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Test Code LABRBCME Red Blood Cell Membrane Evaluation, Blood

Additional Codes

Test Name in EPIC EPIC Test Code Mnemonic Mayo Test ID
RED BLOOD CELL MEMBRANE EVALUATION LABRBCME RBCME RBCME

 

Useful For

Investigation of suspected red cell membrane disorders such as hereditary spherocytosis or hereditary pyropoikilocytosis

Profile Information

Test ID Reporting Name Available Separately Always Performed
FRAGO Osmotic Fragility Yes, (Order FRAG) Yes
SCTRL Shipping Control Vial No Yes
HSEV Spherocytosis Interpretation No Yes
BND3 Band 3 Fluorescence Staining, RBC No Yes
SMPB Peripheral Blood Smear Review No Yes

Testing Algorithm

Osmotic fragility and eosin-5-maleimide (EMA) binding (Band3) flow cytometry testing will always be performed. A normal shipping control is necessary to exclude false-positive results due to preanalytical artifact. Testing will be canceled if no shipping control is received or if the shipping control is abnormal. A consultative interpretation will be provided.

Method Name

HSEV: Consultative Interpretation

FRAGO: Osmotic Lysis

BND3: Flow Cytometry

SMPB: Consultant Review

Reporting Name

RBC Membrane Evaluation, B

Specimen Type

Control
Whole Blood EDTA
Whole Blood Slide


Shipping Instructions


Specimens must arrive within 72 hours of draw.



Necessary Information


Include recent transfusion information.

 

Include most recent CBC results.



Specimen Required


A whole blood EDTA specimen, an EDTA control specimen, and 2 well-made peripheral blood smears (Wright stained or fixed in absolute methanol) are required for testing.

 

Patient:

Specimen Type: Blood

Container/Tube: Lavender top (EDTA)

Specimen Volume: 4 mL

Collection Instructions:

1. Immediately refrigerate specimen after draw. Refrigerate at 0 to 4° C. Do not freeze. Freezing causes sample lysis, and tests will not be performed on hemolyzed specimens.

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

3. Rubber band patient specimen and control vial together. Control must accompany the patient sample at all times to ensure the reliability of testing results.

4. Be sure specimen and control are stored and transported together at refrigerate temperature, carefully following proper handling and shipping instructions.

 

Patient:

Specimen Type: Slides

Container/Tube: Blood smears

Specimen Volume: 2 well-made peripheral blood smears

Collection Instructions: Collect 2 well-made peripheral blood smears (Wright stained or fixed in absolute methanol).

 

Normal Shipping Control:

Specimen Type: Whole blood

Container/Tube: Lavender top (EDTA)

Specimen Volume: 4 mL

Collection Instructions:

1. Draw a control specimen from a normal (healthy), unrelated, nonsmoking person at the same time as the patient.

2. Label clearly on outermost label normal control.

3. Immediately refrigerate specimen after draw. Refrigerate at 0 to 4° C. Do not freeze. Freezing causes sample lysis, and tests will not be performed on hemolyzed specimens.

4. Send specimen in original tube. Do not aliquot.

5. Rubber band patient specimen and control vial together. Control must accompany the patient sample at all times to ensure the reliability of testing results.


Specimen Minimum Volume

2 mL

Specimen Stability Information

Specimen Type Temperature Time
Control Refrigerated 72 hours
Whole Blood EDTA Refrigerated 72 hours
Whole Blood Slide Refrigerated

Reject Due To

Hemolysis

Mild reject; Gross reject

Lipemia

NA

Icterus

NA

Other

Clotted, frozen

Clinical Information

The functional red cell membrane is composed of a cholesterol and phospholipid bilayer anchored by integral proteins to an elastic cytoskeletal network. These interactions form the shape, deformability, and proper ion balance of the cell. Abnormalities in these moieties result in red blood cell membrane disorders.

 

Hereditary spherocytosis (HS) is a common membrane disorder that can be present in many ethnic groups. Its prevalence has been estimated at approximately 1 in 3,000 persons of Northern European ancestry. It is usually associated with visible spherocytes on the peripheral blood smear and can be associated with variable clinical features of hemolysis ranging from completely compensated to mild to severe.

 

Hereditary elliptocytosis (HE) is another fairly common and clinically variable disorder that can range from normal RBC indices in the large majority of cases to rare patients with moderate to severe anemia.

 

Common hereditary elliptocytosis (CHE) is characterized by the presence of elliptocytes on the peripheral blood smear. Mutations associated with HE have been reported in widely variable ethnicities with greater prevalence in populations overlapping the malaria belt.

 

Hereditary pyropoikilocytosis (HPP) is best classified as a severe form of hereditary elliptocytosis. It is uncommon and presents in early childhood as a severe hemolytic anemia. These disorders are associated with marked poikilocytosis on the peripheral blood smear.(1,2) Red cell membrane disorders can result from abnormalities involving several red cell membrane proteins, such as band 3, alpha and beta spectrin, protein 4.1, protein 4.2, glycophorin C, and ankyrin.

 

Most often, red cell membrane disorders are diagnosed in childhood, adolescence, or early adult life. The diagnosis of HS is usually made by a combination of patient and family history, laboratory evidence of hemolysis, and review of a peripheral blood smear. The osmotic fragility (OF) test is usually markedly abnormal in HS cases. However, factors such as age, iron status, and medications can affect the OF test. The OF test is nonspecific and can be increased in acquired disorders such as autoimmune hemolytic anemia. Coombs testing should be negative prior to ordering this test.

