serum samples for this retrospective case - control study were collected in the clinical department of neurology innsbruck between 2005 and 2014 and stored at 80c until use .
serum samples from 366 individuals were included : 51 patients with adem , 41 patients with nmosd , 34 patients with clinically isolated syndrome ( cis ) , and 89 patients with ms .
nine patients with nmdar encephalitis , 94 patients with other neurologic diseases , and 48 healthy individuals were included as controls . most of the patients have previously been included in a study on nmdar antibody assay validation and in other studies on mog and/or aqp4 antibodies .
nmosds were diagnosed as described by wingerchuk et al . in 2007 , adem was diagnosed according to the criteria of the international pediatric ms study group , and ms and cis were diagnosed according to the 2005 revisions to the mcdonald criteria , as previously described .
diagnosis of nmdar encephalitis was based on clinical assessment ( e.g. , new onset of neuropsychiatric symptoms ) and demonstration of antibodies in serum or csf , as recommended recently .
the relative frequency of symptoms suggestive of nmdar encephalitis was selectively increased in our cohort of patients with demyelinating diseases ( figure ) .
of these , 81 presented with seizures and/or behavioral symptoms whereas 134 did not . as expected , all 9 patients with nmda receptor ( nmdar ) encephalitis presented with seizures and/or behavioral symptoms .
ab = antibody ; adem = acute disseminated encephalomyelitis ; aqp4 = aquaporin-4 ; cis = clinically isolated syndrome ; hc = healthy controls ; mog = myelin oligodendrocyte glycoprotein ; ms = multiple sclerosis ; nmdar - e = nmdar encephalitis ; nmosd = neuromyelitis optica spectrum disorder ; ond = other neurologic diseases .
the present study was approved by the ethical committee of innsbruck medical university ( study numbers am3041a and am4059 ) .
for the detection of autoantibodies against nmdar , aqp4 , and mog we used live hek293a cells transfected with the respective complementary dna , as described previously .
the following cutoff values were used : 1:20 for nmdar antibodies , 1:160 for mog antibodies , and 1:20 for aqp4 antibodies . to prove the specificity of antibody binding
one csf sample was tested for nmdar antibodies as described above , except csf was applied undiluted but supplemented with mk-801 ( sigma - aldrich , st .
louis , mo ) and further diluted ( 1:2 , 1:4 , etc . ) in washing buffer containing mk-801 .
nmdar antibody testing with fixed cell - based assay ( cba ) and rat brain immunohistochemistry ( ihc ) was done as described recently .
serum samples for this retrospective case - control study were collected in the clinical department of neurology innsbruck between 2005 and 2014 and stored at 80c until use .
serum samples from 366 individuals were included : 51 patients with adem , 41 patients with nmosd , 34 patients with clinically isolated syndrome ( cis ) , and 89 patients with ms .
nine patients with nmdar encephalitis , 94 patients with other neurologic diseases , and 48 healthy individuals were included as controls . most of the patients have previously been included in a study on nmdar antibody assay validation and in other studies on mog and/or aqp4 antibodies .
nmosds were diagnosed as described by wingerchuk et al . in 2007 , adem was diagnosed according to the criteria of the international pediatric ms study group , and ms and cis were diagnosed according to the 2005 revisions to the mcdonald criteria , as previously described .
diagnosis of nmdar encephalitis was based on clinical assessment ( e.g. , new onset of neuropsychiatric symptoms ) and demonstration of antibodies in serum or csf , as recommended recently .
the relative frequency of symptoms suggestive of nmdar encephalitis was selectively increased in our cohort of patients with demyelinating diseases ( figure ) .
of these , 81 presented with seizures and/or behavioral symptoms whereas 134 did not . as expected , all 9 patients with nmda receptor ( nmdar ) encephalitis presented with seizures and/or behavioral symptoms .
ab = antibody ; adem = acute disseminated encephalomyelitis ; aqp4 = aquaporin-4 ; cis = clinically isolated syndrome ; hc = healthy controls ; mog = myelin oligodendrocyte glycoprotein ; ms = multiple sclerosis ; nmdar - e = nmdar encephalitis ; nmosd = neuromyelitis optica spectrum disorder ; ond = other neurologic diseases .
