A Guide to the Interpretation of CSF Indices

Transcription

1 Guide to the Interpretation of CSF Indices Mandolin Ziadie, MD, Frank H. Wians, Jr., PhD, MT(SCP) (Department of Pathology, The University of Texas Southwestern Medical Center, Dallas, TX) DOI: /0D1FGL7JXE3VQBY Received Revisions Received ccepted Multiple sclerosis (MS) is a disder of the central nervous system attributed to an autoimmune reaction against myelin protein. The most recent diagnostic criteria f MS combine clinical presentation, labaty test data, and radiological findings. We review the labaty tests that contribute to the diagnosis of MS, including identification of oligoclonal bands by cerebrospinal fluid (CSF) electrophesis, quantification of CSF immunoglobulin G and albumin concentrations, and the interpretation of CSF indices. fter reading this article, the reader should understand the novel pattern recognition guide as an aid in the interpretation of CSF indices. Chemistry exam questions and cresponding answer fm are located after the CE update on p Multiple sclerosis is the most common of the demyelinating disders and occurs in approximately 1/1,000 individuals in the United States population. 1 Meover, MS is me prevalent in women and young adults. The diagnosis of MS is based on histy and physical examination findings indicative of multiple demyelinating lesions of the CS that occur separately in time and anatomic site. 2,3 However, due to its highly variable clinical presentation, additional evidence from neuroimaging and labaty studies is often required to diagnose MS. The most recent diagnostic criteria f MS combine clinical presentation, labaty test data, and radiological findings. 4 We review the labaty tests that contribute to the diagnosis of MS, including identification of oligoclonal bands by CSF electrophesis, quantification of CSF immunoglobulin G and albumin concentrations, the interpretation of CSF indices (ie, lbumin Index,, IgG Synthesis Rate, and Local IgG Synthesis), and the use of a novel pattern recognition guide to aid in the interpretation of these indices. CSF Proteins and MS Cerebrospinal fluid is an ultrafiltrate of plasma composed primarily of water, electrolytes, glucose, and protein. lthough derived from the blood, the CSF is separated from it by an anatomic barrier - the blood-brain barrier (BBB) - that limits the exchange of substances between these 2 compartments (ie, the spinal canal and the bloodstream) and creates a protective environment around the CS. Due to this limited exchange, the protein concentration of the CSF is relatively low (mg/dl quantities) compared to that of serum (g/dl quantities). When the BBB is intact (ie, undamaged by disease trauma), the CSF concentration of its constituent proteins is inversely related to the protein s molecular weight and linearly related to the serum concentration of the protein. Therefe, albumin, a protein of relatively low molecular weight (~69 KDa) that constitutes nearly 50% of the total protein concentration in serum from healthy individuals, is the maj protein found in the CSF of these individuals. Conditions (eg, autoimmune inflammaty disders) that compromise the integrity of the BBB those (eg, MS) that result in increased local synthesis of proteins (eg, IgG) within the sheaths of neurons of the CS (ie, intrathecal protein synthesis) negate these relationships. In healthy individuals with an intact BBB, the ratio of CSF IgG:total protein concentration is approximately two-thirds the ratio of serum IgG:total protein concentration. n increase (ie, >0.66) in this value occurs in MS as well as in several other inflammaty CS conditions, including neurosyphilis, subacute sclerosing panencephalitis, and others. 5 In patients with MS, the abnmally increased levels of CSF immunoglobulin are a result of increased intrathecal synthesis of IgG. 6 Increased intrathecal IgG synthesis can be shown both qualitatively by demonstration of oligoclonal bands in CSF electrophesis and quantitatively both with measurement of CSF IgG concentration by immunonephelometry and with the utilization of various indices (see below) to assess the integrity of the BBB and the magnitude of intrathecal IgG synthesis. CSF Electrophesis and the Identification of Oligoclonal Bands When tandem CSF and serum specimens from healthy individuals are subjected to high resolution electrophesis isoelectric focusing, there are no protein bands in the CSF high resolution electrophesis isoelectric focusing electrophetic patterns that are not present in the cresponding serum electrophesis isoelectric focusing patterns. However, when electrophesis (HRE IEF) of tandem CSF and serum specimens from patients with MS is perfmed, additional bands may Downloaded 558 from LBMEDICIE Volume umber 9 September 2005 labmedicine.com on 06 pril 2018

