(2007) 526–530

Clinical Biochemistry 40

Antibodies to β2-glycoprotein-I: Relation of anticardiolipin antibodieswith clinical and laboratory parameters in patients with systemic

lupus erythematosus

Mehmet Sahin a,⁎, Nursen Duzgun b, Sevket Ercan Tunc a, Huseyin Tutkak b

a Department of Internal Medicine, Division of Rheumatology, Faculty of Medicine, Suleyman Demirel University, 32020 Isparta, Turkeyb Department of Clinical Immunology and Rheumatology, Faculty of Medicine, Ankara University, Ankara, Turkey

Received 26 March 2006; received in revised form 7 November 2006; accepted 9 December 2006Available online 14 February 2007

Abstract

Objectives: There are controversial reports on the frequency of antiphospholipid antibodies (aPL) in patients with systemic lupuserythematosus (SLE). Thus, we aimed to determine the frequency and clinical importance of aPL isotypes in Turkish patients with SLE.

Design and methods: Fifty-nine patients with SLE and 41 healthy controls were included. Serum aPL levels were measured both in patientsand healthy subjects by ELISA.

Results: Fifteen of the patients with SLE had the antiphospholipid syndrome (APS) (25.4%). The percentage of anticardiolipin antibody(aCL)-positive SLE patients among all patients was 56%. At least one isotype of anti-β2-glycoprotein I (β2-GPI) antibody was positive in 83% ofpatients. The positivity rates of aCL and anti-β2-GPI antibodies in patients with or without APS were higher than the healthy controls. There werepositive correlations between isotypes of IgM aCL, IgG and IgM anti-β2-GPI and manifestations of APS.

Conclusion: It seems that the isotypes of IgM aCL, IgG and IgM anti-β2-GPI are correlated with manifestations of APS. They may play a rolein pathogenesis and may be helpful in establishing the diagnosis.© 2007 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Keywords: Anticardiolipin antibodies; Antiphospholipid antibodies; Antiphospholipid syndrome; Anti-β2-GPI antibodies; Systemic lupus erythematosus

Introduction

Antiphospholipid antibodies (aPLs) comprise a heteroge-neous group of immunoglobulins that are characterized by theirability to bind anionic phospholipids. The aPL directed towardsphosphatidylserine, phosphatidylethanolamine, phosphatidyl-choline, phosphatidylglycerin, phosphatidylinositol has beeninvestigated [1,2].

Antiphospholipid antibodies have been shown to beassociated with manifestations including arterial and venousthrombosis, recurrent fetal loss and thrombocytopenia inantiphospholipid antibody syndrome (APS) [3,4]. APS hasbeen classified as a primary disease without evidence of

⁎ Corresponding author. Fax: +90 246 2370240.E-mail address: mehmetsahin66@hotmail.com (M. Sahin).

0009-9120/$ - see front matter © 2007 The Canadian Society of Clinical Chemistsdoi:10.1016/j.clinbiochem.2006.12.013

systemic lupus erythematosus (SLE) or other collagenvascular diseases and secondarily, mainly associated withSLE.

It is known that the aPLs are directed not only againstphospholipids but also against a complex of the phospholi-pid-binding plasma proteins (cofactors), such as β2-glyco-protein I (β2−GPI) [5]. The antibody directed against thiscofactor can be detected by ELISA in the absence of phos-pholipids [5,6].

Ethnicity plays a role on the frequency and clinicalimportance of these autoantibodies. Few studies related toaPLs have been done in Turkish patients with SLE [7,8]. In thisstudy, IgA, IgG and IgM isotypes of aCL and anti-β2-GPIantibodies were studied in healthy controls and in SLE patientswith and without APS. In addition, the relationship of APSmanifestations with the presence of aCL and anti-β2-GPIantibodies was investigated.

. Published by Elsevier Inc. All rights reserved.

