Hereditary spherocytosis laboratory findings

Jump to navigation Jump to search

Hereditary spherocytosis Microchapters

Home

Patient Information

Overview

Historical Perspective

Classification

Pathophysiology

Causes

Differentiating Hereditary spherocytosis from other Diseases

Epidemiology and Demographics

Risk Factors

Screening

Natural History, Complications and Prognosis

Diagnosis

Diagnostic Study of Choice

History and Symptoms

Physical Examination

Laboratory Findings

Electrocardiogram

Chest X Ray

CT

MRI

Echocardiography or Ultrasound

Other Imaging Findings

Other Diagnostic Studies

Treatment

Medical Therapy

Surgery

Primary Prevention

Secondary Prevention

Cost-Effectiveness of Therapy

Future or Investigational Therapies

Case Studies

Case #1

Hereditary spherocytosis laboratory findings On the Web

Most recent articles

Most cited articles

Review articles

CME Programs

Powerpoint slides

Images

American Roentgen Ray Society Images of Hereditary spherocytosis laboratory findings

All Images
X-rays
Echo & Ultrasound
CT Images
MRI

Ongoing Trials at Clinical Trials.gov

US National Guidelines Clearinghouse

NICE Guidance

FDA on Hereditary spherocytosis laboratory findings

CDC on Hereditary spherocytosis laboratory findings

Hereditary spherocytosis laboratory findings in the news

Blogs on Hereditary spherocytosis laboratory findings

Directions to Hospitals Treating Hereditary spherocytosis

Risk calculators and risk factors for Hereditary spherocytosis laboratory findings

Editor-In-Chief: C. Michael Gibson, M.S., M.D. [1]