the present study was approved by the ethical committee of innsbruck medical university ( study numbers am3041a and am4059 ) .
for the detection of autoantibodies against nmdar , aqp4 , and mog we used live hek293a cells transfected with the respective complementary dna , as described previously .
the following cutoff values were used : 1:20 for nmdar antibodies , 1:160 for mog antibodies , and 1:20 for aqp4 antibodies . to prove the specificity of antibody binding
one csf sample was tested for nmdar antibodies as described above , except csf was applied undiluted but supplemented with mk-801 ( sigma - aldrich , st .
louis , mo ) and further diluted ( 1:2 , 1:4 , etc . ) in washing buffer containing mk-801 .
nmdar antibody testing with fixed cell - based assay ( cba ) and rat brain immunohistochemistry ( ihc ) was done as described recently .
demographic data and the nmdar , mog , and aqp4 antibody seroprevalence of all patients and controls are shown in the table . demographic data and prevalence of antibodies in patients with demyelinating diseases and controls
serum nmdar antibodies were present in 9/9 ( 100% ) patients with nmdar encephalitis , in only 1/89 ( 1% ) patients with ms , and in none of the other cases .
csf positivity of nmdar antibodies was confirmed in all patients with nmdar encephalitis by fixed cba and/or tissue ihc .
csf from the patient with ms with low titer ( 1:20 ) serum nmdar antibodies at disease onset was negative for nmdar antibodies in our live cba but positive in the fixed cba and ihc .
in contrast , serum was negative in the fixed cba despite being positive in live cba and weakly positive in ihc . at the age of 23 , this woman presented with paresthesia of the left hand .
mri showed multiple ms - typical t2 lesions , and investigation of csf showed the presence of oligoclonal bands and 15 cells/l .
the patient was diagnosed with ms , and complete remission was observed after treatment with high - dose methylprednisolone .
three months later , on coincided with a clear increase in t2 lesions on mri , and she was again treated with methylprednisolone followed by complete remission . however , on reoccurred 3 years after disease onset , and the patient had residual visual deficit despite methylprednisolone treatment .
three months postpartum , at the age of 26 , the patient presented with psychiatric ( dysphoria , logorrhea ) and cognitive ( dyscalculia , frontal executive disorder , psychomotor slowing ) symptoms .
mri showed dramatic progression of t2 lesion load and serum was positive for low titer antinuclear antibodies .
absorption of patient serum with nmdar - transfected cells abolished the nmdar - specific signal in the live cba , whereas incubation with nontransfected cells did not , proving the specificity of antibodies against nmdar .
could be treated for an autoimmune cause of the psychiatric symptoms , she was transferred to an outside psychiatric hospital and no further clinical data were available .
demographic , clinical , and antibody data of all patients with demyelinating cns disorders with seizures and/or behavioral symptoms , which are the major initial symptoms of nmdar encephalitis , are shown in the figure .
overall , we found serum nmdar antibodies in 1 of 215 patients with cns demyelinating diseases , suggesting that the frequency of those autoantibodies is very low ( < 1% ) .
although a low false - positive rate of igg nmdar antibodies was reported recently , in our study serum igg nmdar antibodies were only present in patients with symptoms suggestive of nmdar encephalitis .
the major limitation of our study was the small number of available csf samples , which could have possibly yielded a higher number of nmdar antibody  positive patients with demyelinating diseases when using suitable testing methods .
as demonstrated here , live cba might be more sensitive for detecting low titer serum nmdar antibodies but not for testing csf samples , as currently used .
the disease course of the 1 patient with ms and nmdar antibodies was initially typical for relapsing - remitting ms with residual visual deficits after the second episode of on . because analysis of nmdar antibodies was done retrospectively , the occurrence of antibodies during the first episode of demyelination was not known at that time point .