Table 2Average aPL results (mean±SD) (U/mL) and their statistical comparisons incontrols and patients with SLE

Isotype (U/mL) Control group(n: 41)

SLE (APS−)(n: 44)

SLE (APS+)(n: 15)

IgG-aCL 22.9±13.1 54.8±111.9 48.4±54.4IgM-aCL 16.1±14.3 43.7±77.3 a 63.3±108.2 b

IgA-aCL 21.4±17.1 74.0±135.7 c 55.8±43.6 c

IgG-anti-β2-GPI 1.9±1.4 9.95±22.2 b 15.4±33.3 b

IgM-anti-β2GPI 1.0±0.8 3.9±6.87 16.0±32.4 a

IgA-anti-β2-GPI 1.4±1.1 3.55±3.57 c 5.13±12.2

aPL: antiphospholipid antibody, SLE: systemic lupus erythematosus, APS:antiphospholipid syndrome, aCL: anticardiolipin antibody, anti-β2-GPI:β2-glycoprotein-I antibody.a p<0.05 compared to control group.b p<0.01 compared to control group.c p<0.001 compared to control group.

527M. Sahin et al. / Clinical Biochemistry 40 (2007) 526–530

Materials and methods

Patients

Fifty-nine patients with SLE who fulfilled the AmericanCollege of Rheumatology classification criteria [9] wereincluded in the study. There were 53 (89.8%) females and 6(10.2%) males, and the mean age of patients was 35.0±11.4years (range 14–62). Forty-one age- and sex-matched healthycontrols were also studied. Among the healthy subjects, 35 werefemales (85.4%) and 6 were males (14.6%), and the mean agewas 31.3±9.6 years (range 17–56). Clinical data regardingmanifestations of SLE and APS were collected retrospectivelyfrom medical records. Forty-seven patients were receivingprednisolone <15 mg/day; azathioprine <150 mg/day and/ororal anticoagulant therapy. Exclusion criteria included thepresence of pregnancy and the aged patients younger than 16.This study was designed as a cross-sectional, controlled studyconducted in the department of immunology and rheumatology.The study was approved by the ethics committee, and writteninformed consent was obtained from each patient and healthysubject.

Methods

Antiphospholipid antibodies were determined by ELISA forcardiolipin (CL) and for β2-GPI (Orgentec Diagnostika GmbH,Mainz, Germany) according to manufacturer's recommenda-tions. Sera were stored at −20 °C before the measurements.The calculation of the cut-off units was done according to themanufacturer's instructions. Briefly, microtiter plates coatedwith CL and β2-GPI and then standard solution were used. Thesera of patients and controls were diluted as 1:100 and addedinto each well coated with CL and β2-GPI. A blank wellcoated with standard solution was used. Each specimen wasincubated at room temperature (20–25 °C) for 30 min. Afterwashing with nondetergent buffer, isotype-specific enzyme(horseradish peroxidase) conjugates were added and plateswere washed again (four times). Tetramethyl benzidine inacetate buffer with hydrogen peroxide was added as achromogen and incubated at room temperature for 10 min.

Table 1Clinical and laboratory features of patients SLE (with or without APS)

SLE (all patients) SLE/APS

n: 59 (%) n: 15 (%)

Livedo reticularis 15 (25.4) 12 (80)Hemolytic anemia 7 (11.9) 5 (33.3)Neurological finding 9 (15.3) 4 (26.7)Migraine 8 (13.6) 5 (33.3)Venous thrombosis 12 (20.3) 10 (66.7)Fetal loss 15 (25.4) 11 (73.3)Thrombocytopenia 17 (28.8) 12 (80)aPTT (>36 Sn) 10 (16.9) 7 (46.7)Anti-ds-DNA (>7 IU/mL) 37 (62.7) 10 (66.7)

SLE: systemic lupus erythematosus, APS: antiphospholipid syndrome, aPTT:activated partial thromboplastin time.

