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Background: The added value of antiphosphatidylserine/prothrombin antibodies (aPS/PT) in the diagnostic workup of antiphospholipid syndrome (APS) is unclear. Currently, diagnosis of thrombotic APS (TAPS) and obstetric APS (OAPS) requires persistent presence of lupus anticoagulant (LAC), anticardiolipin (aCL) immunoglobulin (Ig) G/IgM, or anti-β2-glycoprotein I (aβ2GPI) IgG/IgM antibodies., Objectives: To evaluate the role of aPS/PT IgG and IgM in OAPS., Methods: aPS/PT IgG/IgM, aCL IgG/IgM, aβ2GPI IgG/IgM, and LAC were determined in 653 patients (OAPS, TAPS, and controls). In-house aPS/PT cut-off values were calculated, titers and prevalence were compared between OAPS, TAPS, and controls and type of pregnancy morbidity. Sensitivity, specificity, likelihood ratios, and odds ratios (OR) with 95% CI were calculated., Results: In OAPS, aPS/PT IgG and IgM showed an OR of 4.32 (95% CI, 2.54-7.36) and 3.37 (95% CI, 1.93-5.89), respectively, but the association was not independent of LAC. Prevalence and titers of aPS/PT IgG and IgM were lower in OAPS than in patients with TAPS. aPS/PT were more prevalent and showed higher titers in patients with late pregnancy loss than in patients with early pregnancy loss with a positivity of 86.4% and 39.3%, respectively. Higher aPS/PT titers did not increase the likelihood of having OAPS., Conclusion: The added value of aPS/PT testing in the current diagnostic workup of OAPS seems limited compared with LAC, aCL, and aβ2GPI. aPS/PT might be useful in specific subsets of patients with OAPS. However, future multicentric studies are needed to elucidate the risk of less frequent and most severe obstetrical manifestations., Competing Interests: Declaration of competing interests G.W.M. was employed at Guy’s & St. Thomas’ Hospitals at the time of sample collection and reports consultancy fees from Technoclone, D.W. reports personal fees, outside the submitted work, from GlaxoSmithKline. The other authors state that there are no competing interests to disclose., (Copyright © 2023 International Society on Thrombosis and Haemostasis. Published by Elsevier Inc. All rights reserved.)

Background: Diagnosis of antiphospholipid syndrome (APS) requires persistent presence of lupus anticoagulant (LAC), anticardiolipin (aCL) IgG/IgM, or anti-β2 glycoprotein I (aβ2GPI) IgG/IgM antibodies. Other antiphospholipid antibodies (aPL) such as antiphosphatidylserine/prothrombin antibodies (aPS/PT) are promising in assessment of thrombotic APS (TAPS)., Aim: To evaluate the added value of aPS/PT IgG and IgM in TAPS., Material and Methods: aPS/PT IgG/IgM, aCL IgG/IgM, aβ2GPI IgG/IgM, and LAC were determined in 757 patients (TAPS and controls). aPS/PT cut-off values were calculated, and aPS/PT titers and positivity were compared between TAPS and controls, type of thrombosis, and antibody profiles. Likelihood ratios (LR), odds ratios (OR), and aPL score were determined., Results: aPS/PT IgG and IgM were associated with TAPS and triple positivity. In-house calculated cut-offs were higher for IgM (43 units), compared to manufacturer's cut-off (30 units). Thresholds of 90 (IgG) and 200 (IgM) units were determined as high-titer cut-off. Higher aPS/PT titers were observed in triple positive patients and showed higher LR and OR for TAPS. aPS/PT was independently associated with TAPS when adjusted for aCL/aβ2GPI, but not when adjusted for LAC. In isolated LAC positive patients, aPS/PT was positive in 27.1% TAPS patients and in 77.3% patients with autoimmune disease. Diagnostic value of aPL score did not differ with and without including aPS/PT., Conclusion: aPS/PT positivity, especially with high antibody titer, is associated with TAPS diagnosis. Analysis on top of current laboratory criteria is not essential in TAPS diagnosis, but aPS/PT could be useful in patients with thrombosis and a double positive aPL profile (aCL+/aβ2GPI+)., (© 2022 International Society on Thrombosis and Haemostasis.)