csf was positive for nmdar antibodies at disease onset , in line with studies suggesting csf to be more sensitive than serum . with the onset of psychiatric and cognitive symptoms ,
this case demonstrates that nmdar autoantibodies can be linked with demyelination , which has been shown in a recent study investigating a cohort of 691 patients with nmdar encephalitis , identifying a subgroup of patients with demyelinating episodes .
most reported cases ( 18/23 ) of demyelinating syndromes overlapping with the presence of nmdar antibodies were positive for at least 1 antibody associated with demyelination , namely antibodies against mog or aqp4 . in our case , those antibodies were not found . of note ,
nmdar encephalitis  related symptoms occurred 3 months after the patient gave birth to a healthy child , paralleling a recently described case , although our case showed a less severe disease course .
we found nmdar antibodies to be rare in demyelinating diseases , even when focusing on patients with symptoms suggestive of nmdar encephalitis . in the case
described here , the presence of nmdar antibodies preceded the occurrence of symptoms associated with nmdar encephalitis .
thus , serum nmdar antibodies could have diagnostic value for an overlapping nmdar encephalitis  related disease course in patients with demyelinating diseases .
conceived and designed the experiments : m. ramberger , g.b . , m. reindl . performed the experiments : m. ramberger , k.s . ,
this study was supported by research grants from the fonds zur frderung der wissenschaftlichen forschung , austria ( fwf graduate program w1206 spin ) and the austrian federal ministry of science and economy ( grant big wig ms ) .
m. ramberger , g. bsteh , k. schanda , and r. hftberger report no disclosures .
k. rostsy is on the scientific advisory board for novartis and received speaker honoraria from merck - serono .
lutterotti received travel funding from european charcot foundation and fundacio gaem and received research support from austrian ms society .
f. deisenhammer is on the scientific advisory board for biogen - idec , novartis pharma , genzyme , merck , and teva ; received travel funding and/or speaker honoraria from bayer healthcare , biogen - idec , novartis , and genzyme ; was a section editor for multiple sclerosis and related disorders ; and received research support from biogen - idec and innovative medicine initiatives .
m. reindl is an academic editor for plos one , was on the editorial board for current medicinal chemistry and autoimmune diseases , and received research support from fonds zur frderung der wissenschaftlichen forschung and austrian federal ministry of science , research and economy .
reindl and medical university of innsbruck offer testing for mog antibodies for research purposes only ( free of charge ) .
the university hospital and medical university of innsbruck receive payments for antibody assays ( nmdar , aqp4 , and other autoantibodies ) and for aqp4 antibody validation experiments organized by euroimmun .
 objectives : to analyze the frequency of nmda receptor ( nmdar ) antibodies in patients with various inflammatory demyelinating diseases of the cns and to determine their clinical correlates.methods:retrospective case - control study from 2005 to 2014 with the detection of serum igg antibodies to nmdar , aquaporin-4 , and myelin oligodendrocyte glycoprotein by recombinant live cell - based immunofluorescence assays . 
 fifty - one patients with acute disseminated encephalomyelitis , 41 with neuromyelitis optica spectrum disorders , 34 with clinically isolated syndrome , and 89 with multiple sclerosis ( ms ) were included . due to a known association of nmdar antibodies with seizures and behavioral symptoms , patients with those clinical manifestations 
 were preferentially included and are therefore overrepresented in our cohort . 
 nine patients with nmdar encephalitis , 94 patients with other neurologic diseases , and 48 healthy individuals were used as controls.results:nmdar antibodies were found in all 9 patients with nmdar encephalitis but in only 1 of 215 ( 0.5% ) patients with inflammatory demyelination and in none of the controls . 
 this patient had relapsing - remitting ms with nmdar antibodies present at disease onset , with an increase in nmdar antibody titer with the onset of psychiatric symptoms and cognitive deficits.conclusion:in demyelinating disorders , nmdar antibodies are uncommon , even in those with symptoms seen in nmdar encephalitis . 
METHODS
Patients and controls.
Standard protocol approvals, registrations, and patient consents.
Live cell-based assays for antibody detection.
RESULTS
DISCUSSION
AUTHOR CONTRIBUTIONS
STUDY FUNDING
DISCLOSURE