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Test Code AIAES Axonal Neuropathy, Autoimmune/Paraneoplastic Evaluation, Serum


Ordering Guidance


Multiple neurological phenotype-specific autoimmune/paraneoplastic evaluations are available. For more information as well as phenotype-specific testing options, see Autoimmune Neurology Test Ordering Guide.

 

When more than one evaluation is ordered on the same order number, the duplicate test will be canceled.

 

For a list of antibodies performed with each evaluation, see Autoimmune Neurology Antibody Matrix.

 

This test should not be requested for patients who have recently received radioisotopes, therapeutically or diagnostically, because of potential assay interference. The specific waiting period before specimen collection will depend on the isotope administered, the dose given, and the clearance rate in the individual patient. Specimens will be screened for radioactivity prior to analysis. Radioactive specimens received in the laboratory will be held 1 week and assayed if sufficiently decayed or canceled if radioactivity remains.



Necessary Information


Provide the following information:

-Relevant clinical information

-Ordering healthcare professional's name, phone number, mailing address, and email address



Specimen Required


Patient Preparation: For optimal antibody detection, collect specimen before starting immunosuppressant medication or intravenous immunoglobulin treatment.

Supplies: Sarstedt Aliquot Tube, 5 mL (T914)

Collection Container/Tube:

Preferred: Red top

Acceptable: Serum gel

Submission Container/Tube: Plastic vial

Specimen Volume: 4 mL Serum

Collection Instructions: Centrifuge and aliquot serum into a plastic vial.


Secondary ID

606974

Useful For

Evaluation of patients who present with a subacute neurological disorder of undetermined etiology, especially those with known risk factors for cancer using serum specimens

 

Directing a focused search for cancer

 

Investigating neurological symptoms that appear during, or after, cancer therapy, and are not explainable by metastasis

 

Differentiating autoimmune neuropathies from neurotoxic effects of chemotherapy

 

Detecting early evidence of cancer recurrence in previously seropositive patients

Profile Information

Test ID Reporting Name Available Separately Always Performed
AIAEI Autoimmune Axonal Interp, S No Yes
AMPHS Amphiphysin Ab, S No Yes
AGN1S Anti-Glial Nuclear Ab, Type 1 No Yes
ANN1S Anti-Neuronal Nuclear Ab, Type 1 No Yes
ANN3S Anti-Neuronal Nuclear Ab, Type 3 No Yes
APBIS AP3B2 IFA, S No Yes
CS2CS CASPR2-IgG CBA, S No Yes
CRMS CRMP-5-IgG, S No Yes
IG5CS IgLON5 CBA, S No Yes
LG1CS LGI1-IgG CBA, S No Yes
PCAB2 Purkinje Cell Cytoplasmic Ab Type 2 No Yes

Reflex Tests

Test ID Reporting Name Available Separately Always Performed
AGNBS AGNA-1 Immunoblot, S No No
AGNTS AGNA-1 Titer, S No No
APHTS Amphiphysin Ab Titer, S No No
AMIBS Amphiphysin Immunoblot, S No No
AN1BS ANNA-1 Immunoblot, S No No
AN1TS ANNA-1 Titer, S No No
AN2BS ANNA-2 Immunoblot, S No No
AN3TS ANNA-3 Titer, S No No
APBCS AP3B2 CBA, S No No
APBTS AP3B2 IFA Titer, S No No
CRMTS CRMP-5-IgG Titer, S No No
IG5TS IgLON5 IFA Titer, S No No
MP1BS MAP1B Immunoblot, S No No
PC2TS PCA-2 Titer, S No No
CRMWS CRMP-5-IgG Western Blot, S No No

Testing Algorithm

If the indirect immunofluorescence assay (IFA) patterns suggest antiglial nuclear antibody-1 (AGNA-1), then the AGNA-1 antibody IFA titer and AGNA-1 antibody immunoblot will be performed at an additional charge.

 

If the IFA patterns suggest antineuronal nuclear antibody type 1 (ANNA-1), then the ANNA-1 IFA titer, ANNA-1 immunoblot, and ANNA-2 immunoblot will be performed at an additional charge.

 

If the IFA pattern suggests ANNA-3 antibody or the client requests it, then the ANNA-3 IFA titer will be performed at an additional charge.

 

If the IFA pattern suggests PCA-2 antibody, then the PCA-2 antibody IFA titer and MAP1B immunoblot will be performed at an additional charge.

 

If the IFA patterns suggest amphiphysin antibody, then the amphiphysin antibody IFA titer and amphiphysin antibody immunoblot will be performed at an additional charge.