Overview

Laboratory Findings

Initial testing

  • Neonates
  • The evaluation of a neonate with suspected HS depends on whether a parent (or both parents) are known to have HS.
  • If an infant with hyperbilirubinemia has a known family history of HS, then the likelihood of HS is high, and we rely heavily on the RBC indices. As noted above, an MCHC ≥36 g/dL is highly suggestive of HS.
  • If an infant with hyperbilirubinemia or hemolytic anemia does not have a known family history of HS, then a number of other possible diagnoses must be considered.
  • Appropriate therapy should not be delayed while determining the underlying cause; likewise, the importance of making the diagnosis of HS should be emphasized regardless of the management interventions needed.
  • Hemolytic anemia with a negative Coombs test and a high MCHC (eg, >36 g/dL) is consistent with HS but must be considered in the context of the entire clinical picture.
  • It may be difficult to assess spherocytes on the peripheral blood smear in a neonate, either because neonates with HS may have fewer spherocytes or because spherocytic cells are often present after birth in neonates without HS.[3] If the infant is well, it is reasonable to postpone testing until approximately six months of age or older, at which time the RBC morphology will be easier to assess.[4]
  • Older children and adults
  • Hereditary spherocytosis may be suspected in a patient of any age who has evidence of hemolysis (eg, elevated serum LDH, elevated indirect bilirubin, reduced haptoglobin, increased reticulocyte count) or hemolytic anemia that is Coombs-negative and not explained by another condition.
  • HS may also be suspected in an individual who presents with a complication of hemolysis, such as splenomegaly, pigmented gallstones, or an abrupt drop in hemoglobin level when the bone marrow cannot compensate for hemolysis (eg, during a viral illness, pregnancy, or other condition). In such cases, a CBC will be obtained and RBC indices will be available; the reticulocyte count should also be measured if not done already.
  • Evidence consistent with HS as the likely diagnosis in an older child or adult include the following:
    • Positive family history of HS, although this is not always present as some cases arise as new mutations and not all individuals will have a complete family history available.
    • Chronic hemolytic anemia, although in mild cases, there may be chronic compensated hemolysis without anemia.
    • Jaundice and/or splenomegaly, although these may be absent if the hemolysis is mild.
    • Spherocytes on the peripheral blood smear. The percentage of spherocytes is variable. The typical reticulocyte count in older children and adults with HS is approximately 5 to 20 percent, but it may be as high as 20 to 30 percent in severe cases. Certain genetic defects have been associated with specific spherocyte morphologies, although the diagnostic value of these findings has not been rigorously tested.[5][6][7][8]
    • Pincered or notched spherocytesBand 3 deficiency
    • Acanthocytic spherocytes – Spectrin deficiency
    • Dense and irregularly shaped cells – Spectrin/ankyrin deficiency
    • Elliptocytic spherocytes – Spherocytic elliptocytosis
    • RBC indices consistent with spherocytosis (eg, MCHC >36 g/dL; normal to slightly low MCV). The MCV and red cell distribution width (RDW) may be increased by greater degrees of reticulocytosis in older children and adults; thus, the MCHC is the most useful of the RBC indices. The combination of increased MCHC and increased RDW further improves diagnostic performance.[9] If reticulocyte indices are available, a higher-than-average reticulocyte MCHC and a low reticulocyte MCV are also consistent with HS.[10]
  • In cases that are unclear or if additional diagnostic confirmation is needed, specialized testing can be pursued.
  • Confirmatory tests
  • A number of tests are available for confirming the diagnosis of HS. We perform confirmatory testing in all cases, although some experts may omit this testing, especially in resource-limited settings and/or if there are classic clinical findings in an individual with a known family history of HS.
