Test Id : MAS1

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

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

If the IFA pattern suggests 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-2 antibodies, then the ANNA-2 IFA titer, ANNA-2 immunoblot, and ANNA-1 immunoblot will be performed at an additional charge.

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

If the IFA pattern suggests adaptor protein 3 beta 2 (AP3B2) antibodies, then the AP3B2 cell-binding assay (CBA) and AP3B2 IFA titer will be performed at an additional charge.

If the CRMP-5-IgG Western blot is positive, then the CRMP-5-IgG IFA titer will be performed at an additional charge.

If the IFA pattern suggests Purkinje cytoplasmic antibody type 1 (PCA-1), then the PCA-1 immunoblot and PCA-1 IFA titer will be performed at an additional charge.

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

If gamma-aminobutyric acid B (GABA-B) receptor antibody CBA result is positive, then the GABA-B-receptor antibody IFA titer will be performed at an additional charge.

If the dipeptidyl-peptidase-like protein-6 (DPPX) antibody CBA result is positive, then the DPPX antibody IFA titer will be performed at an additional charge.

If the IFA pattern suggests metabotropic glutamate receptor 1 (mGluR1) antibody, then the mGluR1antibody CBA and mGluR1 antibody IFA titer will be performed at an additional charge.

If the IFA pattern suggests glial fibrillary acidic protein (GFAP) antibody, then the GFAP antibody CBA and GFAP antibody IFA titer will be performed at an additional charge.

If the neuromyelitis optica/aquaporin-4-IgG (NMO/AQP4-IgG) fluorescence-activated cell sorting (FACS) screen assay requires further investigation, then the NMO/AQP4-IgG FACS titration assay will be performed at an additional charge.

If the myelin oligodendrocyte glycoprotein (MOG) FACS screen assay requires further investigation, then the MOG FACS titration assay will be performed at an additional charge.

If the IFA pattern suggests neuronal intermediate filament (NIF) antibody, then the alpha internexin CBA, NIF heavy chain CBA, NIF light chain CBA, and NIF antibody IFA titer will be performed at an additional charge.

If the IFA pattern suggests neurochondrin antibody, then the neurochondrin antibody CBA and neurochondrin IFA titer will be performed at an additional charge.

If the IFA pattern suggests septin-7 antibody, then the septin-7 CBA and septin-7 IFA titer will be performed at an additional charge.

If the IFA pattern suggests tripartite motif-containing protein 46 (TRIM46) antibody, then the TRIM46 antibody CBA and TRIM46 IFA titer will be performed at an additional charge.

For more information see Autoimmune/Paraneoplastic Myelopathy Evaluation Algorithm-Serum.

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.

Provide the following information:

-Relevant clinical information

-Ordering provider name, phone number, mailing address, and e-mail address

For optimal antibody detection, specimen collection is recommended prior to initiation of 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

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

• Autoimmune/Paraneoplastic Myelopathy Evaluation Algorithm-Serum

If not ordering electronically, complete, print, and send a Neurology Specialty Testing Client Test Request (T732) with the specimen.

2 mL

Evaluating patients with suspected autoimmune myelopathy, myelitis, and paraneoplastic myelopathy using serum specimens

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

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

If the IFA pattern suggests 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-2 antibodies, then the ANNA-2 IFA titer, ANNA-2 immunoblot, and ANNA-1 immunoblot will be performed at an additional charge.

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

If the IFA pattern suggests adaptor protein 3 beta 2 (AP3B2) antibodies, then the AP3B2 cell-binding assay (CBA) and AP3B2 IFA titer will be performed at an additional charge.

If the CRMP-5-IgG Western blot is positive, then the CRMP-5-IgG IFA titer will be performed at an additional charge.

If the IFA pattern suggests Purkinje cytoplasmic antibody type 1 (PCA-1), then the PCA-1 immunoblot and PCA-1 IFA titer will be performed at an additional charge.

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

If gamma-aminobutyric acid B (GABA-B) receptor antibody CBA result is positive, then the GABA-B-receptor antibody IFA titer will be performed at an additional charge.

If the dipeptidyl-peptidase-like protein-6 (DPPX) antibody CBA result is positive, then the DPPX antibody IFA titer will be performed at an additional charge.

If the IFA pattern suggests metabotropic glutamate receptor 1 (mGluR1) antibody, then the mGluR1antibody CBA and mGluR1 antibody IFA titer will be performed at an additional charge.

If the IFA pattern suggests glial fibrillary acidic protein (GFAP) antibody, then the GFAP antibody CBA and GFAP antibody IFA titer will be performed at an additional charge.

If the neuromyelitis optica/aquaporin-4-IgG (NMO/AQP4-IgG) fluorescence-activated cell sorting (FACS) screen assay requires further investigation, then the NMO/AQP4-IgG FACS titration assay will be performed at an additional charge.