The reaction was stopped with phosphoric acid for determiningthe isotypes of aCL and stopped with hydrochloric acid fordetermining the isotypes of anti β2-GPI. The opticalabsorbance measured at 450 nm by Easy Reader. The valuesof IgA, IgG, and IgM aCL and anti-β2-GPI antibodies wereexpressed as units compared with those of the standard serum.A positive result indicated a value in units more than 3 SDover the mean value obtained with control sera (41 healthyvolunteers). Antibodies to DNA (anti-dsDNA) were detectedusing radioimmunoassay (Amerlex, Amersham England) [10].The lupus anticoagulant was detected by the prolongation ofactivated partial thromboplastin time (aPTT) in a mixing test,using the 1/5 diluted aPTT reagent (Automated aPTT, GeneralDiagnostics, Durham, NC, USA), which was based on themethods of Colaco and Elkon [11].

Statistical analyses

All results are expressed as means±SD. Data were ana-lyzed using the Mann–Whitney U test and Student's t tests.P-values of less than 0.05 were regarded as significant.Analysis of correlations was performed using Spearman's rankcorrelation test. Data were analyzed using the SPSS statisticalprogram (version 9.05 software, SPSS Inc., Chicago, Illinois,USA).

Results

The clinical and laboratory features of patients with SLE(with or without APS) were shown in Table 1. Fifteen patientshad the diagnosis of secondary APS according to the proposedcriteria [12].

The distribution and relationship of aCL, and anti-β2-GPIantibodies in controls and SLE patients with and without APSwere shown in Table 2. IgA isotype of aCL and anti-β2-GPIantibodies were significantly (p<0.001) higher in the patientsthan in the controls. IgG anti-β2-GPI values were also found tobe statistically (p<0.01) higher in the patient group than in thecontrol group. Besides, compared to controls, IgM aCL(p<0.01) and IgM anti-β2-GPI (p<0.05) values were also

Table 3The value of aCL and anti-β2-GPI antibodies in cases with and without APS

aCL Anti-β2-GPI APS (n)%

(+) (−)

+ + 5 (33.3) 8 (18.2)+ − 1 (6.07) 7 (16.5)− + 6 (40) 5 (11.3)− − 3 (20) 24 (54)Total 15 (25.4) 44 (74.6)

APS: antiphospholipid syndrome, aCL: anticardiolipin antibody, anti-β2-GPI:β2-glycoprotein-I antibody.

528 M. Sahin et al. / Clinical Biochemistry 40 (2007) 526–530

significantly higher in the patient group. IgG aCL values werehigher in the patient group as compared to the control groupalthough the difference was not statistically significant(p>0.05).

Among the 59 patients, aCL antibodies were found in 33patients (56%) and anti-β2-GPI antibodies were found in 49patients (83%). Eight patients (13.6%), 13 patients (22%) and12 patients (20.3%) showed IgG aCL, IgM aCL and IgA aCLpositivity, respectively. The positivity numbers of IgG, IgA andIgM anti-β2-GPI antibodies were 21 (35.6%), 17 (28.8%) and11 (18.6%), respectively. The number of all positive isotypes ofaCL was only 3 (5.08%) although anti β2-GPI was found in 9patients (15.25%). The positivity of all isotypes in aCL was notseen in patients with SLE/APS although all isotypes of ant-β2-GPI antibodies were positive in 2 (14%) patients.

General distributions of aCL and anti-β2-GPI antibodies inpatients with and without APS were shown in Table 3. Only onepatient with APS (6.07%) had positive aCL and negative anti-β2-GPI antibodies. Six patients with APS (40%) had negativeaCL and positive anti-β2-GPI antibodies. Of these patients 9(60%) had positive aCL and 11 (73.3%) had positive anti-β2-GPI antibodies, regardless of isotypes.

The relationship of clinical and laboratory findings with aPLisotypes in SLE patients was shown in Table 4. The statisticalcorrelations were found between IgA aCL and migraine. Therewas also a correlation between IgM aCL and both IgM anti-β2-GPI and livedo reticularis (p<0.001).

