Test ID: ADE
Autoimmune Dysautonomia Evaluation, Serum

Investigating idiopathic dysautonomic symptoms

Directing a focused search for cancer in patients with idiopathic dysautonomia

Investigating autonomic symptoms that appear in the course or wake of cancer therapy, and are not explainable by recurrent cancer or metastasis. Detection of autoantibodies in this profile helps differentiate autoimmune dysautonomia from the effects of chemotherapy.

If IFA (ANN1S/83381) patterns are indeterminate, paraneoplastic autoantibody Western blot is performed at an additional charge.

If IFA patterns suggest CRMP-5-IgG, CRMP-5-IgG Western blot is performed at an additional charge.

If IFA patterns suggest amphiphysin antibody, amphiphysin Western blot is performed at an additional charge.

If ACh receptor binding antibody is >0.02 or if striational antibodies are > or =1:60, ACh receptor modulating antibodies and CRMP-5-IgG Western blot are performed at an additional charge.

See Autoimmune Dysautonomia Evaluation Testing Algorithm in Special Instructions.

• Paraneoplastic Evaluation Algorithm
• Autoimmune Dysautonomia Evaluation Testing Algorithm

CCN/81184, GD65S/81596, ARBI/8338, ARMO/83378, GANG/84321, VGKC/89165, CCPQ/81185: Radioimmunoassay (RIA)

WBN/83108, CRMWS/83107, ABLOT/89381: Western blot

ANN1S/83381: Indirect Immunofluorescence Assay (IFA)

STR/8746: Enzyme Immunoassay (EIA)

PAINT/29347: Interpretive Comment

Container/Tube:

Preferred: Red top

Acceptable: Serum gel

Specimen Volume: 4 mL

Additional Information: Include relevant clinical information, name, phone number, mailing address, and e-mail address (if applicable) of ordering physician.

Autoimmune dysautonomia encompasses disorders of peripheral autonomic synapses, ganglionic neurons, autonomic nerve fibers and central autonomic pathways mediated by neural-specific IgG or effector T cells. These disorders may be idiopathic or paraneoplastic, subacute or insidious in onset, and may present as a limited disorder or generalized pandysautonomia. Pandysautonomia is usually subacute in onset and severe, and includes impaired pupillary light reflex, anhidrosis, orthostatic hypotension, cardiac arrhythmias, gastrointestinal dysmotility, sicca manifestations, and bladder dysfunction. Limited dysautonomia is confined to 1 or just a few domains, is often mild and may include sicca manifestations, postural orthostatism and cardiac arrhythmias, bladder dysfunction or gastrointestinal dysmotilities. Diagnosis of limited dysautonomia requires documentation of objective abnormalities by autonomic reflex testing, thermoregulatory sweat test or gastrointestinal motility studies.

The most commonly encountered autoantibody marker of autoimmune dysautonomia is the neuronal ganglionic alpha-3- (acetylcholine receptor) autoantibody. This autoantibody to date is the only proven effector of autoimmune dysautonomia. A direct relationship has been demonstrated between antibody titer and severity of dysautonomia in both alpha-3-AChR-immunized animals and patients with autoimmune dysautonomia. Patients with high alpha-3-AChR autoantibody values (>1.0 nmol/L) generally have profound pandys autonomia. Dysautonomic patients with lower alpha-3-AChR autoantibody values (0.03-0.99 nmol/L) have limited dysautonomia.

Importantly, cancer is detected in 30% of patients with alpha-3-AChR autoantibody. Cancers recognized most commonly include small-cell lung carcinomas, thymoma, adenocarcinomas of breast, lung, prostate and gastrointestinal tract, and lymphoma. Cancer risk factors include past or family history of cancer, history of smoking or social/environmental exposure to carcinogens. Early diagnosis and treatment of the neoplasm favors neurologic improvement and lessens morbidity.

Autoantibodies to other onconeural proteins shared by neurons, glia or muscle (eg, antineuronal nuclear antibody-type 1 [ANNA-1], CRMP-5-IgG, N-type voltage-gated calcium channel, muscle AChR and sarcomeric [striational antigens]) serve as additional markers of paraneoplastic or idiopathic dysautonomia. A specific neoplasm is often predictable by the individual patient’s autoantibody profile.

