Test ID: XHIM
X-Linked Hyper IgM Syndrome, Blood
Secondary ID 
A test code used for billing and in test definitions created prior to November 2011
NY State Approved Indicates the status of NY State approval and if the test is orderable for NY State clients.
Useful For Suggests clinical disorders or settings where the test may be helpful
Screening for XL-HIGM or CD40L deficiency, primarily in male patients <10 years of age
Ascertaining XL-HIGM carrier status in females of child-bearing age <45 years of age
Method Name A short description of the method used to perform the test
Flow Cytometry
Reporting Name A shorter/abbreviated version of the Published Name for a test; an abbreviated test name
Aliases Lists additional common names for a test, as an aid in searching
CD40 Activation
CD40 Ligand
Memory Cells
T cell Activation
IgM (Immunoglobulin)
Specimen Type Describes the specimen type needed for testing
Specimen Required Defines the optimal specimen. This field describes the type of specimen required to perform the test and the preferred volume to complete testing. The volume allows automated processing, fastest throughput and, when indicated, repeat or reflex testing.
Specimen must arrive within 24 hours of draw. Send specimen Sunday through Thursday only. Draw and package specimen as close to shipping time as possible. Ship specimen overnight.
Container/Tube: Green top (sodium heparin)
Specimen Volume: 4 mL
Additional Information:
1. Ordering physician's name and phone number are required.
2. For serial monitoring, we recommend that specimen draws be performed at the same time of day.
Specimen Minimum Volume Defines the amount of specimen required to perform an assay once, including instrument and container dead space. Submitting the minimum specimen volume makes it impossible to repeat the test or perform confirmatory or perform reflex testing. In some situations, a minimum specimen volume may result in a QNS (quantity not sufficient) result, requiring a second specimen to be collected.
Reject Due To Identifies specimen types and conditions that may cause the specimen to be rejected
| Hemolysis | Mild OK; Gross reject |
| Lipemia | Mild OK; Gross reject |
| Icterus | NA |
| Other | NA |
Specimen Stability Information Provides a description of the temperatures required to transport a specimen to the laboratory. Alternate acceptable temperature(s) are also included.
| Specimen Type | Temperature | Time |
|---|---|---|
| WB Sodium Heparin | Ambient (preferred) | 72 hours |
| Refrigerated | 72 hours |
Clinical Information Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test
CD154 (CD40 ligand: CD40L) is required for the interaction of T cells and B cells as part of the normal adaptive immune response. Activation of T cells leads to the expression of the CD40L molecule on the cell surface. CD40L binds the CD40 receptor that is always present on B cells, monocytes, and macrophages (regardless of environmental conditions). This interaction of CD40L with CD40 is important in B-cell proliferation, differentiation, and class-switch recombination (isotype class-switching).
Patients with X-linked hyper-IgM (XL-HIGM) syndrome have defective CD40L expression on their activated helper CD4 T cells.(1,2) This leads to defective B-cell responses and the absence of immunoglobulin class-switching. These features are typified in these patients by a profound reduction or absence of isotype class-switched memory B cells (CD19+CD27+IgM-IgD-) with low or absent secreted IgG and IgA, and normal or elevated serum IgM levels.(1,2) Due to the impairment of T-cell function and macrophage activation, XL-HIGM patients are particularly prone to opportunistic infections with Pneumocystis jiroveci, Cryptosporidium, and Toxoplasma gondii.(1)
To date, more than 100 unique mutations of CD40LG, the gene that encodes CD40L, have been described, affecting the intracellular, transmembrane and, more commonly, extracellular domain containing the CD40-binding region.
A defect in surface expression of CD40L on activated CD4 T cells can be demonstrated using an anti-CD40L antibody and flow cytometry.(3,4) Since certain CD40LG mutations can maintain surface protein expression, albeit with loss of function, it is important to also evaluate CD40L-binding capacity to eliminate the possibility of false-negative results. A soluble recombinant, chimeric receptor protein, CD40-uIg, is incorporated into the assay, which assesses CD40L function by determining receptor-binding activity. Approximately 20% of XL-HIGM patients have activated CD4 T cells with normal surface expression of CD40L, but aberrant function.(4)
XL-HIGM is a severe type of primary immunodeficiency that affects males, and most patients are diagnosed within a few months to the first year of life. Females are typically carriers and asymptomatic. Consequently, this test is only indicated in young males (<10 years of age) or, to identify carriers, in females of child-bearing age (<45 years).
Reference Values Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.
Present
Interpretation Provides information to assist in interpretation of the test results
This is a qualitative assay; CD40L-protein expression and function is reported as present or absent. Absence of CD40L-protein expression and function is consistent with XL-HIGM. In females, the presence of 2 populations-normal and abnormal-is consistent with carrier status.
Most patients with XL-HIGM have absent or significantly reduced CD40L expression on their activated CD4 T cells. Patients with normal CD40L expression, but abnormal function, show an absence of binding with soluble chimeric CD40-uIg antibody, substantiating a diagnosis of XL-HIGM. Females who are carriers for this disease will show a typical bimodal pattern of CD40L expression, with 50% of the T cells lacking any CD40L expression. In the case of aberrant protein function, a similar profile will be obtained with the CD40-uIg antibody.
CD69 is a marker for T-cell activation and serves as a positive control; in the absence of induced CD69 expression on T cells, the presence of XL-HIGM cannot be assessed.
Cautions Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances
This test is typically not indicated in males >10 years of age or females beyond child-bearing age (>45 years). For questions about appropriate test selection, contact Mayo Medical Laboratories.