2 be observed in the CSF electrophetic pattern that are not present in the cresponding serum electrophetic pattern (Figure 1). Meover, when 2 me distinct bands are present in the CSF electrophetic pattern that are absent from the patient s serum electrophetic pattern, obtained on serum tested simultaneously in an adjacent lane to the patient s CSF sample, the CSF sample is considered positive f oligoclonal bands. The presence of oligoclonal bands suggests increased intrathecal IgG synthesis, which may be suppted independently by the values f the patient s CSF indices, provided that the patient s BBB is intact, so that any increased concentration of IgG in the patient s CSF can be attributed to intrathecal synthesis and not passive diffusion from the serum into the CSF due to a damaged BBB. 7 It is imptant to remember that oligoclonal bands are detected by high resolution electrophesis isoelectric focusing in the CSF of only 60% to 80% of patients with MS. 6,7 However, a recent study in patients with confirmed MS has demonstrated that immunofixation electrophesis with immunoblotting increases the analytical sensitivity of this procedure over high resolution electrophesis f detecting oligoclonal bands from 60% (HRE) to 90% (IFE-immunoblot assay). 7 It is equally imptant to remember that oligoclonal bands can occur in the CSF of patients with disders other than MS, including subacute sclerosing panencephalitis, Guillain- BarrJ syndrome, neurosyphilis, vasculitis, and IDS. Thus, the presence of CSF oligoclonal bands by any method is not diagnostic of MS. In addition, if BBB damage is present, a common complication in long-standing MS, oligoclonal bands that may be present in the CSF can be obscured and lead to false negative results. 8 evertheless, because such a high percentage of patients with MS demonstrate oligoclonal bands upon CSF electrophesis, this method can provide valuable infmation in suppt of a diagnosis of MS. Meover, to avoid bias in the interpretation of the presence absence of oligoclonal bands by electrophetic methods, this decision should be made independently of the assessment of values f CSF indices. CSF Indices To augment the infmation provided by CSF electrophesis, various CSF indices have been developed, the most useful of which are: lbumin Index,, IgG Synthesis Rate, and Local IgG Synthesis. These indices rely on the measurement of Figure 1_IEF gel of serum and CSF samples from the same patient. The CSF sample demonstrates the presence of at least 5 oligoclonal bands that are not present in the serum sample. CSF and serum IgG and albumin concentrations and fmulas (Table 1) designed to provide values that represent the integrity of the BBB and the magnitude of intrathecal IgG synthesis. These fmulas 7-15 have been developed to crect CSF IgG concentration f the contribution from serum, especially in cases where the BBB is significantly impaired damaged, in der to determin the amount produced locally ( intrathecally) in the CS. We focus here on only those fmulas used in our institution. 16 lbumin Index Because increased levels of CSF IgG are significant only when leakage from serum can be excluded, the lbumin Index is used to assess the integrity of the BBB. lbumin is not produced intrathecally. Therefe, its concentration in the CSF is based solely on diffusion from the serum. The ratio ( quotient) of CSF:serum concentrations of albumin (Q lb ) is constant in healthy individuals with an intact BBB. 12 Damage to the BBB results in leakage of albumin from serum into the CSF and an increased value f the lbumin Index. 