Background: Regional citrate anticoagulation (RCA) is recommended for continuous renal replacement therapy (CRRT). However, filter life varies and premature filter clotting can occur. The aims of this explorative prospective study were to investigate the effects of RCA on thrombin generation, fibrinolysis and platelet function in critically ill patients receiving CRRT, to compare clotting parameters between systemic and intra-circuit blood samples, and to screen participants for coagulation disorders. We recruited critically ill adult patients admitted to a 30-bedded Intensive care unit in a tertiary care hospital who required CRRT with RCA for acute kidney injury (AKI). Patients with pre-existing thrombotic, bleeding tendencies or a CRRT duration less than 48 h were excluded. We measured coagulation and thrombophilia parameters at baseline. Thrombin generation, D-dimer and platelet function were measured pre-CRRT and at 12, 24, 36, 48 and 72 h after commencing CRRT using blood samples taken from the arterial line and the circuit., Results: At baseline, all eleven patients (mean age 62.4 years, 82% male) had Factor VIII and von Willebrand Factor concentrations above reference range and significantly increased peak thrombin generation. During CRRT, there were no significant changes in systemic maximum peak thrombin generation, time to peak thrombin generation, fibrinogen, D-dimer and platelet function analysis. We observed no significant difference between paired samples taken from the patient's arterial line and the circuit., Conclusions: Critically ill patients with AKI requiring CRRT are hypercoagulable. Citrate used for anticoagulation during CRRT does not affect thrombin generation, D-dimer or platelet function. Systemic clotting parameters reflect intra-circuit results., Trial Registration: ClinicalTrials.gov Identifier: NCT02486614. Registered 01 July 2015-Registered after recruitment of first patient. https://clinicaltrials.gov/ct2/show/NCT02486614., (© 2022. The Author(s).)

Background: Antiβ2glycoprotein I (aβ2GPI) and anticardiolipin (aCL) IgG/IgM show differences in positive/negative agreement and titers between solid phase platforms. Method-specific semiquantitative categorization of titers could improve and harmonize the interpretation across platforms., Aim: To evaluate the traditional 40/80-unit thresholds used for aCL and aβ2GPI for categorization into moderate/high positivity with different analytical systems, and to compare with alternative thresholds., Material and Methods: aCL and aβ2GPI thresholds were calculated for two automated systems (chemiluminescent immunoassay [CLIA] and multiplex flow immunoassay [MFI]) by receiver operating characteristic curve analysis on 1108 patient samples, including patients with and without antiphospholipid syndrome (APS), and confirmed on a second population (n = 279). Alternatively, regression analysis on diluted standard material was applied to identify thresholds. Thresholds were compared to 40/80 threshold measured by an enzyme-linked immunosorbent assay (ELISA). Additionally, likelihood ratios (LR) were calculated., Results: Threshold levels of 40/80 units show poor agreement between ELISA and automated platforms for classification into low/moderate/high positivity, especially for aCL/aβ2GPI IgG. Agreement for semiquantitative interpretation of antiphospholipid antibodies (aPL) IgG between ELISA and CLIA/MFI improves with alternative thresholds. LR for aPL IgG increase for thrombotic and obstetric APS based on 40/80 thresholds for ELISA and adapted thresholds for the other systems, but not for IgM., Conclusion: Use of 40/80 units as medium/high thresholds is acceptable for aCL/aβ2GPI IgG ELISA, but not for CLIA and MFI. Alternative semiquantitative thresholds for non-ELISA platforms can be determined by a clinical approach or by using monoclonal antibodies. Semiquantitative reporting of aPL IgM has less impact on increasing probability for APS., (© 2021 International Society on Thrombosis and Haemostasis.)