Discussion

We evaluated the frequencies of aCL and anti-β2-GPIantibodies as well as isotype distribution and their correlations

Table 4Statistical comparison of aPL isotypes with findings of APS

Finding IgA-aCL IgG-aCL IgM-aCL

Livedo reticularis – – <0.001Recurrent abortion – – <0.05Thrombosis – – <0.05Neurological disorder – – –Migraine <0.001 – <0.01ThrombocytopeniaHemolytic anemia – – –Prolonged aPTT – – <0.05

aPL: antiphospholipid antibody, APS: antiphospholipid syndrome, aCL: anticardiolthromboplastin time.

with the main manifestations of the APS in Turkish patientswith SLE. In different populations, the average prevalence ofaPL in patients with SLE varies between 17% and 61%[4,13,14]. The IgG aCL in SLE was found to be more than bothIgM and IgA aCL antibodies. We determined that theprevalence of IgG aCL in the present study was lower than inthe previous studies [15,16] although IgM aCL was thepredominant isotype and the prevalence of IgA aCL was higherthan the other series [15,16]. The difference of results may bedue to the modality used in the measurement of antibodies, tothe study population, ethnic–racial differences and variation ofpositive aCL criteria with respect to investigators [17]. To ourknowledge, scarce studies are present analyzing the prevalenceof these autoantibodies in different ethnic groups. IgA aCLlevels were also found highly positive (61%) in AfricaCaribbean patients with SLE [18]. In contrast to our study, in91 Chinese patients with SLE a low prevalence was found inIgM and IgA aCL as 1.1% and 4.4%, respectively [19]. Theresults suggested that genetic and environmental factors may beresponsible for the different reports in the different ethnicities orgeographic populations [18]. Similarly, there are publishedseries which suggested that aCL levels were correlated with thedisease activity [14,15].

Recently, it has been reported that the detection of antibodiesto β2-GPI is a better indicator in APS manifestations than thedetection of aCL [15]. Antibodies against β2-GPI have beenreported to have a percentage from zero to 90 in patients withAPS [20–22], and the percentage has been related with variousclinical findings. We found at least one isotype positivityagainst anti-β2-GPI in 49 of 59 SLE patients (83%). However,all three isotypes of anti-β2-GPI in SLE patients were positivein only 9 patients (15.25%). The frequencies of anti-β2-GPIantibodies and its isotypes in our patients were similar to a well-known report [21]. There was a higher frequency in IgG isotypeof anti-β2-GPI antibody in our patients compared to IgM andIgA isotypes of anti-β2-GPI antibody. Fanopoulos et al. [15]reported that the frequency of anti-β2-GPI isotypes, particularlyIgA isotype was higher in patients with SLE than the controlgroup. Nevertheless, the difference in the frequency and levelsbetween other reports and the current study may be attributed todisease activity. It has been reported that some patients withmanifestations of APS had reference levels of aCL althoughthey have abnormal levels of anti-β2-GPI [9,23–25]. In thepresent study, anti-β2-GPI antibodies were positive in 11

IgA-anti-β2-GPI IgG-anti-β2-GPI IgM-anti-β2-GPI

– <0.01 <0.001– – <0.05– <0.05 <0.05– – –– <0.05 <0.05

<0.05– – –– <0.05 –

ipin antibody, anti-β2-GPI: β2-glycoprotein-I antibody, aPTT: activated partial

529M. Sahin et al. / Clinical Biochemistry 40 (2007) 526–530

(28.4%) of 38 (64.4%) aCL-negative SLE patients. It has beenreported that anti-β2-GPI can be accepted as a specific markerboth in SLE and APS [20,22,25].