GANGLIONIC ACETYLCHOLINE RECEPTOR (ALPHA3) AUTOANTIBODY

< or =0.02 nmol/L

NEURONAL VOLTAGE-GATED POTASSIUM CHANNEL (VGKC) AUTOANTIBODY

< or =0.02 nmol/L

N-TYPE CALCIUM CHANNEL ANTIBODY

< or =0.03 nmol/L

P/Q-TYPE CALCIUM CHANNEL ANTIBODY

< or =0.02 nmol/L

ACETYLCHOLINE RECEPTOR (MUSCLE AChR) BINDING ANTIBODY

< or =0.02 nmol/L

GLUTAMIC ACID DECARBOXYLASE (GAD65) ANTIBODY ASSAY

< or =0.02 nmol/L

ANTINEURONAL NUCLEAR ANTIBODY-TYPE 1 (ANNA-1)

<1:240

Neuron-restricted patterns of IgG staining that do not fulfill criteria for ANNA-1 may be reported as "unclassified antineuronal IgG." Complex patterns that include non-neuronal elements may be reported as "uninterpretable."

STRIATIONAL (STRIATED MUSCLE) ANTIBODIES

<1:60

Antibodies directed at onconeural proteins shared by neurons, muscle and glia are valuable serological markers of a patient’s immune response to cancer. These autoantibodies are not found in healthy subjects, and are usually accompanied by subacute neurological symptoms and signs. It is not uncommon for more than 1 autoantibody to be detected in patients with autoimmune dysautonomia. These include:

-Plasma membrane cation channel antibodies (neuronal ganglionic [alpha-3] and muscle [alpha-1] acetylcholine receptor [AChR]; neuronal calcium channel N-type or P/Q-type, and neuronal voltage-gated potassium channel [VGKC] antibodies). All of these autoantibodies are potential effectors of autonomic dysfunction.

-Antineuronal nuclear autoantibody-type 1 (ANNA-1)

-Neuronal and muscle cytoplasmic antibodies (CRMP-5 IgG, glutamic acid decarboxylase [GAD65] and striational)

A rising autoantibody titer in previously seropositive patients suggests cancer recurrence.

Negative results do not exclude autoimmune dysautonomia or cancer.

1. Vernino S, Low PA, Fealey RD, et al: Autoantibodies to ganglionic acetylcholine receptors in autoimmune autonomic neuropathies. N Engl J Med 2000;343:847-855

2. O'Suilleabhain P, Low PA, Lennon VA: Autonomic dysfunction in the Lambert-Eaton myasthenic syndrome: serologic and clinical correlates. Neurology 1998;50:88-93

3. Dhamija R, Tan KM, Pittock SJ, et al: Serological profiles aiding the diagnosis of autoimmune gastrointestinal dysmotility. Clin Gastroenterol Hepatol 2008;6:988-992

4. McKeon A, Lennon VA, Lachance DH, et al: The ganglionic acetylcholine receptor autoantibody: oncological, neurological and serological accompaniments. Arch Neurol 66(6):735-741

Indirect immunofluorescence assay (IFA)

Before testing, patient's serum is preabsorbed with liver powder to remove non-organ-specific autoantibodies. After applying to a composite substrate of frozen mouse tissues (brain, kidney, and gut) and washing, fluorescein-conjugated goat antihuman IgG is applied to detect the distribution and pattern of patient IgG binding. (Pittock SJ, Kryzer TJ, Lennon VA: Paraneoplastic antibodies coexist and predict cancer, not neurological syndrome. Ann Neurol 2004; 56: 715-719)

Radioimmunoassay (RIA)

