Alpha Hemoglobin DNA Sequence

Interpretation

Guidelines

• Genetics and Solid Tumor Diagnostic Testing

Method

DNA sequencing of the coding regions and introns of both the alpha1 (HBA1) and alpha 2 globin (HBA2) genes in both directions to detect point mutations causing alpha globin variants and some types of alpha thalassemia.

This test detects alpha globin variants and non-deletional alpha thalassemia mutations (e.g. Hb Constant Spring). The coding regions and introns of both HBA1 and HBA2 genes are sequenced in bidirectionally. The reference mRNA sequences are HBA1 (NM_000558.3) and HBA2 (NM_000517.4) with nucleotide 1 corresponding to the A of the AUG initiation codon (nucleotide 38 of both reference sequences) and codon 1 corresponding to the valine encoded by GTG (nucleotides 41-43 of both reference sequences). The sensitivity of this test for detecting nucleotide substitutions, small insertions and deletions and in the alpha 1 (HBA1) and alpha 2 (HBA2) genes is theoretically >98%. This test will not detect mutations that lie outside of the sequenced regions, nor large HBA1 and HBA2 gene deletions (e.g. –a3.7, – –SEA). This test was developed and its performance characteristics determined by the Department of Laboratory Medicine at the University of Washington.

Reference Range

See individual components

Ref. Range Notes

No mutation detected.

References

• Huisman, T. H. J., Carver, M. F. H., and Efremov, G. D. (1996). A Syllabus of Hemoglobin Variants (Augusta, GA: The Sickle Cell Anemia Foundation).
• Higgs DR, et al. A review of the molecular genetics of the human alpha-globin gene cluster. Blood 1989, 73:1081-104. 2649166 .
• Skogerboe KJ, et al. Screening for alpha-thalassemia. Correlation of hemoglobin H inclusion bodies with DNA-determined genotype. Arch Pathol Lab Med 1992, 116:1012-8. 1329692
• Molchanova TP, Pobedimskaya DD, and Postnikov YuV. A simplified procedure for sequencing amplified DNA containing the alpha 2- or alpha 1-globin gene. Hemoglobin 1994, 18:251-5. 7928384
• Old JM. Screening and genetic diagnosis of haemoglobin disorders. Blood Rev 2003, 17:43-53. 12490210
• Chui DH, Fucharoen S, and Chan V. Hemoglobin H disease: not necessarily a benign disorder. Blood 2003, 101:791-800. 12393486
• Chan V, et al. Molecular defects in Hb H hydrops fetalis. Br J Haematol 1997, 96:224-8. 9029003
• Globin Gene Server: http://globin.cse.psu.edu/

Ordering & Collection

Specimen Type

Blood/Cultured amniocytes or chorionic villus cells/Extracted DNA from blood, chorionic villi, and amniocyte. Direct chorionic villi, amniocyte, or amniotic fluid testing require Genetics Director approval. Please call the lab at 206-598-7021

Collection

Acceptable:

1. Whole blood:5 mL lavender top (EDTA) tube or yellow (ACD) top tube or 2 mL microtainer lavender top tube
   
   
2. Extracted DNA from blood, chorionic villi, and amniocytes: 500 ng (concentration >10 ng/uL)
   
   
3. Cultured amniocytes/chorionic villi: MCC is required for testing fetal samples. See MCC OLTG.
   
   
4. Also acceptable, but requires the Genetics Director's approval and a backup culture. Direct chorionic villi and/or TISSUE: Send 20mg of tissue in a sterile tube or RPMI culture media
   
   *NOTE: If a fetal sample (cultured amniocytes or chorionic villi) was received, add MCC to the order. Prenatal testing requires concomitant testing for maternal cell contamination (see Online Test Guide, MCC for ordering and specimen requirements). See Special Instructions.

Unacceptable: Heparin green top tubes, buccal swab

Forms & Requisitions

Genetics Requisition

Handling Instructions

SPS specimen handling:

1. Whole blood sample: store in the refrigerator
   
   
2. Cultured amniocytes/chorionic villi: store at room temperature. Call the Genetics lab upon receipt (206)598-7021.
   
   
3. Extracted DNA: store in the refrigerator

Quantity

Requested: Entire specimen
Minimum: Blood: 1 mL. If volume is less than 1mL, do not cancel. Send to Genetics lab. Confluent cultured cells: One (1) T25 flask. Extracted DNA: 250 ng

Processing

Receiving Instructions

If fetal tissue (cultured amniocytes or chorionic villi) was received for prenatal testing, consultation with the laboratory is required. Please notify the Genetics lab about prenatal studies via email at geneticshelp@uw.edu or call 206-598-7021.

For clients outside of UW, please include the most recent CBC and Hb electrophoresis result/s (if available), and/or any relevant clinical history.

Misc Sendout

Performance

Lab Department

Genetics(GEN)

Frequency

Performed weekly. Results within 3 weeks.

Available STAT?

No

Performing Location(s)

UW-MT	Genetics Attention: Genetics Lab Clinical lab, Room NW220 University of Washington Medical Center 1959 NE Pacific Street Seattle, WA 98195 Tel: 206-598–6429 M–F (7:30 AM–4:00 PM) Fax: 206-616-4584 Lab email: cgateam@uw.edu Tel (EXOME only): 206-543-0459	Faculty Jillian Buchan, PhD, FACMG Runjun Kumar, MD, PhD Regina Kwon, MD, MPH Christina Lockwood, PhD, DABCC, DABMGG Abbye McEwen, MD, PhD Colin Pritchard, MD, PhD Vera Paulson, MD, PhD Eric Konnick, MD, MS He Fang, PhD

Billing & Coding

CPT Codes

81259

LOINC

60540-2

Interfaced Order Code

UOW2161

Billing Comments

For pricing information, contact Client Support Services 206-520-4600 or 800-713-5198.