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Unit Code 80013:
Paraneoplastic Autoantibody Evaluation, Spinal Fluid

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Method Description

Indirect Immunofluorescence Assay (IFA)

The patient's sample is tested by a standardized IFA

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. Autoantibody of ANNA-1 specificity is panneuronal in

reactivity; it binds to the cytoplasm and nucleus (sparing nucleolus)

of central and peripheral nervous system neurons. Autoantibody of ANNA-2

specificity has a similar binding pattern but reactivity is restricted to

neurons of the central nervous system; neurons of the peripheral

nervous system are nonreactive. Autoantibody of PCA-1 specificity bind

distinctively to endoplasmic reticulum in Purkinje cells, molecular

neurons, and other large neurons in the central and peripheral nervous

system. Samples that are scored positive for any neuronal

nuclear or cytoplasmic autoantibody are titrated to an endpoint on

mouse Purkinje cells or peripheral neurons. Interference by coexisting

nonneuron-specific autoantibodies can usually be eliminated by

serologic absorption. Occasionally Western blot analysis is required.

(Yu Z, Kryzer TJ, Griesmann GE, Kim KK, et al: CRMP-5 neuronal

autoantibody:  marker of lung cancer and thymoma-related

autoimmunity. Ann Neurol 2001;49:146-154)

 

Western Blot

Western blot testing is indicated in the infrequent situation that

interference by coexisting nonneuronal-specific autoantibodies

precludes IFA interpretation. It is not cost-effective for routine

serologic screening. However, results obtained by Western

blot analyses in Mayo Clinic's Neuroimmunology Laboratory have

shown 100% concordance with IFA results for PCA-1, and superior

sensitivity of the IFA for detection of ANNA-1 and ANNA-2.

 

An aqueous extract of adult rat cerebellar proteins is used as the

source of neuronal antigens. Western blot is performed on denatured

and reduced proteins separated by electrophoresis on 10% poly-

acrylamide gel. (Vernino S, Lennon VA:  New Purkinje cell antibody

[PCA-2]:  marker of lung cancer-related neurological autoimmunity.

Ann Neurol 2000 March;47[3]:297-305)

 

Immunoprecipitation Assay

(125)I-labeled recombinant human GAD65 and nonimmune human

serum are incubated with the patient's diluted CSF. Antihuman

IgG and IgM are then added to form an immunoprecipitate. After

washing the precipitated immune complexes, specific antibodies

are detected by counting gamma-emission from the pellet's bound

I-GAD65. (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 December;73[12]:1161-1166)

Performing Laboratory Location

Rochester

Key