Goat antihuman IgG and IgM is used as precipitant in all assays. Cation channel protein antigens are solubilized from neuronal or muscle membrane, in non-ionic detergent, and complexed with a selective high-affinity ligand labeled with iodine (I). I-labelled recombinant human GAD65 antigen is used to confirm GAD65 autoantibody (when suspected from immunofluorescent staining pattern). (Griesmann GE, Kryzer TJ, Lennon VA: Autoantibody profiles of myasthenia gravis and Lambert-Eaton myasthenic syndrome. In Manual of Clinical and Laboratory Immunology, 6th edition. Edited by NR Rose, RG Hamilton, et al. Washington, DC, ASM Press, 2002, pp 1005-1012; Walikonis JE, Lennon VA: Radioimmunoassay for glutamic acid decarboxylase [GAD65] autoantibodies as a diagnostic aid for stiff-man syndrome and a correlate of susceptibility to type 1 diabetes mellitus. Mayo Clin Proc 1998; 73[12]:1161-1166)

Muscle acetylcholine receptor (AChR) modulating antibodies are detected by incubating the patient's serum for 14 hours with viable, non-innervated, monolayer cultures of human muscle cells. Percent loss of surface AChR is quantitated by probing with (125)I-alpha-bungarotoxin. (Howard FM Jr, Lennon VA, Finley J, et al: Clinical correlations of antibodies that bind, block, or modulate human acetylcholine receptors in myasthenia gravis. Ann NY Acad Sci 1987; 505:526-538)

Enzyme Immunoassay (EIA)

A mixture of sarcomeric proteins extracted from innervated rat skeletal muscle is used as antigen to detect striational antibodies (IgG, IgM, and IgA). (Cikes N, Momoi MY, Williams CL, et al: Striational autoantibodies: quantitative detection by enzyme immunoassay in myasthenia gravis, thymoma, and recipients of D-penicillamine or allogeneic bone marrow. Mayo Clin Proc 1988;63:474-481)

Western Blot (WB)

WB is performed when immunofluorescence assay (IFA) screening for antineuronal nuclear antibody-type 1 (ANNA-1) is not interpretable due to interfering autoantibodies. A mixture of neuronal antigens extracted aqueously from adult rat cerebellum is denatured, reduced, and separated by electrophoresis on 10% polyacrylamide gel. Full-length recombinant human CRMP-5 antigen is used to confirm CRMP-5-IgG. (Yu Z, Kryzer TJ, Griesmann GE, et al: CRMP-5 neuronal autoantibody: marker of lung cancer and thymoma-related autoimmunity. Ann Neurol 2001;49[2]:145-154)

ANNA-1: Monday through Thursday, Sunday; 11:30 a.m.

Striational (striated muscle) antibodies: Monday through Thursday, Sunday; 11 a.m.

N-type calcium channel antibody: Monday, Wednesday, Friday; 2 p.m.

Acetylcholine receptor (muscle AChR) binding antibody: Monday through Thursday, Saturday; 2 p.m.

Ganglionic acetylcholine receptor (alpha3) autoantibody: Tuesday, Thursday, Sunday; 2 p.m.

Neuronal (VGKC) autoantibody: Tuesday, Thursday, Sunday; 2 p.m.

GAD65 antibody assay: Monday through Thursday, Sunday; 6 a.m.

P/Q-type calcium channel antibody: Monday, Wednesday, Friday; 6 a.m.

Paraneoplastic autoantibody Western blot: Monday, Wednesday, Friday; 8 a.m.

Acetylcholine receptor (muscle) modulating antibodies: Monday through Thursday; 11 a.m.

CRMP-5-IgG Western blot: Monday through Friday; 8 a.m.

Amphiphysin Western blot: Tuesday, Thursday; 6 a.m.

83519-59-ACh receptor (muscle) binding antibody

83519-59-AChR ganglionic neuronal antibody

83519-59-Neuronal VGKC autoantibody

83519-59-N-type calcium channel antibody

83519-59-P/Q-Type Calcium Channel Ab

83520-Striational (striated muscle) antibodies

86256-ANNA-1

86341-GAD65 antibody assay

83519-59-ACh receptor (muscle) modulating antibodies (if appropriate)

84182-Paraneoplastic autoantibody Western blot confirmation (if appropriate)

84182-CRMP-5-IgG Western blot (if appropriate)

84182-Amphiphysin Western Blot (if appropriate)