If the myelin oligodendrocyte glycoprotein (MOG) FACS screen assay requires further investigation, then the MOG FACS titration assay will be performed at an additional charge.

If the IFA pattern suggests neuronal intermediate filament (NIF) antibody, then the alpha internexin CBA, NIF heavy chain CBA, NIF light chain CBA, and NIF antibody IFA titer will be performed at an additional charge.

If the IFA pattern suggests neurochondrin antibody, then the neurochondrin antibody CBA and neurochondrin IFA titer will be performed at an additional charge.

If the IFA pattern suggests septin-7 antibody, then the septin-7 CBA and septin-7 IFA titer will be performed at an additional charge.

If the IFA pattern suggests tripartite motif-containing protein 46 (TRIM46) antibody, then the TRIM46 antibody CBA and TRIM46 IFA titer will be performed at an additional charge.

For more information see Autoimmune/Paraneoplastic Myelopathy Evaluation Algorithm-Serum.

Patients with autoimmune myelopathy present with subacute onset and rapid progression of spinal cord symptoms with one or more of the following: weakness, gait difficulties, loss of sensation, neuropathic pain, and bowel and bladder dysfunction. Clinical history and examination, spinal cord magnetic resonance imaging, and cerebrospinal fluid (CSF) testing may provide clues to an autoimmune diagnosis. Autoimmune myelopathy evaluation of both serum and CSF can assist in the diagnosis (paraneoplastic or idiopathic autoimmune) and aid distinction from other causes of myelopathy (multiple sclerosis, sarcoidosis, vascular disease). Early testing may assist in early diagnosis of occult cancer, prompt initiation of immune therapies, or both.

Reflex Information

*Methodology abbreviations:

Immunofluorescence assay (IFA)

Cell-binding assay (CBA)

Flow cytometry (FCM)

Radioimmunoassay (RIA)

Immunoblot (IB)

Western blot (WB)

Neuron-restricted patterns of IgG staining that do not fulfill criteria for ANNA-1, ANNA-2, ANNA-3, CRMP-5-IgG, PCA-1, or PCA-2 may be reported as "unclassified anti-neuronal IgG." Complex patterns that include non-neuronal elements may be reported as "uninterpretable."

A positive result is consistent with a diagnosis of autoimmune myelopathy in the appropriate clinical context.

Negative results do not exclude a diagnosis of autoimmune myelopathy.

Intravenous immunoglobulin treatment prior to the serum collection may cause a false-positive result.

1. Dubey D, Pittock SJ, Krecke KN, et al. Clinical, radiologic, and prognostic features of myelitis associated with myelin oligodendrocyte glycoprotein autoantibody. JAMA Neurol. 2019;76(3):301-309

2. Zalewski NL, Flanagan EP. Autoimmune and paraneoplastic myelopathies. Semin Neurol. 2018;38(3):278-289

3. Flanagan EP, Hinson SR, Lennon VA, et al. Glial fibrillary acidic protein immunoglobulin G as biomarker of autoimmune astrocytopathy: Analysis of 102 patients. Ann Neurol. 2017;81(2):298-309

4. Keegan BM, Pittock SJ, Lennon VA. Autoimmune myelopathy associated with collapsin response-mediator protein-5 immunoglobulin G. Ann Neurol. 2008;63(4):531-534

5. Weinshenker BG, Wingerchuk DM, Vukusic S, et al. Neuromyelitis optica IgG predicts relapse after longitudinally extensive transverse myelitis. Ann Neurol. 2006;59(3):566-569

Cell-Binding Assay:

Patient specimen is applied to a composite slide containing transfected and nontransfected HEK-293 cells. After incubation and washing, fluorescein-conjugated goat-antihuman IgG is applied to detect the presence of patient IgG binding.(Package insert: IIFT: Neurology Mosaics, Instructions for the indirect immunofluorescence test. EUROIMMUN; FA_112d-1_A_UK_C13, 02/25/2019)

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 are scored 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)

Radioimmunoassay:

(125)I-labeled recombinant human antigens or labeled receptors are incubated with patient specimen. After incubation, anti-human IgG is added to form an immunoprecipitate. The amount of (125)I-labeled antigen in the immunoprecipitate is measured using a gamma-counter. The amount of gamma emission in the precipitate is proportional to the amount of antigen-specific IgG in the specimen. Results are reported as units of precipitated antigen (nmol) per liter of patient sample.(Griesmann GE, Kryzer TJ, Lennon VA: Autoantibody profiles of myasthenia gravis and Lambert-Eaton myasthenic syndrome. In: Rose NR, Hamilton RG, eds. Manual of Clinical and Laboratory Immunology. 6th ed. ASM Press; 2002:1005-1012; 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; Jones AL, Flanagan EP, Pittock SJ, et al. Responses to and outcomes of treatment of autoimmune cerebellar ataxia in adults. JAMA Neurol. 2015;72[11]:1304-1312. doi:10.1001/jamaneurol.2015.2378)