Background: Lupus anticoagulant (LA) assays are compromised in anticoagulated patients, and existing strategies to overcome the interferences have limitations. The prothrombin-activating Taipan snake venom time (TSVT) screening test and ecarin time (ET) confirmatory test are innately insensitive to vitamin K antagonists (VKA) and direct factor Xa inhibitors (DFXaI)., Objectives: Validate standardized TSVT/ET reagents for LA detection, in a multicenter, multiplatform study., Patients/methods: Six centers from four countries analyzed samples with TSVT/ET from 81 nonanticoagulated patients with LA, patients with established antiphospholipid syndrome (APS), and proven persistent LA who were either not anticoagulated (n = 120) or were anticoagulated with VKAs (n = 180) or DFXaIs (n = 71). Additionally, 339 nonanticoagulated LA-negative patients, and 575 anticoagulated non-APS patients (172 VKA, 403 DFXaI) were tested. Anticoagulant spiking experiments were performed and 112 samples containing potential interferences (i.e., direct thrombin inhibitors) were tested. Results were evaluated against locally derived cutoffs. Imprecision was evaluated., Results: Cutoffs were remarkably similar despite use of different analyzers and donor populations. Cutoffs for TSVT ratio, ET ratio, percent correction, and normalized TSVT ratio/ET ratio ranged between 1.08 and 1.10, 1.09 and 1.12, 9.3% and 14.8%, and 1.10 and 1.15, respectively. Coefficients of variation for TSVT and ET ratios were ≤5.0%. TSVT/ET exhibited sensitivity, specificity, and negative and positive predictive values of 78.2%/95.0%/86.3%/91.5%, respectively, with established APS as the LA-positive population, and 86.9%/95.0%/76.8%/97.4%, respectively, with triple-positive APS. Interference was seen with direct thrombin inhibitors, unfractionated heparin, and low molecular weight heparins, but not VKAs or DFXaIs., Conclusions: TSVT/ET are validated for LA detection in nonanticoagulated patients and those on VKAs or DFXaIs., (© 2021 International Society on Thrombosis and Haemostasis.)

Background: Lupus anticoagulants (LA) are detected by prolongation of clotting times for dilute Russell's viper venom time (dRVVT) and activated partial thromboplastin time (APTT) screening tests. Direct oral anticoagulants (DOACs) can interfere with both screening and confirmatory tests. The present study aimed to investigate the influence of direct factor Xa inhibitors (DiXaIs) on screen, confirm and mixing tests and establish a method for differentiation from other sample types., Materials and Methods: A total of 257 samples including nonanticoagulated LA positive, LA positive with DiXaIs, factor deficiency, FVIII inhibitors, warfarin and non-APS DiXaIs were tested. APTT reagents Cephen LS/Cephen and dRVVT reagents LA1/LA2 were used, respectively, to screen/confirm the study group. Index of circulating anticoagulant (ICA) was calculated from clotting times based on the following formula as ICA screening and ICA confirmation. ICA= (1:1 Mix sample - Normal pooled plasma) / Screen patient x 100. An ICA matrix was established which suggested the presence of a DiXaI when both ICA screening and confirmation were above the cut-off. When only ICA screening is elevated, LA is suspected., Results: Sensitivity and specificity of the ICA matrix were 52.2% and 92.8% for DiXaIs and 38.1% and 96.7% for LA in APTT, and 61.2% and 92.9% for DiXaIs and 22.2% and 88.4% for LA in dRVVT, respectively., Conclusion: The ICA matrix achieved high specificity with a lower apparent sensitivity for DiXaI samples comparatively to other devices but due only to less interferences: the matrix could contribute to differentiating DiXaIs from LA in samples where anticoagulation status is unknown., (© 2021 John Wiley & Sons Ltd.)