 

If the IFA patterns suggest collapsin response-mediator protein-5 (CRMP-5)-IgG, then the CRMP-5-IgG IFA titer and CRMP-5-IgG Western blot will be performed at an additional charge.

 

If the IFA pattern suggests adaptor protein 3 beta2 (AP3B2) antibody, then the AP3B2 antibody IFA titer and AP3B2 antibody CBA will be performed at an additional charge.

 

If the IgLON family member 5 antibody (IgLON5) by CBA result is positive, then the IgLON5 antibody IFA titer will be performed at an additional charge.

 

For more information see:

-Autoimmune/Paraneoplastic Axonal Neuropathy Evaluation Algorithm.

-Acquired Neuropathy Diagnostic Algorithm

Method Name

AIAEI: Technical Interpretation

 

AMPHS, AGN1S, ANN1S, ANN3S, APBIS, CRMS, PCAB2, AGNTS, AN1TS, AN3TS, APBTS, APHTS, CRMTS, IG5TS, PC2TS: Indirect Immunofluorescence Assay (IFA)

 

CS2CS, IG5CS, LG1CS, APBCS: Cell Binding Assay (CBA)

 

CRMWS: Western Blot (WB)

 

AGNBS, AMIBS, AN1BS, AN2BS, MP1BS: Immunoblot (IB)

Reporting Name

Axonal, Autoimm/Paraneo, S

Specimen Type

Serum

Specimen Minimum Volume

Serum: 2 mL

Specimen Stability Information

Specimen Type Temperature Time
Serum Refrigerated (preferred) 28 days
  Frozen  28 days
  Ambient  72 hours

Reject Due To

Gross hemolysis Reject
Gross lipemia Reject
Gross icterus Reject

Clinical Information

Neuropathy patients have variable sensory disturbance (loss or exaggerated sensation) with pain, weakness, and autonomic involvements such as sweat abnormalities, gastrointestinal dysfunction, and lightheadedness on standing. These symptoms result from injury to the distal nerves, roots, ganglia, or their gathering points (nerve plexus in the thighs and arms). Patients may have symmetric or asymmetric involvements of the extremities, trunk, and head, including extraocular muscles. Subacute onsets and asymmetric involvements favor inflammatory or immune causes over inherited or metabolic forms. Depending on the specific inflammatory or immune mediated causes other parts of the nervous system may also be affected (brain, cerebellum, spinal cord).

 

In the evaluation of patients with immune mediated autoantibody neuropathies, nerve conduction studies and needle electromyography can help to classify the neuropathy as either primary axonal, primary demyelinating, or mixed axonal and demyelinating. This evaluation focuses on persons with primary axonal forms.

 

Well established neuronal autoantibodies responsible for axonal neuropathies include antineuronal nuclear antibody (ANNA1 and 3), Purkinje cytoplasmic antibody type 2 (PCA2), amphiphysin antibody, collapsin response mediator protein 5 (CRMP5) antibody, leucine-rich glioma inactivated 1 protein (LGI1) antibody, and contactin-associated response protein 2 (CASPR2) antibody. Other autoantibodies have preliminary evidence to support their association with neuropathy, including antiglial nuclear antibody (AGNA), and antineuronal nuclear antibody type 2 (ANNA2).

 

A patient's humoral and cellular immune response leads to the neurological syndrome. This may be related to an underlying cancer or unidentified antigen trigger. If related to cancer, it may be a new or recurrent malignancy, is usually limited in metastatic volume, and is often occult by standard imaging procedures. Autoantibodies specific for onconeural proteins found in the plasma membrane, cytoplasm, and nucleus of neurons, glia, or muscle are generated in this immune response and serve as serological markers of paraneoplastic autoimmunity. Cancers recognized in this context most commonly are small-cell lung carcinoma, thymoma, ovarian (or related Mullerian) carcinoma, breast carcinoma, and Hodgkin lymphoma. Pertinent childhood neoplasms recognized thus far include neuroblastoma, thymoma, Hodgkin lymphoma, and chondroblastoma.

