Multiple Sclerosis Journal - October 2017 - 1498

Multiple Sclerosis Journal 23(11)
both quantitatively by calculating the IgG index
([CSF/serum quotient IgG]/Q alb; reference < 0.7)20
and qualitatively by the determination of oligoclonal
bands (OCB) using isoelectric focusing followed by
immunoblotting.21
KFLC and LFLC were measured in serum and CSF by
immunonephelometry (Freelite®, The Binding Site
Group Ltd., Birmingham, UK) on a BNII analyser
(Siemens Healthcare Diagnostics, Marburg, Germany).
We determined the CSF/serum quotients (Q FLC) of
KFLC and LFLC and calculated the indices of KFLC
and LFLC by correcting for the albumin quotient
(index FLC = Q FLC/Q alb). Next, the proportion of
kappa to lambda was determined in serum and CSF
separately, that is, [serum KFLC]/[serum LFLC] and
[CSF KFLC]/[CSF LFLC].
MRI
All patients underwent MRI of the brain at baseline on
a 3 T Tim Trio system (Siemens Medical Systems,
Erlangen, Germany) using a 12-element phased-array
head coil. For structural imaging we used a T1-weighted
three-dimensional (3D) Magnetization Prepared Rapid
Acquisition Gradient Echo (MPRAGE) sequence (repetition time (TR)/echo time (TE)/inversion time (TI)/
flip angle (FA) = 1.9 s/2.19 ms/0.9 s/9°; isotropic resolution = 1 mm) and a T2-weighted two-dimensional (2D)
fast fluid-attenuated inversion recovery (FLAIR)
sequence (TR/TE/TI = 9000 ms/70 ms/2500 ms, in plane
resolution = 0.9 × 0.9 mm2, slice thickness = 3 mm). A
follow-up MRI scan using the identical imaging protocol was performed in 56 (91.8%) patients.
We determined the following morphologic markers of
progression of tissue damage as previously
described.22,23 In brief, for assessing T2 lesion load
(T2LL), MS lesions were outlined on a transparency
overlaid on hard copies of the FLAIR sequence.
Using these templates, lesion masks were then created
using the DISPImage program.24 To measure the percentage of brain volume change (PBVC), we co-registered follow-up MRI scans from each individual to
baseline MRI, applying SIENA.25
All imaging analyses were performed by trained and
experienced MRI technicians, blinded to clinical data.
Statistical analyses
Statistical analyses were performed using SPSS
Statistics (version 22.0, IBM Corp. Armonk, NY,

1498

USA) and GraphPad Prism (version 5.00, GraphPad
Software, San Diego, CA, USA).
All variables were tested for normal distribution
using the Kolmogorov-Smirnov test. Group differences were determined by either chi-square test for
categorical data or Mann-Whitney U test for continuous variables. Differences between more than
two groups were defined by applying Kruskal-
Wallis test followed by post hoc Dunn's multiple
comparison test.
We performed Spearman correlations to determine
the correlation coefficients for serum and CSF variables with demographic, clinical and MRI data.
Pearson partial correlations corrected for time
between two MR images taken were applied on longitudinal MRI data. Significance level was set at 5%
(p < 0.05). The diagnostic value was determined for
index KFLC by applying the empirically well-established threshold published for potential diagnostic
use (index KFLC > 5.95) on our data.
The prognostic value of FLC was determined by comparing CIS-CDMS converters versus CIS non-converters by binary logistic and Cox regression analyses,
and hazard graphs.
Results
Subject description
The patient group (n = 61) consisted of 48 patients
with CIS and 13 patients with RRMS. The control
group (n = 60) consisted of 14 patients with cranial/
peripheral palsy (non-inflammatory neurological disease controls), 31 with headache and 15 with sensory
disturbance (both symptomatic controls). The demographic and clinical data of both groups are shown in
Table 1. Patients and controls were comparable
regarding age and gender distribution.
CSF samples were obtained at the time of diagnosis in
CIS. For MS patients, median disease duration at the
time of sample collection, that is, time between presentations of first symptoms and lumbar puncture, was
1.2 years (IQR, 0.8-4.2 years). Active disease (clinical attack within 14 days prior to lumbar puncture)
was present in 33 (54.1%) patients. Among all CIS
patients 23 (47.9%) converted to CDMS during follow-up (median, 4.8 years; IQR, 1.5-6.5 years).
Active disease at the time of lumbar puncture was
present in 13 of 23 converters and 14 of 25 non-converters. Diagnostic variables of CSF and serum

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