In this study, the relation of aPLs with manifestations in APSwas also evaluated. We found that the antibodies may have animportant role in pathogenesis of clinical and laboratorymanifestations in patients with SLE and APS. Increased IgMaCL, IgM and IgG anti-β2-GPI antibodies were related with aclinical history of thrombosis. Thrombosis was specificallyassociated with IgA aCL [26], IgG aCL [27] and all aPL [28],whereas in an other study no apparent relation to titer was alsoobserved [29]. Treatment regimes may have led to thesedifferent results. Several cases have been reported, in whichaCL titers have been shown to fluctuate markedly over time[30].

It has been reported that the detection of anti-β2-GPIantibodies has a positive association with thrombotic events inpatients with primary or secondary APS (with SLE) than thedetection of aCL antibodies [5,25]. It inhibits intrinsic clottingpathway and ADP-induced platelet aggregation [25]. It isproposed that anti-β2-GPI antibodies may be used as ascreening test in evaluation of the risk of thrombosis in patientswith APS [31]. Similarly, Inanc et al. [32] found that IgG anti-β2-GPI antibodies are more frequent in SLE patients withthrombotic/neurological events. They proposed that IgG anti-β2-GPI may be a marker for prediction of thrombotic/neurological events in SLE. In another study, a correlationwas demonstrated between IgM anti-β2-GPI isotype andthrombosis [33]. Also, an association between IgA anti-β2-GPI and thrombosis was shown [22]. In this study, IgM β2-GPIwas higher in patients with SLE/APS than the other isotypesand IgM and IgG anti-β2-GPI antibodies were associated withthe presence of thrombosis. These results suggested that IgMand IgG β2-GPI antibodies may play a role in the pathogenesisof thrombosis and may be useful to predict thrombosisformation.

Another manifestation of APS suggesting that aPLs play arole in pathogenesis is fetal loss [4]. In many studies, there wasa correlation between isotypes of aCL (especially IgG) and fetalloss [20,34]. They suggested that aCL positivity increases theincidence of fetal loss in patients with SLE and determinationof aCL titer is useful in estimation of fetal loss in SLE [14,32].Recurrent abortion occurred in 25.4% of the patients with SLEand it was related with IgM-aCL and the similar result wasfound by the study of Day et al. [24]. In most published paperson the subject, the number of patients is small and pregnancydata are insufficient. In most studies an association betweenanti-β2-GPI antibodies and fetal loss in patients with SLE andprimary APS was reported [35]. In this study, we found acorrelation between IgM anti-β2-GPI antibody and fetal loss.Mok et al. [28] also showed a relation between IgM anti-β2-GPI antibodies and preeclampsia. Our data suggested that IgManti-β2-GPI antibody may play a role in the pathogenesis ofabortus.

Livedo reticularis is one of the clinical manifestations ofAPS. We found statistically significant correlation betweenlivedo reticularis and IgM aCL, IgM and IgG anti-β2-GPI in

SLE patients. The fact that there is a prominent associationbetween livedo reticularis and particularly IgM isotypes of twoaPL groups in our study suggested that these antibodies mayplay a role in the pathogenesis of livedo reticularis.

In conclusion, we found higher levels of aPLs in Turkishpatients with SLE than the healthy controls. Especially, anti-β2-GPI antibodies were found more frequently than aCLs inthe patients with SLE (with or without APS). Different reportsin the papers may reflect the effect of ethnicity and it may havean important effect in controlling aPL production and clinicalrelevance. However, IgM aCL, IgG and IgM anti-β2-GPIisotypes may be helpful in diagnosis of SLE patients withAPS.

References

[1] Mc Neıl HP, Chesterman CN, Krılıs SA. Immunology and clinicalimportance of antiphospholipid antibodies. Adv Immunol 1991;49:193–279.

[2] von Landenberg P, Schölmerich J, Kempis JV, Lackner KJ. Thecombination of different antiphospholipid antibody subgroups in the seraof patients with autoimmune disease is a strong predictor for thrombosis.Immunobiology 2003;207:65–71.

[3] Sammaritano LR, Gharavi AE, Lockshin MD. Antiphospholipid antibodysyndrome: immunologic and clinical aspects. Semin Arhritis Rheum1990;20(2):81–96.