 

This evaluation focuses on those antibodies with known associations with varied forms of peripheral axonal neuropathy. Seropositive patients usually present with subacute neurological symptoms of radiculopathy; plexopathy; or sensory, sensorimotor, or autonomic neuropathy, with or without a neuromuscular transmission disorder, such as neuromuscular hyperexcitability. Other peripheral manifestations include cranial neuropathies, especially loss of vision, hearing, smell, or taste. Commonly beyond the peripheral manifestation are encephalopathy, seizures, cerebellar ataxia, and myelopathy. Initial signs may be subtle, but a subacute multifocal and progressive syndrome usually evolves. Sensorimotor neuropathy and cerebellar ataxia are common presentations, but the clinical picture in some patients is dominated by striking gastrointestinal dysmotility and limbic encephalopathy. Some patients may present with mostly pain and have a limited small fiber neuropathy with or without autonomic symptoms.

 

Cancer risk factors include previous or family history of cancer, history of smoking, or social or environmental exposure to carcinogens.

Reference Values

Test ID

Reporting name

Methodology

Reference value

AIAEI

Autoimmune Axonal Interp, S

Technical interpretation

Interpretive report

AMPHS

Amphiphysin Ab, S

IFA

Negative

AGN1S

Anti-Glial Nuclear Ab, Type 1

IFA

Negative

ANN1S

Anti-Neuronal Nuclear Ab, Type 1

IFA

Negative

ANN3S

Anti-Neuronal Nuclear Ab, Type 3

IFA

Negative

APBIS

AP3B2 IFA, S

IFA

Negative

CS2CS

CASPR2-IgG CBA, S

CBA

Negative

CRMS

CRMP-5-IgG, S

IFA

Negative

IG5CS

IgLON5 CBA, S

CBA

Negative

LG1CS

LGI1-IgG CBA, S

CBA

Negative

PCAB2

Purkinje Cell Cytoplasmic Ab Type 2

IFA

Negative

 Reflex Information:

Test ID

Reporting name

Methodology

Reference value

AGNBS

AGNA-1 Immunoblot, S

IB

Negative

AGNTS

AGNA-1 Titer, S

IFA

<1:240

APHTS

Amphiphysin Ab Titer, S

IFA

<1:240

AMIBS

Amphiphysin Immunoblot, S

IB

Negative

AN1BS

ANNA-1 Immunoblot, S

IB

Negative

AN1TS

ANNA-1 Titer, S

IFA

<1:240

AN2BS

ANNA-2 Immunoblot, S

IB

Negative

AN3TS

ANNA-3 Titer, S

IFA

<1:240

APBCS

AP3B2 CBA, S

CBA

Negative

APBTS

AP3B2 IFA Titer, S

IFA

<1:240

CRMTS

CRMP-5-IgG Titer, S

IFA

<1:240

CRMWS

CRMP-5-IgG Western Blot, S

WB

Negative

IG5TS

IgLON5 IFA Titer, S

IFA

<1:240

MP1BS

MAP1B Immunoblot, S

IB

Negative

PC2TS

PCA-2 Titer, S

IFA

<1:240

 

*Methodology abbreviations:

Cell-binding assay (CBA)

Immunoblot (IB)

Immunofluorescence assay (IFA)

Western blot (WB)

Interpretation

Antibodies directed at onconeural proteins shared by neurons, glia, muscle, and certain cancers are valuable serological markers of a patient's immune response to cancer. They are not found in healthy individuals and are usually accompanied by subacute neurological signs and symptoms. Several autoantibodies have a syndromic association, but no autoantibody predicts a specific neurological syndrome. More than one paraneoplastic autoantibody may be detected and associated with specific cancers.

Method Description

Cell-Binding Assay:

Patient sample is applied to a composite slide containing transfected and nontransfected cells. After incubation and washing, fluorescein-conjugated goat-antihuman IgG is applied to detect the presence of patient IgG binding.(Klein CJ, Lennon VA, Aston PA, et al. Insights from LGI1 and CASPR2 potassium channel complex autoantibody subtyping. JAMA Neurol. 2013;70(2):229-234; package insert: IIFT: Neurology Mosaics, Instructions for the indirect immunofluorescence test. EUROIMMUN; FA_112d-1_A_UK_C13, 02/2019)

 

Indirect Immunofluorescence Assay:

The patient's sample is tested by a standardized immunofluorescence assay that uses a composite frozen section of mouse cerebellum, kidney, and gut tissues. After incubation with sample and washing, fluorescein-conjugated goat-antihuman IgG is applied. Neuron-specific autoantibodies are identified by their characteristic fluorescence staining patterns. Samples that score positive for any neuronal nuclear or cytoplasmic autoantibody are titrated to an endpoint. Interference by coexisting non-neuron-specific autoantibodies can usually be eliminated by serologic absorption.(Honorat JA, Komorowski L, Josephs KA, et al. IgLON5 antibody: neurological accompaniments and outcomes in 20 patients. Neurol Neuroimmunol Neuroinflamm. 2017;4[5]:e385. doi:10.1212/NXI.0000000000000385)