13 The reference interval f the lbumin Index is Thus, in a patient with an increased CSF IgG concentration, an lb Index <9.0 suggests an intact BBB and makes the possibility of significant BBB damage as the cause of an elevated CSF IgG concentration less likely. n lbumin Index >9.0 is indicative of BBB damage and increases the likelihood that an increased CSF concentration is due to leakage of serum IgG across a damaged BBB. Therefe, CSF IgG results, and any CSF indices that require them, are generally unreliable and inconclusive when the lbumin Index is abnmal. Table 1_Fmulas and Reference Intervals f CSF Indices CSF Index Fmula Reference Interval a lbumin Index Q lb x 1,000 <9.0 o significant impairment of BBB Slight impairment Moderate impairment Severe impairment >100.0 Total breakdown b Q IgG /Q lb {IgG csf (IgG s / 369) ([lb csf (lb s / 230)] x (IgG s / lb s ) x 0.43)} x mg/day {Q IgG 0.8[Q lb 2 + (15 x 10-6 )] x 10-3 } x [IgG] s <0.0 mg/dl a Reference intervals provided f each CSF index are based on non-bloody CSF samples obtained from a lumbar puncture and cannot be applied to samples taken from other regions of the spinal column. Red cell contamination (ie, >0.2%) of the CSF falsely elevates values f CSF indices and, in most cases, makes evaluation of local IgG production impossible. 10 b Q IgG = [IgG] csf / [IgG] s ; Q lb = [lb] csf / [lb] s BBB, blood-brain barrier; Q, quotient; IgG, immunoglobulin G; lb, albumin; csf, cerebrospinal fluid; s, serum; SR, synthesis rate; loc, local Downloaded labmedicine.com from September 2005 Volume 36 umber 9 LBMEDICIE 559 on 06 pril 2018

3 Similar to albumin, IgG enters the CSF from the serum. When the BBB is intact, the CSF:serum IgG ratio is relatively constant. Unlike albumin, however, IgG can also be produced intrathecally. Thus, an altered CSF:serum IgG ratio quotient (Q IgG ) can result from a damaged BBB and/ increased local IgG production. However, when an elevated CSF IgG concentration is due to a damaged BBB, values f both Q IgG and Q lb will be increased compared to values f these quotients in individuals with an intact BBB. In contrast, when the BBB is intact, an increased CSF IgG concentration is most likely due to increased intrathecal IgG production and only Q IgG, and not Q lb, will be increased. t an cut-off value of 0.70, this index has a diagnostic sensitivity f MS of ~92%, making it comparable to isoelectric focusing with immunoblotting in the detection of patients with MS. 7,11 It is imptant to note, however, that certain conditions, including some causes of BBB damage, intrathecal hemrhage, and traumatic lumbar puncture can falsely elevate the. Because these conditions do not cause Q IgG to exceed Q lb, however, they do not raise the IgG index above 1.0. Thus, all values greater than 1.0 are definitive of locally increased IgG synthesis. 10 IgG Synthesis Rate The fmula f calculating IgG synthesis rate uses constants that crect f the average nmal serum:csf ratios of IgG and albumin (ie, 369 and 230, respectively), the molecular weight ratio (ie, 0.43) of these 2 proteins, and the daily production of CSF volume (ie, the fact, 5). 8 The fmula is based on the concept that, in conditions in which BBB damage occurs, the degree of increased permeability of the BBB to albumin is directly proptional to the degree of increased permeability to IgG. By subtracting from the CSF IgG concentration, the amount of IgG due to entry into the CSF from the serum when the BBB has nmal permeability and when its permeability is increased, this fmula estimates the amount of IgG that is produced intrathecally. 15 Unftunately, this fmula has the greatest tendency to yield falsely elevated results in conditions of BBB impairment. 13 Therefe, this tendency must be kept in mind when interpreting values f IgG synthesis rate. Local IgG Synthesis The fmula f Local IgG Synthesis is used to calculate the minimum amount of local IgG synthesis in the CS, using empirically derived crection facts to account f the loss of molecular size selectivity by the BBB. 14 Similar to the, Local IgG Synthesis has high diagnostic sensitivity and a low false positive rate; however, in the presence of BBB impairment, it has an increased rate of false negative results. 13,15 Values f Local IgG Synthesis above the upper limit of nmal (ie, >0.0 mg/dl) in a patient with an increased provides strong evidence f increased intrathecal IgG synthesis. 