The test must be performed on fresh, heparinized whole blood cells for appropriate CD40L expression on activated CD4 T cells; specimen handling instructions must be followed. T-cell activation is variable on specimens tested between 48 and 72 hours after blood collection. These specimens will be analyzed and results will be reported after the laboratory director's review. Specimens received >72 hours after collection will be rejected and the assay will not be performed.
Patients with normal CD40L expression and normal receptor binding with the CD40-uIg antibody, yet presenting with the clinical phenotype of HIGM syndrome, should be evaluated for autosomal recessive forms of this syndrome including mutations in CD40, AICDA (AID), and UNG.(1,2) A combination of clinical features and laboratory analyses should permitidentification of an underlying HIGM defect, if present.
The other form of XL-HIGM involving mutations in the NEMO (NF-kappa B essential modulator) gene (official symbol IKBKG) can be easily discriminated from the CD40LG deficiency due to the unusual and characteristic clinical findings including abnormal development of ectoderm-derived skin structures and immunodeficiency with increased susceptibility to mycobacterial infections.(1,2)
Previous studies have reported mutations involving splice sites that result in the generation of small amounts of wild-type CD40L, associated with a milder clinical phenotype.(4) In these cases, the CD40-uIg fusion protein may show some binding, albeit at lower intensity and, therefore, the final molecular diagnosis depends on sequencing of the CD40LG gene.
This is not a confirmatory test for CD40L deficiency, and genetic testing must be performed to determine the specific mutation involved. Information about genetic testing for CD40L deficiency is available by contacting Mayo Medical Laboratories.
Clinical Reference Provides recommendations for further in-depth reading of a clinical nature
1. Etzioni A, Ochs HD: The hyper IgM syndrome-an evolving story. Pediatr Res 2004:56(4):519-525
2. Durandy A, Peron S, Fischer A: Hyper-IgM syndromes. Curr Opin Rheumatol 2006;18(4):369-376
3. Lee WI, Torgerson TR, Schumacher MJ, et al: Molecular analysis of a large cohort of patients with the hyper immunoglobulin M (IgM) syndrome. Blood 2005;105(5):1881-1890
4. Seyama K, Nonoyama S, Gangsaas I, et al: Mutations of the CD40 ligand gene and its effect on CD40 ligand expression in patients with X-linked hyper IgM syndrome. Blood 1998;92:2421-2434
Method Description Describes how the test is performed and provides a method-specific reference
The assay measures the expression of CD40L on activated CD4 T cells. Heparinized whole blood is incubated with phorbol myristate acetate (PMA) and ionomycin (calcium ionophore) for lymphocyte activation. The red blood cells are lysed and the remaining white blood cells are stained with a 4-color panel of antibodies on a single platform. The assay involves 4 tubes, which include an unstimulated control for both the CD40L and CD40-uIg antibodies. CD69 expression is measured as a positive control for appropriate T-cell activation. A combination of CD3, CD8, CD154 (CD40L), and CD40-uIg antibodies enables assessment of CD40L expression and binding (with CD40-uIg) on total T cells (CD3+), suppressor T cells (CD3+CD8+), and helper T cells (CD3+CD8-). A normal, healthy control will be included with each experiment to ensure the optimal performance of the assay. (O'Gorman MR, Zaas D, Paniagua M, et al: Development of a rapid whole blood flow cytometry procedure for the diagnosis of X-linked hyper-IgM syndrome patients and carriers. Clin Immunol Immunopathol 1997 November;85[2]:179-181; and Mayo unpublished method)
Day(s) and Time(s) Test Performed Outlines the days and times the test is performed. This field reflects the day and time the sample must be in the testing laboratory to begin the testing process and includes any specimen preparation and processing time required before the test is performed. Some tests are listed as continuously performed, which means assays are performed several times during the day.
Monday through Friday
Do not send specimen after Thursday. Specimen must be received by 10 a.m. on Friday.
Analytic Time Defines the amount of time it takes the laboratory to setup and perform the test. This is defined in number of days. The shortest interval of time expressed is "same day/1 day," which means the results may be available the same day that the sample is received in the testing laboratory. One day means results are available 1 day after the sample is received in the laboratory.
Maximum Laboratory Time Defines the maximum time from specimen receipt at Mayo Medical Laboratories until the release of the test result
Specimen Retention Time Outlines the length of time after testing that a specimen is kept in the laboratory before it is discarded
Performing Laboratory Location The location of the laboratory that performs the test
Test Classification Provides information regarding the medical device classification for laboratory test kits and reagents. Tests may be classified as cleared or approved by the US Food and Drug Administration (FDA) and used per manufacturer's instructions, or as products that do not undergo full FDA review and approval, and are then labeled as an Analyte Specific Reagent (ASR), Investigation Use Only (IUO) product, or a Research Use Only (RUO) product.
CPT Code Information Provides guidance in determining the appropriate Current Procedural Terminology (CPT) code(s) information for each test or profile. The listed CPT codes reflect Mayo Medical Laboratories interpretation of CPT coding requirements. It is the responsibility of each laboratory to determine correct CPT codes to use for billing.
88184-Flow cytometry, cell surface, cytoplasmic
88185 x 6-Each additional marker
LOINC® Code Information Provides guidance in determining the Logical Observation Identifiers Names and Codes (LOINC) values for the result codes returned for this test or profile.
| Result ID | Reporting Name | LOINC Code |
|---|---|---|
| 82964 | CD40 Ligand Expression | In Process |
| 29040 | CD40muIg (Function) | In Process |
| 23901 | Interpretation | 69052-9 |
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