 

Western Blot:

Neuronal antigens extracted aqueously from adult rat cerebellum, full-length recombinant human collapsin response-mediator protein-5 (CRMP-5), or full-length recombinant human amphiphysin protein is denatured, reduced, and separated by electrophoresis on 10% polyacrylamide gel. IgG is detected autoradiographically by enhanced chemiluminescence.(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]:146-154; Dubey D, Jitprapaikulsan J, Bi H, et al. Amphiphysin-IgG autoimmune neuropathy: A recognizable clinicopathologic syndrome. Neurology. 2019;93[20]:e1873-e1880. doi:10.1212/WNL.0000000000008472)

 

Immunoblot for amphiphysin, ANNA-1, and ANNA-2:

All steps are performed at ambient temperature (18-28° C) utilizing the EUROBlot One instrument. Diluted patient sample is added to test strips (strips containing recombinant antigen manufactured and purified using biochemical methods) in individual channels and incubated for 30 minutes. Positive samples will bind to the purified recombinant antigen and negative samples will not bind. Strips are washed to remove unbound antibodies and then are incubated with antihuman IgG antibodies (alkaline phosphatase-labelled) for 30 minutes. The strips are again washed to remove unbound antihuman IgG antibodies and nitroblue tetrazolium chloride/5-bromo-4-chloro-3-indolylphosphate substrate is added. Alkaline phosphatase enzyme converts the soluble substrate into a colored insoluble product on the membrane to produce a black band. Strips are digitized via picture capture on the EUROBlot One instrument and evaluated with the EUROLineScan software.(O'Connor K, Waters P, Komorowski L, et al. GABAA receptor autoimmunity: A multicenter experience. Neurol Neuroimmunol Neuroinflamm. 2019;6[3]:e552. doi:10.1212/NXI.0000000000000552)

 

Immunoblot for MAP1B:

Test strips are incubated with appropriately diluted sample, prepared to meet the assay's required end-dilution for each specimen type. Samples and controls are added directly to the strips, ensuring they are fully covered with dilution buffer, then mixed gently and incubated for 60 minutes at room temperature on a rocking platform. After incubation, the strips undergo a series of wash steps using diluted wash buffer, followed by a 30-minute incubation with a ready-to-use alkaline phosphatase-labeled IgG conjugate. The strips are washed again using the same procedure, then incubated with a ready-to-use substrate solution for 20 minutes at room temperature. The reaction is stopped by rinsing with distilled water through several wash cycles. Strips are then placed on absorbent material and allowed to dry fully, generally requiring at least two hours. Results are interpreted only after the strips have dried completely, and dried strips are stored protected from light.(Package insert: PCA-2 line assay. ravo Diagnostika GmbH; 05/2023)

Day(s) Performed

Profile tests: Monday through Sunday; Reflex tests: Varies

Performing Laboratory

Mayo Clinic Laboratories in Rochester

Test Classification

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

CPT Code Information

86255 x 10

84182 AGNBS (if appropriate)

86256 AGNTS (if appropriate)

86256 APHTS (if appropriate)

84182 AMIBS (if appropriate)

84182 AN1BS (if appropriate)

86256 AN1TS (if appropriate)

84182 AN2BS (if appropriate)

86256 AN3TS (if appropriate)

86255 APBCS (if appropriate)

86256 APBTS (if appropriate)

86256 CRMTS (if appropriate)

84182 CRMWS (if appropriate)

86256 IG5TS (if appropriate)

84182 MP1BS (if appropriate)

86256 PC2TS (if appropriate)

LOINC Code Information

Test ID Test Order Name Order LOINC Value
AIAES Axonal, Autoimm/Paraneo, S 94695-4

 

Result ID Test Result Name Result LOINC Value
606950 IgLON5 CBA, S 96478-3
615863 AP3B2 IFA, S 101907-4
89080 AGNA-1, S 84927-3
81722 Amphiphysin Ab, S 72327-0
80150 ANNA-1, S 33615-6
83137 ANNA-3, S 43102-3
83077 CRMP-5-IgG, S 72504-4
83138 PCA-2, S 84925-7
64279 LGI1-IgG CBA, S 94287-0
64281 CASPR2-IgG CBA, S 94285-4
606975 Autoimmune Axonal Interp, S 69048-7
618900 IFA Notes 48767-8