13,17 List of bbreviations bnmal IDS cquired immunodeficiency syndrome lb lbumin BBB Blood-brain barrier CS Central nervous system CSF Cerebrospinal fluid ELP Electrophesis HRE High resolution electrophesis IEF Isoelectric focusing IFE Immunofixation electrophesis Local immunoglobulin G (synthesis) LIS Labaty Infmation System MS Multiple sclerosis mal o-bands Oligoclonal bands Q lb lbumin quotient Q IgG IgG quotient SR Synthesis rate SSPE Subacute sclerosing panencephalitis UL Upper limit of nmal Interpretation of CSF Indices Made Simple Based on the interpretation of the 4 CSF indices as either mal (values within the reference interval f the index) bnmal, with abnmal defined as only values f an index that exceed the UL f that index, the number of possible permutations of and f 4 indices is (Table 2). Using this infmation, we interpreted each of the 16 possible patterns f and f all 4 CSF indices and created a guide that provides the rationale f the interpretation of each pattern (Table 3). Using this approach, interpretation of values f CSF indices, printed on our labaty rept in the der:, lbumin Index, IgG Synthesis Rate, Local IgG Synthesis, is perfmed simply by assigning an an to the value f each index and then interpreting the 4-letter pattern accding to the infmation provided in Table 3. Once the appropriate interpretation is selected, this infmation is matched with the interpretive code used to enter this interpretation into our LIS. Summary Qualitative (identification of oligoclonal bands by high resolution electrophesis, isoelectric focusing, immunofixation electrophesis with immunoblotting) and quantitative (determination of CSF indices) methods f the evaluation of increased Table 2_ll Possible Permutations of and Values f 4 CSF Indices Pattern lb Index = nmal [within nmal reference range limit(s)]; = abnmal [exceeds UPPER LIMIT of nmal reference range; abnmally low values are unimptant in the assessment of increased intrathecal synthesis of IgG] Downloaded 560 from LBMEDICIE Volume umber 9 September 2005 labmedicine.com on 06 pril 2018

4 Table 3_Interpretation and Rationale f ll Patterns of mal and bnmal Results f CSF Indices Pattern o. Pattern Interpretation Rationale lb Index lb Index lb Index lb Index lb Index lb Index lb Index lb Index lb Index lb Index lb Index lb Index lb Index lb Index lb Index lb Index ot indicative of local b synthesis ot indicative of local b synthesis ot indicative of local b synthesis Indicative of local b synthesis Suspicious of local b synthesis Results inconclusive, Q lb abnmal Results inconclusive, Q lb abnmal Results inconclusive, Q lb abnmal Indicative of local b synthesis Indicative of local b synthesis Suspicious of local b synthesis a Indicative of local b synthesis a Indicative of local b synthesis Indicative of local b synthesis Indicative of local b synthesis Results inconclusive, Q lb abnmal b Indicative of local b synthesis b Results inconclusive, Q lb abnmal b Indicative of local b synthesis b Values f all CSF indices are nmal. lthough the lb Index is abnmal, indicating BBB impairment, values f all other indices are nmal. When values f the and are nmal, intrathecal IgG synthesis is unlikely. In this pattern, the principal indicats of no local b synthesis are the nmal values f the and. The high false positive rate associated with makes an abnmal value f this indice a po indicat of local b synthesis when the other 2 indices [ and ] are nmal. The nmal lb index indicates an intact BBB. Thus, the elevated cannot be attributed to BBB impairment. The abnmal, which has a much lower false positive rate than the, indicates a high likelihood of local b synthesis. The nmal lb index indicates an intact BBB. Thus, the elevated cannot be attributed to BBB impairment making intrathecal IgG synthesis highly likely. Without supptive evidence from the IgG index, however, this pattern can only be interpreted as "suspicious" f local b synthesis. The lb Index is abnmal, indicating an abnmal BBB (eg, due to inflammation, neoplasia, traumatic lumbar puncture, etc). Increased intrathecal IgG synthesis is suggested by the abnmal and, but the possibility that these abnmal