 

The addition of eosin-5-maleimide (EMA) binding (Band3) flow cytometry to this profile increases specificity if a typical moderately decreased pattern is present. Hereditary pyropoikilocytosis can have normal or only mildly increased OF results and often displays a very dim and sometimes dual peak pattern with EMA-binding testing. Common hereditary elliptocytosis cases are not discriminated from normal patients in either OF and EMA binding (band3) testing and this profile does not add confirmatory information for HE.

Reference Values

≥12 months:

0.50 g/dL NaCl (unincubated): 3-53% hemolysis

0.60 g/dL NaCl (incubated): 14-74% hemolysis

0.65 g/dL NaCl (incubated): 4-40% hemolysis

0.75 g/dL NaCl (incubated): 1-11% hemolysis 

An interpretive report will be provided.

 

Reference values have not been established for patients who are <12 months of age.

Interpretation

An interpretive report will be provided.

Cautions

This test is not useful for hereditary elliptocytosis.

Interfering factors:

-Recent transfusion

-Oral contraceptives

-H2 blockers

Infrequently, other hemolytic disorders may also be associated with positive osmotic fragility results, as in patients with congenital nonspherocytic hemolytic anemia due to glucose-6-phosphate dehydrogenase or pyruvate kinase deficiency. Eosin-5-maleimide (EMA) binding (band 3) results can be masked by reticulocytosis. Results must be interpreted within the clinical, familial, and peripheral blood smear findings.

 

Patients with an immunohemolytic anemia, or who have recently received a blood transfusion may also have increased RBC lysis.

 

The shipping control specimen is required to adequately interpret these cases, as temperature extremes can increase fragility of the specimen. 

 

Resulting Cautions:

Osmotic fragility results will be reported if the shipping control is normal.

 

If the shipping control is abnormal and the osmotic fragility results are within normal range, the results will be reported; however, a comment will be added to the report indicating that the shipping control was not entirely satisfactory.

 

The test will be cancelled if the patient specimen and shipping control are both abnormal.

Method Description

Osmotic Fragility:

Specimens for erythrocyte osmotic fragility tests are anticoagulated with EDTA. Osmotic lysis is performed using sodium chloride (NaCl) solution, 0.50 g/dL. An incubated fragility test is performed following 24-hour incubation at 37° C at the following NaCl concentrations: 0.60, 0.65, and 0.75 g/dL. Results are reported and interpreted.(Larson CJ, Scheidt R, Fairbanks VF: The osmotic fragility test for hereditary spherocytosis: use of EDTA-anticoagulated blood stored at 4° C for up to 96 hours. Am Soc Clin Pathol Meeting Abstract, 1988; Larson CJ, Scheidt R, Fairbanks VF: The osmotic fragility test for hereditary spherocytosis: objective criteria for test interpretation. Am Soc Clin Pathol Meeting Abstract, 1988)

 

Band 3:

Eosin-5-maleimide (EMA) is a fluorescent dye that binds to Lys-430 of the extracellular loop of the band 3 protein. Using a 1-color flow cytometry method (number of events plotted against fluorescence), the fluorescent intensity of EMA-stained RBC, is assessed and compared to normal-value patients.(King MJ, Behrens J, Rogers C, et al: Rapid flow cytometric test for the diagnosis of membrane cytoskeletal associated hemolytic anemia. Br J Haematol 2000;111:924-933)

 

Peripheral Blood Smear Review:
A hematopathologist who is an expert in these disorders evaluates the slides and an interpretive report is issued.

Day(s) and Time(s) Performed

Monday through Saturday

Analytic Time

4 days (not reported on Saturday or Sunday)

Specimen Retention Time

7 days

Performing Laboratory

Mayo Medical Laboratories in Rochester

CPT Code Information

85557-Osmotic fragility

88184-Flow cytometry; first cell surface, cytoplasmic or nuclear marker x 1

85060-Morphology review

LOINC Code Information

Test ID Test Order Name Order LOINC Value
RBCME RBC Membrane Evaluation, B In Process

 

Result ID Test Result Name Result LOINC Value
83141 Band 3 Fluorescence Staining, RBC In Process
9064 Osmotic Fragility, RBC 34964-7
SCTRL Shipping Control Vial 40431-9
13065 Spherocytosis Interpretation 50595-8
37406 Peripheral Blood Smear Review 59465-5
37436 Reviewed By 18771-6
3306 Osmotic Fragility, 0.50 g/dL NaCl 23915-2
3307 Osmotic Fragility, 0.60 g/dL NaCl 23917-8
3308 Osmotic Fragility, 0.65 g/dL NaCl 23919-4
3309 Osmotic Fragility, 0.75 g/dL NaCl 30543-3
3310 Osmotic Fragility Comment 59466-3

Forms

1. Red Cell Membrane Disorders Information (T706) in Special Instruction

2. If not ordering electronically, complete, print, and send a Benign Hematology Test Request Form (T755) with the specimen (http://www.mayomedicallaboratories.com/it-mmfiles/benign-hematology-test-request-form.pdf)

Test Classification

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