[4] Love PE, Santoro SA. Antiphospholipid antibodies: anticardiolipin and thelupus anticoagulant in systemic lupus erythematosus (SLE) and in nonSLE disorders. Ann Intern Med 1990;112:682–98.

[5] Inanç M, Radway-Bright EL, Isenberg DA. β2-Glycoprotein I andantiβ2glycoproteýn I antibodies: where are we now? Br J Rheumatol1997;36:1247–57.

[6] Matsuura E, Igarashi Y, Yasuda T, Triplett DA, Koike T. Anticardiolipinantibodies recognize β2-glycoprotein I structure altered by interacting withoxygen modified solid phase surface. J Exp Med 1994;179:457–62.

[7] Sahin M, Duzgun N, Tunc SE, Tutkak H. Clinical manifestations andantiphosphatidylserine antibodies in patients with systemic lupus erythe-matosus: is there an association? Clin Rheumatol 2007;26:154–60.

[8] Tokay S, Direskeneli H, Yurdakul S, Akoglu T. Anticardiolipin antibodiesin Behcet's disease: a reassessment. Rheumatology (Oxford) 2001;40:192–5.

[9] Tan EM, Cohen ES, Fries CF, Masi AT. The 1982 revised criteria for theclassification of systemic lupus erythematosus. Arthritis Rheum 1982;25:1271–7.

[10] Clarke MC, Carr R, Burdash NM, Chen Z, Ainsworth SK. A comparisonof three anti-double stranded DNA antibody assays on sera from SLE andother diseases. Diagn Immunol 1986;4:288–93.

[11] Colaco CB, Elkon KB. The lupus anticoagulant: a disease marker inantinuclear antibody negative lupus that is cross-reactive with autoanti-bodies to double-stranded DNA. Arthritis Rheum 1985;28:67–74.

[12] Wilson WA, Gharavi AE, Koike T, et al. International consensus statementon preliminary classification criteria for definite antiphospholipidsyndrome: report on international workshop. Arthritis Rheum 1999; 42:1309–11.

[13] Petri M. Epidemiology of the antiphospholipid antibody syndrome.J Autoimmun 2000;15:45–151.

[14] Sanfilippo SS, Khamashta MA, Atsumı T, et al. Antibodies to β2-glycoprotein I: a potential marker for clinical features of antiphospholipidantibody syndrome in patients with systemic lupus erythematosus.J Rheumatol 1998;25:2131–4.

[15] Fanopoulos D, TeodorescuMR, Varga J, TeodorescuM. High frequency ofabnormal levels of IgA anti-β2-glycoprotein I antibodies in patients withsystemic lupus erythematosus: relationship with antiphospholipid syn-drome. J Rheumatol 1998;25:675–80.

[16] Merkel PA, Chang Y, Pierangali SS, et al. The prevalence and clinical

530 M. Sahin et al. / Clinical Biochemistry 40 (2007) 526–530

associations of anticardiolipin antibodies in a large inception cohort ofpatients with connective tissue disease. Am J Med 1996;101:576–83.

[17] Aquirre V, Cuchacovich R, Barria L, et al. Prevalence and isotypedistribution of antiphospholipid antibodies in Chilean patients withsystemic lupus erythematosus. Lupus 2001;10:75–80.

[18] Molina JF, Gutıerrez-Urena S, Molina J, et al. Variability of anticardiolipinantibody isotype distribution in 3 geographic populations of patients withsystemic lupus erythematosus. J Rheumatol 1997;24:291–6.

[19] Wong KL, Liu HW, Ho K, Chan K, Wong R. Anticardiolipin antibodiesand lupus anticoagulant in Chinese patients with systemic lupuserythematosus. J Rheumatol 1991;18:1187–92.

[20] Amengual O, Atsumi T, Khamashta MA, Koike T, Hughes GRV.Specificity of ELISA for antibody to β2 glycoprotein-I in patients withantiphospholipid syndrome. Br J Rheumatol 1996;35:1239–43.