values are due to leakage across a damaged BBB cannot be ruled out. Thus, the results are inconclusive. In general, the same is true f all patterns that combine an abnmal lb Index with 1 me additional abnmal values f the remaining indices. However, there are exceptions to this rule (cf, patterns 10, 14, 15, 16). The lb Index is abnmal, indicating BBB impairment. The abnmal suggests local b synthesis; however, in vivo contamination of the CSF with plasma due to BBB impairment cannot be excluded as the cause of the findings f the CSF indices associated with this pattern. The lb Index is abnmal, indicating BBB impairment. The combination of nmal values f and and an abnmal value f is possible in the setting of increased local b production, but the lack of supptive evidence (ie, an abnmal IgG index) restricts the interpretation to "inconclusive." ll indices suggestive of local b synthesis are abnmal in the presence of a nmal lb Index (intact BBB). These findings are indicative of intrathecal ( local) synthesis of IgG. The lb Index is abnmal, indicating BBB impairment. lthough BBB impairment may cause a false positive, the probability that values f both the and are abnmal due to BBB impairment is low. Therefe, local b synthesis is likely. When only the value f the is abnmal, the degree of abnmality is key. n greater than the upper limit of nmal (0.7) but less than a cutoff value of 1.0 is suspicious f local b synthesis, despite the nmal values f and. n IgG index value =1.0 is incontrovertible evidence f local b synthesis, even in the presence of hemrhage BBB impairment. Values f both and are abnmal with a nmal lb Index (intact BBB.) The combination of abnmal values f these 2 indices [ and ] has a low false positive rate. Therefe, local b synthesis is likely. Values f both the and are abnmal with a nmal lb Index (intact BBB). This pattern is consistent with increased local b synthesis. The lb Index is abnmal, indicating BBB impairment. The probability that values f both and are abnmal solely due to BBB impairment is low. Therefe, local b synthesis is likely. The lb Index is abnmal, indicating BBB impairment. BBB impairment has been associated with increased rates of false positive and false negative IgG (loc) values. Thus, the abnmal and nmal values are inconclusive in the assessment of local b synthesis. However, when the IgG index value is >1.0, there is incontrovertible evidence f local b synthesis, even in the presence of hemrhage BBB impairment. The lb Index is abnmal, indicating BBB impairment. nmal value with an abnmal lb Index is not reliable in the assessment of local b synthesis. Therefe, this pattern must be interpreted as inconclusive with regard to local b synthesis. However, when the IgG index value is >1.0, there is incontrovertible evidence f local b synthesis, even in the presence of hemrhage BBB impairment. a If is , interpret pattern as "Suspicious f local b synthesis;" when the IgG index is =1.0, interpret pattern as "Indicative of local b synthesis." b If the IgG index value is =1.0, interpret pattern as "Indicative of local b synthesis." Otherwise, interpret as "Results inconclusive, Q lb abnmal." = nmal [within nmal reference range limit(s)]; = abnmal [exceeds UPPER LIMIT of nmal reference range; abnmally decreased values f any index are unimptant in the assessment of increased intrathecal IgG synthesis]; BBB, blood-brain barrier; = immunoglobulin G synthesis rate; = local (ie, intrathecal) IgG synthesis. 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5 Example (to illustrate manual calculation of each of the 4 CSF Indices using the fmulas provided in Table 1 and the Interpretive guide provided in Table 3) [lb] s = 4.4 g/dl = 4400 mg/dl [IgG] s = 950 mg/dl [lb] csf = 44 mg/dl [IgG] csf = 8.4 mg/dl Q lb = [lb] csf / [lb] s = 44 / 4400 = Q IgG = [IgG] csf / [IgG] s = 8.4 / 950 = lb Index = Q lb x 1000 = x 1000 = 10.0 = Q IgG / Q lb = / = = {IgG csf (IgG s / 369) ([lb csf (lb s / 230)] x (IgG s / lb s ) x 0.43)} x 5 = {[8.4 (950 / 369)] ([44 (4400 / 230)] x (950 / 4400) x 0.43)} x 5 = {[ ] ([ ] x (0.216) x 0.43)} x 5 = {[5.826] ([24.87] x x 0.43)} x 5 = { } x 5 = {3.516} x 5 = mg/day = {Q IgG 0.8[Q lb 2 + (15 x 10-6 )] x 10-3 } x [IgG] s = { [(0.0100) ] } x 950 = { [ ] } x 950 = { [ ] } x 950 = { ( ) } x 950 = { } x 950 = { } x 950 = mg/dl Pattern lb Index = 10.0 = bnmal = 0.88 = bnmal = 17.6 mg/day = bnmal = 2.0 mg/dl = bnmal Using the Interpretive Guide provided in Table 3, this pattern (#10) is consistent with local ( intrathecal) IgG synthesis. The interpretive comments included on the patient s rept are: indicative of local b synthesis clinical crelation required IgG concentration in CSF are useful tools in the evaluation of patients suspected of having MS. Used together, but interpreted independently, they provide clinicians with useful infmation to complement patient histy, clinical, imaging, and other labaty data in such patients. To assist clinicians and labaty professionals in the assessment of values f CSF indices, we developed a simple and practical guide to perfm this function that provides the rationale f the interpretation of all possible combinations of nmal and abnmal values f these indices. Of course, the reliability of the infmation provided by this guide is critically dependent on the accuracy of the values f the CSF indices calculated, in most labaties, using fmulas incpated into the labaty s LIS. LM cknowledgements: We gratefully acknowledge the contributions of Drs Daniel Fajardo and Dennis Wooten to the creation of the Interpretive Guide f the Interpretation of CSF Indices shown in Table oonan CW, Kathman SJ, White MC. Prevalence estimates f MS in the United States and evidence of an increasing trend f women. eurol. 2002;58: Frosch MP, nthony DC, Girolami UD. The central nervous system. In: Kumar V, bbas K, Fausto, eds. Robbins and Cotran Pathologic Basis of Disease, 7 th ed. Philadelphia: Elsevier, 2005: Ferri FF, ed. Ferri s best test: a practical guide to clinical labaty medicine and diagnostic imaging. Philadelphia: Elsevier; 2004: McDonald WI, Compston, Edan G, et al. Recommended diagnostic criteria f multiple sclerosis: Guidelines from the International Panel on the diagnosis of multiple sclerosis. nn eurol. 2001; 50: Fishman R. Cerebrospinal fluid in diseases of the nervous system. 2nd ed. Philadelphia: W.B. Saunders Company; Hershey L, Trotter JL. The use and abuse of the cerebrospinal fluid IgG profile in the adult: a practical evaluation. nn eurol. 1980;4: Ftini S, Sander EL, Weinshenker BG, et al. Cerebrospinal fluid oligoclonal bands in the diagnosis of multiple sclerosis. m J Clin Pathol. 2003;120: Tourtellote WW, Staugaitis SM, Walsh MJ, et al. The basis of intra-bloodbrain-barrier IgG synthesis. nn eurol. 17:21-27, Olek MJ. Diagnosis, features, and prognosis of MS. In: Olek MJ, ed. Multiple sclerosis: etiology, diagnosis, and new treatment strategies. Totowa, H: Humana Press; 2005: Peter JB, Bowman RL, Boman Jr. RL, et al. Blood plasma contamination of CSF: Effect on CS IgG synthesis rate and IgG index [abstract]. mer J Clin Pathol. 1987; 87: Blennow K, Fredman P, Wallin, et al. Fmulas f the quantitation of intrathecal IgG production: their validity in the presence of blood-brain barrier damage and their utility in multiple sclerosis. J eurol Sci. 1994;121: Tibbling G, Link H, Ohman S. Principles of albumin and IgG analyses in neurological disders. I. Establishment of reference values. Scand J Clin Lab Invest. 1977; 37: Peter J, Bowman RL. Intra-blood-brain barrier synthesis of IgG: comparison of IgG synthesis fmulas in a computer model and in 1,629 consecutive specimens. eurol. 1992;42: Reiber H, Felgenhauer K. Protein transfer at the blood cerebrospinal fluid barrier and the quantitation of the humal immune response within the central nervous system. Clin Chim cta. 1987; 163: Syndulko K, Tourtellotte WW, Conrad J, et al. Multiple Sclerosis Study Group, lpha Interferon Study Group, zathioprine Study Group. Transblood-brain-barrier albumin leakage and comparisons of intrathecal IgG synthesis calculations in multiple sclerosis patients. J eurimmunol. 1993;46: Wians FH Jr, Baskin LB. Electrophetic methods f the evaluation of proteins in human body fluids. Diag Endo Metab. 1998;16: Souverign JHM, Smith WG, Peet R, et al. Intrathecal IgG synthesis: Its detection by isoelectric focusing and IgG index. J eurol Sci. 1989;93: Downloaded 562 from LBMEDICIE Volume umber 9 September 2005 labmedicine.com on 06 pril 2018