  • Available tests include the following:
  • EMA binding
  • If specialized testing is indicated, EMA binding is our preferred test. EMA (eosin-5-maleimide) is an eosin-based fluorescent dye that binds to RBC membrane proteins, especially band 3 and Rh-related proteins.[11]
  • The mean fluorescence of EMA-labeled RBCs from individuals with HS is lower than controls, and this reduction in fluorescence can be detected in a flow cytometry-based assay. Two case series of individuals with HS have found the EMA fluorescence in individuals with HS to be approximately two-thirds that of controls [97,98]. Samples can be stored and tested; one of the studies also analyzed the effect of delayed testing and found that samples stored for 24 hours in the darkness gave similar results to those tested immediately.[12]
  • Advantages of EMA binding include its high sensitivity and specificity; rapid turnaround time (approximately two hours); and need for only a minimal amount of blood (a few microliters), which is especially advantageous for testing neonates [99-101].
  • In addition, EMA testing can be used to identify HS in a patient who has recently received a transfusion [102]. In various studies, the sensitivity and specificity of the test appear to be in the ranges of 93 to 96 and 93 to 99 percent, respectively.[13][14]
  • EMA binding may also be positive in some forms of hereditary elliptocytosis (HE; eg, hereditary pyropoikilocytosis [HPP] and Southeast Asian ovalocytosis [SAO]), in individuals with congenital dyserythropoietic anemia (CDA) type II, and/or in autoimmune hemolytic anemia. False negative results may be seen in mild cases of HS.
  • Osmotic fragility
  • If EMA binding is not available, osmotic fragility testing (OFT) is another useful specialized test for HS. In this test, RBCs are incubated in hypotonic buffered salt solutions of various osmolarities, and the fraction of hemoglobin released (due to hemolysis) is measured.
  • The test takes advantage of the increased sensitivity of spherocytes to hemolysis, which is due to their reduced surface area to volume (SA/V) ratio. Incubation of patient samples for 24 hours prior to testing may accentuate osmotic fragility and improve diagnostic yield.
  • The OFT has relatively low sensitivity and specificity. It fails to identify a significant number of individuals with HS, and, particularly in the newborn, it may be positive in other conditions including immune hemolytic anemia, hemolytic transfusion reactions, RBC enzyme defects such as glucose-6-phosphate dehydrogenase (G6PD) deficiency, and unstable hemoglobin variants.[15] In one series of 86 individuals with HS, only 57 (66 percent) had positive osmotic fragility testing.[16]
  • Glycerol lysis
  • The glycerol lysis test (GLT) and the acidified GLT (AGLT) are modifications of the OFT that add glycerol (in the GLT) or glycerol plus a sodium phosphate (to lower the pH to 6.85, in the AGLT) to the hypotonic buffered salt solutions in which the patient's RBCs are incubated.[17][18]
  • Like the OFT, these tests may also be positive in acquired spherocytosis conditions such as AIHA.
  • The "pink test" is a modification of the GLT in which the final extent of hemolysis is measured in a blood sample incubated in the glycerol solution at pH 6.66. A further modification has been proposed (the direct pink test) in which the test sample is obtained from fingerprick (or heel puncture in newborns), rather than venipuncture, and incubated directly in the glycerol solution; this requires only a few microliters of blood.[19]
  • Cryohemolysis
  • In the cryohemolysis test, RBCs are suspended in a hypertonic solution, briefly heated to 37°C, then cooled to 4°C for 10 minutes.[20]
  • The ease of performance and the wide separation in degree of hemolysis between spherocytes and normal cells are two attractive features of this test.[21]
  • This test has limited availability in the United States.
  • Plasma membrane electrophoresis[22][23]

