a disease-causing variant in the F9 gene

 

Determining the disease-causing alteration within the F9 gene to delineate the underlying molecular defect in a male patient with a laboratory diagnosis of hemophilia B

 

Identifying the causative alteration for prognostic and genetic counseling purposes

 

Assessing hemophilia B carrier status for female patients with a family history of hemophilia B

 

Prenatal testing for hemophilia B when a familial F9 variant has been previously identified in a family member

Genetics Test Information

This test utilizes next-generation sequencing to detect single nucleotide and copy number variants in the F9 gene associated with hemophilia B (also known as factor IX deficiency). See Method Description for additional details.

 

Identification of a disease-causing variant may assist with diagnosis, prognosis, clinical management, recurrence risk assessment, familial screening, and genetic counseling for hemophilia B.

Reflex Tests

Test ID Reporting Name Available Separately Always Performed
CULFB Fibroblast Culture for Genetic Test Yes No
CULAF Amniotic Fluid Culture/Genetic Test Yes No
MATCC Maternal Cell Contamination, B Yes No

Testing Algorithm

The clinical workup for hemophilia B in symptomatic male patients should begin with special coagulation testing for factor IX (FIX) activity. Genetic testing is indicated if FIX activity is less than 40% of normal (Note: reference range may vary depending on the locally established reference range) and von Willebrand factor antigen testing is normal. For more information see Hemophilia Testing Algorithm.

 

FIX clotting activity does not correlate as well with bleeding severity in female patients, and therefore is unreliable in the detection of female carriers of hemophilia B. Carrier status is determined by identification of a heterozygous disease-causing variant in F9 by molecular genetic testing. For female patients with a suspected or confirmed diagnosis of hemophilia B in a family member, carrier testing is recommended as specified by the Hemophilia Carrier Testing Algorithm.(1-3)

 

Acquired (nongenetic) causes of hemophilia B that should be excluded prior to genetic testing include vitamin K deficiency autoimmune disorders, malignancy, and infections such as HIV and hepatitis B.

 

For prenatal specimens only:

Prenatal genetic testing is not routinely performed without the prior identification of a familial hemophilia alteration in an affected male relative or a female relative who is a confirmed carrier of the alteration. Requests for this prenatal testing without a known familial alteration are performed at the discretion of the Molecular Hematopathology Laboratory Director.

 

-If amniotic fluid (nonconfluent cultured cells) is received, amniotic fluid culture/genetic test will be added at an additional charge.

-If chorionic villus specimen (nonconfluent cultured cells) is received, fibroblast culture for genetic test will be added at an additional charge.

 

For any prenatal specimen that is received, maternal cell contamination testing will be performed at an additional charge.

Method Name

Sequence Capture and Targeted Next-Generation Sequencing (NGS) followed by Polymerase Chain Reaction (PCR) and Sanger Sequencing

Reporting Name

F9 Gene, Full Gene NGS

Specimen Type

Varies

Specimen Minimum Volume

Blood: 1 mL; Amniotic fluid: 10 mL; Other specimen types: see Specimen Required

Specimen Stability Information

Specimen Type Temperature Time Special Container
Varies Varies

Reject Due To

All specimens will be evaluated at Mayo Clinic Laboratories for test suitability.

Reference Values

An interpretive report will be provided.

Method Description

Next-generation sequencing (NGS) and/or Sanger sequencing are performed to test for the presence of variants in coding regions and intron/exon boundaries of the F9 gene, as well as some other regions that have known disease-causing variants. The human genome reference GRCh37/hg19 build was used for sequence read alignment. At least 99% of the bases are covered at a read depth over 30X. Sensitivity is estimated at above 99% for single nucleotide variants, above 94% for deletions-insertions (delins) less than 40 base pairs (bp), above 95% for deletions up to 75 bp, and insertions up to 47 bp. NGS and/or a polymerase chain reaction-based quantitative method is performed to test for the presence of deletions and duplications in the F9 gene.

 

There may be regions of the F9 gene that cannot be effectively evaluated by sequencing or deletion and duplication analysis as a result of technical limitations of the assay, including regions of homology, high guanine-cytosine (GC) content, and repetitive sequences.(Unpublished Mayo method)

 

The reference transcript for the F9 gene is NM_000133.4. Reference transcript numbers may be updated due to transcript re-versioning. Always refer to the final patient report for gene transcript information referenced at the time of testing. Confirmation of select reportable variants may be performed by alternate methodologies based on internal laboratory criteria.

Day(s) Performed

Varies

Performing Laboratory

Mayo Clinic Laboratories in Rochester

CPT Code Information

81238

88233-Tissue culture, skin, solid tissue biopsy (if appropriate)

88240-Cryopreservation (if appropriate)

88235-Amniotic fluid culture (if appropriate)

81265-Maternal cell contamination (if appropriate)