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, Kim K, Benarroch EE, Lennon VA. 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:

All steps are performed at room temperature (18-28 degrees C) utilizing the EUROBlot One instrument. Diluted patient specimen (1:101) is added to test strips (strips containing recombinant antigen manufactured and purified using biochemical methods) in individual channels and incubated for 30 minutes. Positive specimens will bind to the purified recombinant antigen and negative specimens will not bind. Strips are washed to remove unbound antibodies and then incubated with anti-human IgG antibodies (alkaline phosphatase-labeled) for 30 minutes. The strips are again washed to remove unbound anti-human IgG antibodies and nitroblue tetrazolium chloride/5-bromo-4-chloro-3-indolyl phosphate 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)

Neuromyelitis Optica -IgG Fluorescence-Activated Cell Sorting Assay/Flow Cytometry:

Human embryonic kidney cells (HEK 293) are transfected transiently with a plasmid (pIRES2- Aequorea coerulescens green fluorescent protein) encoding both green fluorescent protein (AcGFP) and aquaporin-4 (AQP4)-M1. After 36 hours, a mixed population of cells (transfected expressing AQP4 on the surface and AcGFP in the cytoplasm and nontransfected lacking AQP4 and AcGFP) are lifted and resuspended in live cell-binding buffer. Cells are incubated with patient serum and an AlexaFluor 647-labeled secondary antibody is added. Two populations are gated based on AcGFP expression: positive (high AQP4 expression) and negative (low or no AQP4 expression). Positivity is based on the ratio (positive >2.0) of the average median fluorescence intensity (MFI) of each cell population (MFI GFP positive:MFI GFP negative).(Fryer JP, Lennon VA, Pittock SJ, et al. AQP4 autoantibody assay performance in clinical laboratory service. Neurol Neuroimmunol Neuroinflamm. 2014;1[1]:e11. doi:10.1212/NXI.0000000000000011)

Myelin Oligodendrocyte Glycoprotein -IgG1 Fluorescence-Activated Cell Sorting Assay/Flow Cytometry:

HEK 293 is transfected transiently with a DNA plasmid that allows coexpression of both a reporter fluorescent protein (AcGFP) and full-length myelin oligodendrocyte glycoprotein (MOG). After 36 hours, a mixed population of cells (transfected expressing MOG on the surface and AcGFP in the cytoplasm and nontransfected lacking MOG and AcGFP) are lifted and resuspended in live cell-binding buffer. Cells are incubated with patient serum and an AlexaFluor 647 labeled secondary antibody is added. Two populations are gated based on AcGFP expression: positive (high MOG expression) and negative (low or no MOG expression). Positivity is based on the ratio (positive >2.5) of the average MFI of each cell population (MFI GFP positive:MFI GFP negative).(Unpublished Mayo method)

Profile tests: Monday through Sunday; Reflex tests: Varies

• Authorized users can sign in to Test Prices for detailed fee information.
• Clients without access to Test Prices can contact Customer Service 24 hours a day, seven days a week.
• Prospective clients should contact their account representative. For assistance, contact Customer Service.

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.

86255 x 16

86053

84182

86363

86341

84182 AGNBS (if appropriate)

86256 AGNTS (if appropriate)

86255 AINCS (if appropriate)

84182 AMIBS (if appropriate)

84182 AN1BS (if appropriate)

86256 AN1TS (if appropriate)

84182 AN2BS (if appropriate)

86256 AN2TS (if appropriate)

86256 AN3TS (if appropriate)

86255 APBCS (if appropriate)

86256 APBTS (if appropriate)

86256 APHTS (if appropriate)

86256 CRMTS (if appropriate)
86256 DPPTS (if appropriate)

86256 GABIS (if appropriate)

86255 GFACS (if appropriate)

86256 GFATS (if appropriate)

86255 GL1CS (if appropriate)

86256 GL1TS (if appropriate)

86363 MOGTS (if appropriate)

86255 NCDCS (if appropriate)

86256 NCDTS (if appropriate)

86255 NFHCS (if appropriate)

86255 NFLCS (if appropriate)

86256 NIFTS (if appropriate)
86053 NMOTS (if appropriate)

84182 PC1BS (if appropriate)

86256 PC1TS (if appropriate)

86256 PC2TS (if appropriate)

86255 SP7CS (if appropriate)

86256 SP7TS (if appropriate)

86255 T46CS (if appropriate)

86256 T46TS (if appropriate)