[21] Lockshin MD. Antiphospholipid antibody. JAMA 1997;277:1549–51.[22] Tsutsumı A, Matsuura E, Ichıkawa K, Fujısaku A, MukaıM, Koıke T. IgA

class anti β2-glycoprotein I in patients with systemic lupus erythematosus.J Rheumatol 1998;25:74–8.

[23] Cabiedes J, Cabral AR, Alarcon-Segovia D. Clinical manifestations of theantiphospholipid syndrome in patients with systemic lupus erythematosusassociate more strongly with anti β2-glycoprotein I than with antipho-spholipid antibodies. J Rheumatol 1995;22:1899–906.

[24] Day MH, Thıagarajan P, Ahn C, Reveılle JD, Tınker KF, Arnett FC.Autoantibodies to β2-glycoprotein I in systemic lupus erythematosus andprimary antiphospholipid antibody syndrome: clinical correlations incomparison with other antiphospholipid antibody tests. J Rheumatol1998;25:667–74.

[25] Teixido M, Font J, Reverter JC, et al. Anti β2-glycoprotein I antibodies: auseful marker for the antiphospholipid syndrome. Br J Rheumatol1997;36:113–6.

[26] Moore JE, Mohr CF. Biologically false positive serologic test for syphilis.JAMA 1952;150:467–73.

[27] Harris EN, Chan JK, Asherson RA, Aber VR, Gharavi AE, Hughes GRV.

Thrombosis, recurrent fetal loss and thrombocytopenia. Predictive value ofthe anticardiolipin test. Arch Intern Med 1986;146:2153–6.

[28] Mok MY, Chan EY, Fong DY, Leung KF, Wong WS, Lau CS. Antipho-spholipid antibody profiles and their clinical associations in Chinesepatients with systemic lupus erythematosus. J Rheumatol 2005;32:622–8.

[29] Loizou S, Cofiner C, Weetman AP, Walport MJ. Immunoglobulin class andIgG subclass distribution of anticardiolipin antibodies in patients withsystemic lupus erythematosus and associated disorders. Clin Exp Immunol1992;90:434–9.

[30] Tomas JF, Alberca I, Tabernero MD, Cordero M, Delpıno-Montes J,Vıcente V. Natural anticoagulant proteins and antiphospholipid antibodiesin systemic lupus erythematosus. J Rheumatol 1998;25:57–62.

[31] Alarcon-Segovia D, Mestanza M, Cabiedes J, Cabral A. The antipho-spholipid/cofactor syndromes: II. A variant in patients with systemic lupuserythematosus with antibodies to β2-glycoprotein I but no antibodiesdetectable in standard antiphospholipid assays. J Rheumatol 1997; 24:1545–51.

[32] Inanc M, Donohoe S, Ravirajan CT, Radway-Bright EL, Mackıe I, MachınS, et al. Anti β2-glycoprotein I, anti prothrombin and anticardiolipinantibodies in a longitudinal study of patients with systemic lupuserythematosus and the antiphospholipid syndrome. Br J Rheumatol1998;37:1089–94.

[33] Romero FI, Amengual O, Atsumı T, Khamashta MA, Tınahones FJ,Hughres GRV. Arterial disease in lupus and secondary antiphospholipidsyndrome: association with anti β2-glycoprotein I antibodies but not withantibodies against oxidized low density lipoprotein. Br J Rheumatol1998;37:883–8.

[34] Cuadrado MJ, Lopez-Pedrera C. Antiphospholipid syndrome. Clin ExpMed 2003;3:129–39.

[35] Mc Nally T, Purdy G, Mackie IJ, Machin SJ, Isenberg DA. The use of ananti β2glycoprotein I assay for discrimination between anticardiolipinantibodies associated with infection and increased risk of thrombosis. Br JHaematol 1995;91:471–3.