Background:  Anticardiolipin (aCL) and anti-β2 glycoprotein I (aβ2GPI) immunoglobulin A (IgA) antiphospholipid antibodies (aPL) have shown to associate with thrombosis and pregnancy morbidity. However, inclusion of IgA aPL in the classification criteria of the antiphospholipid syndrome (APS) has been debated. We investigated the value of aCL and aβ2GPI IgA aPL in the detection of thrombosis and pregnancy morbidity in addition to the current aPL panel for APS., Methods:  We included 1,068 patients from eight European medical centers: 259 thrombotic APS patients, 122 obstetric APS patients, 204 non-APS thrombosis patients, 33 non-APS obstetric patients, 60 APS patients with unspecified clinical manifestations, 196 patients with autoimmune diseases, and 194 controls. aCL and aβ2GPI IgG/M/A were detected with four commercial assays and lupus anticoagulant was determined by the local center., Results:  Positivity for IgA aPL was found in 17 to 26% of the patients with clinical manifestations of APS and in 6 to 13% of the control population. Both aCL and aβ2GPI IgA were significantly associated with thrombosis and pregnancy morbidity. Isolated IgA positivity was rare in patients with clinical manifestations of APS (0.3-5%) and not associated with thrombosis and/or pregnancy morbidity. Addition of IgA to the current criterion panel did not increase odds ratios for thrombosis nor pregnancy morbidity., Conclusion:  aCL and aβ2GPI IgA are associated with clinical manifestations of APS. However, isolated IgA positivity was rare and not associated with thrombosis or pregnancy morbidity. These data do not support testing for aCL and aβ2GPI IgA subsequent to conventional aPL assays in identifying patients with thrombosis or pregnancy morbidity., Competing Interests: G. W. Moore reports consultancy fees from Technoclone, outside the submitted work. W. Chayoua, D. Yin, and B. de Laat are employees of Synapse Research Institute, part of Diagnostica Stago SAS. The other authors state that they have no relevant conflict of interest., (Thieme. All rights reserved.)

Background: Multicenter studies reveal that diagnostic efficacy of lupus anticoagulant (LA) assays is enhanced if cut-offs are locally generated. However, a potential confounder is the inevitable use of separate normal donor populations., Objectives: Generate cut-offs for multiple LA reagents with the same analyzer and normal donor plasmas., Methods: Cut-offs for screen ratio, confirm ratio, percent correction of screen ratio by confirm ratio, and normalized screen/confirm ratio (NSCR) were derived from the same 50 normal donor plasmas for screen and confirm pairs for two dilute Russell's viper venom time reagents, LA1/LA2 and HEMOCLOT™ LA-S/LA-C, and two APTTs, Actin FSL/FS and Cephen LS/Cephen. The cut-offs were challenged with plasmas from 20 triple-positive APS patients and 25 plasmas from LA-negative, thrombotic patients., Results: Cut-offs for screen ratio, confirm ratio, percent correction, and NSCR, respectively, were 1.12/1.08/8.3/1.09 for LA1/LA2; 1.17/1.10/13.6/1.13 for HEMOCLOT™ LA-S/LA-C; 1.12/1.13/9.7/1.10 for Actin FSL/FS; 1.09/1.13/11.0/1.11 for Cephen LS/Cephen. LA1 and LA-S screens were elevated in 19/20 and 16/20 triple-positive plasmas, respectively, while 20/20 were detected with both via integrated interpretation ie, percent correction or NSCR irrespective of screen elevation. Actin FSL and Cephen LS screens were elevated in 17/20 and 19/20 triple-positive plasmas, respectively, while one more LA was detected with Actin FSL via integrated interpretation, but not for Cephen LS. Integrated interpretation suggested 5/25 LA-negative plasmas contained weak LA (two with Actin FSL/FS, two with LA1/LA2, one with LA-S/LA-R)., Conclusions: Employing the same normal donor plasmas and analytical platform does not compensate for between-reagent differences when generating LA assay cut-offs., (© 2019 International Society on Thrombosis and Haemostasis.)