References

  1. Farias, Mariela Granero (2017). "Advances in laboratory diagnosis of hereditary spherocytosis". Clinical Chemistry and Laboratory Medicine (CCLM). 55 (7). doi:10.1515/cclm-2016-0738. ISSN 1437-4331.
  2. Christensen RD, Yaish HM, Gallagher PG (2015). "A pediatrician's practical guide to diagnosing and treating hereditary spherocytosis in neonates". Pediatrics. 135 (6): 1107–14. doi:10.1542/peds.2014-3516. PMC 4444801. PMID 26009624.
  3. King MJ, Garçon L, Hoyer JD, Iolascon A, Picard V, Stewart G; et al. (2015). "ICSH guidelines for the laboratory diagnosis of nonimmune hereditary red cell membrane disorders". Int J Lab Hematol. 37 (3): 304–25. doi:10.1111/ijlh.12335. PMID 25790109.
  4. Bolton-Maggs PH, Langer JC, Iolascon A, Tittensor P, King MJ, General Haematology Task Force of the British Committee for Standards in Haematology (2012). "Guidelines for the diagnosis and management of hereditary spherocytosis--2011 update". Br J Haematol. 156 (1): 37–49. doi:10.1111/j.1365-2141.2011.08921.x. PMID 22055020.
  5. Jarolim P, Murray JL, Rubin HL, Taylor WM, Prchal JT, Ballas SK; et al. (1996). "Characterization of 13 novel band 3 gene defects in hereditary spherocytosis with band 3 deficiency". Blood. 88 (11): 4366–74. PMID 8943874.
  6. Hassoun H, Vassiliadis JN, Murray J, Njolstad PR, Rogus JJ, Ballas SK; et al. (1997). "Characterization of the underlying molecular defect in hereditary spherocytosis associated with spectrin deficiency". Blood. 90 (1): 398–406. PMID 9207476.
  7. Becker PS, Tse WT, Lux SE, Forget BG (1993). "Beta spectrin kissimmee: a spectrin variant associated with autosomal dominant hereditary spherocytosis and defective binding to protein 4.1". J Clin Invest. 92 (2): 612–6. doi:10.1172/JCI116628. PMC 294892. PMID 8102379.
  8. Coetzer TL, Lawler J, Liu SC, Prchal JT, Gualtieri RJ, Brain MC; et al. (1988). "Partial ankyrin and spectrin deficiency in severe, atypical hereditary spherocytosis". N Engl J Med. 318 (4): 230–4. doi:10.1056/NEJM198801283180407. PMID 2961992.
  9. Michaels LA, Cohen AR, Zhao H, Raphael RI, Manno CS (1997). "Screening for hereditary spherocytosis by use of automated erythrocyte indexes". J Pediatr. 130 (6): 957–60. PMID 9202619.
  10. Bolton-Maggs PH, Langer JC, Iolascon A, Tittensor P, King MJ, General Haematology Task Force of the British Committee for Standards in Haematology (2012). "Guidelines for the diagnosis and management of hereditary spherocytosis--2011 update". Br J Haematol. 156 (1): 37–49. doi:10.1111/j.1365-2141.2011.08921.x. PMID 22055020.
  11. Ciepiela O, Kotuła I, Górska E, Stelmaszczyk-Emmel A, Popko K, Szmydki-Baran A; et al. (2013). "Delay in the measurement of eosin-5′-maleimide (EMA) binding does not affect the test result for the diagnosis of hereditary spherocytosis". Clin Chem Lab Med. 51 (4): 817–23. doi:10.1515/cclm-2012-0240. PMID 23023797.
  12. Ciepiela O, Kotuła I, Górska E, Stelmaszczyk-Emmel A, Popko K, Szmydki-Baran A; et al. (2013). "Delay in the measurement of eosin-5′-maleimide (EMA) binding does not affect the test result for the diagnosis of hereditary spherocytosis". Clin Chem Lab Med. 51 (4): 817–23. doi:10.1515/cclm-2012-0240. PMID 23023797.
  13. King MJ, Behrens J, Rogers C, Flynn C, Greenwood D, Chambers K (2000). "Rapid flow cytometric test for the diagnosis of membrane cytoskeleton-associated haemolytic anaemia". Br J Haematol. 111 (3): 924–33. PMID 11122157.
  14. Kar R, Mishra P, Pati HP (2010). "Evaluation of eosin-5-maleimide flow cytometric test in diagnosis of hereditary spherocytosis". Int J Lab Hematol. 32 (1 Pt 2): 8–16. doi:10.1111/j.1751-553X.2008.01098.x. PMID 18782334.
  15. King MJ, Garçon L, Hoyer JD, Iolascon A, Picard V, Stewart G; et al. (2015). "ICSH guidelines for the laboratory diagnosis of nonimmune hereditary red cell membrane disorders". Int J Lab Hematol. 37 (3): 304–25. doi:10.1111/ijlh.12335. PMID 25790109.
  16. Cynober T, Mohandas N, Tchernia G (1996). "Red cell abnormalities in hereditary spherocytosis: relevance to diagnosis and understanding of the variable expression of clinical severity". J Lab Clin Med. 128 (3): 259–69. PMID 8783633.
  17. Eber SW, Pekrun A, Neufeldt A, Schröter W (1992). "Prevalence of increased osmotic fragility of erythrocytes in German blood donors: screening using a modified glycerol lysis test". Ann Hematol. 64 (2): 88–92. PMID 1554800.
  18. Stoya G, Gruhn B, Vogelsang H, Baumann E, Linss W (2006). "Flow cytometry as a diagnostic tool for hereditary spherocytosis". Acta Haematol. 116 (3): 186–91. doi:10.1159/000094679. PMID 17016037.
  19. Judkiewicz L, Szczepanek A, Bugała I, Bartosz G (1987). "Modified end-point glycerol hemolysis assay as a screening test for hereditary spherocytosis that requires no venipuncture". Am J Hematol. 26 (1): 89–91. PMID 3631064.
  20. Streichman S, Gescheidt Y (1998). "Cryohemolysis for the detection of hereditary spherocytosis: correlation studies with osmotic fragility and autohemolysis". Am J Hematol. 58 (3): 206–12. PMID 9662272.
  21. Romero RR, Poo JL, Robles JA, Uriostegui A, Vargas F, Majluf-Cruz A (1997). "Usefulness of cryohemolysis test in the diagnosis of hereditary spherocytosis". Arch Med Res. 28 (2): 247–51. PMID 9204617.
  22. Olga Ciepiela (2018). "Old and new insights into the diagnosis of hereditary spherocytosis". Annals of translational medicine. 6 (17): 339. doi:10.21037/atm.2018.07.35. PMID 30306078. Unknown parameter |month= ignored (help)
  23. Paola Bianchi, Elisa Fermo, Cristina Vercellati, Anna P. Marcello, Laura Porretti, Agostino Cortelezzi, Wilma Barcellini & Alberto Zanella (2012). "Diagnostic power of laboratory tests for hereditary spherocytosis: a comparison study in 150 patients grouped according to molecular and clinical characteristics". Haematologica. 97 (4): 516–523. doi:10.3324/haematol.2011.052845. PMID 22058213. Unknown parameter |month= ignored (help)

Template:WH Template:WS