Background: Lupus anticoagulant (LA) is classified in the antibody family that is recognized in antiphospholipid syndrome. Mixing tests are recommended for LA detection, and either a mixing test-specific cutoff (MTC) or index of circulating anticoagulant (ICA) is used for the interpretation. Although we previously showed MTC had higher sensitivity for LA than ICA, there are few studies investigating specificity., Objectives: To investigate specificity of multiple activated partial thromboplastin time (APTT) and diluted Russell's viper venom time (dRVVT) reagents for inhibitors using plasmas with non-LA causes of prolonged clotting times, interpreted with MTC and ICA., Methods: Seventy-six factor-deficient samples (either artificially prepared, hereditary deficiency, or warfarin), and 12 inhibitors (either coagulation factor inhibitors, rivaroxaban, or apixaban) were used. Samples were tested with 4 APTTs, 1 dilute APTT (dAPTT), and 2 dRVVT reagents, and all elevated screen ratios were followed up with mixing tests. Frequencies of corrected and not-corrected results were calculated., Results: The frequency of MTC and ICA corrected results, suggesting factor deficiency, were 5% to 43% and 79% to 100%, respectively, except for dAPTT, where MTC and ICA performed similarly. Frequencies of MTC and ICA not-corrected results, suggesting inhibition, were 29% to 100% and 25% to 67%, respectively., Conclusions: The data indicate that MTC has a tendency to generate not-corrected mixing tests in factor-deficient, warfarin, and other inhibitor samples, while ICA exhibited higher specificity. When we perform the mixing test and interpret the data, it is important to understand the characteristics of the indexes for maximizing the diagnostic potential of mixing test., (© 2019 The Authors. Research and Practice in Thrombosis and Haemostasis published by Wiley Periodicals, Inc on behalf of International Society on Thrombosis and Haemostasis.)

Background:  The anti-phospholipid syndrome (APS) is characterized by thrombosis and/or pregnancy morbidity with persistent presence of anti-phospholipid antibodies (aPL). Laboratory criteria include aPL detection by coagulation tests for lupus anticoagulant (LAC) or solid phase assays measuring anti-β2 glycoprotein I (aβ2GPI) or anti-cardiolipin (aCL) immunoglobulin (Ig) G/IgM antibodies. External quality control programs illustrate that commercially available aPL assays produce variable results., Objective:  We aimed to investigate the agreement and diagnostic accuracy of solid phase assays., Materials and Methods:  In this multi-centre study, 1,168 patient samples were tested on one site for aCL and aβ2GPI IgG/IgM antibodies by four solid phase test systems. Samples included APS patients, controls and monoclonal antibodies (MoAB) against different epitopes of β2GPI. LAC was determined by the local centre., Results:  aCL IgM assays resulted in the most discrepancies (60%), while aCL IgG and aβ2GPI IgM assays resulted in lower discrepancies (36%), suggesting better agreement. Discrepant samples displayed lower median aPL titers. Dependent on the solid phase test system, odds ratios (ORs) for thrombosis and pregnancy morbidity ranged from 1.98 to 2.56 and 3.42 to 4.78, respectively. Three platforms showed lower sensitivity for MoAB directed against the glycine (Gly) 40-arginine (Arg) 43 epitope of domain I of β2GPI., Conclusion:  Poor agreement was observed between different commercially available aCL and aβ2GPI IgG/IgM assays, hampering uniformity in the identification of aPL-positive patients. Clinical association was globally concordant between solid phase test systems considering results of the four aPL together. An assay sensitive in detecting the MoAB against Gly40-Arg43 of domain I of β2GPI reached the highest OR for thrombosis., Competing Interests: None declared., (Georg Thieme Verlag KG Stuttgart · New York.)

Introduction: A prolonged activated partial thromboplastin time (APTT) may be indicative of a specific or multiple factor deficiency, therapeutic anticoagulation, presence of a nonspecific factor inhibitor, or lupus anticoagulant (LA). Recently, pairing of the LA-sensitive APTT and standard APTT reagents, Cephen LS and Cephen, respectively, has been shown to be effective in LA detection. The present study aimed to evaluate the usefulness of this reagent pair for discriminating between causes of APTT elevation and the detection of LA in conjunction with dilute Russell's viper venom time (dRVVT)., Methods: Plasma samples from 50 normal and 105 non-anticoagulated LA-positive patients in routine dRVVT and/or dilute APTT (dAPTT) via the percent correction formula were employed. Cephen LS/Cephen and dRVVT reagents LA1/LA2 were used to screen/confirm, respectively. Thirty-four symptomatic LA-negative, 25 warfarinised non-antiphospholipid syndrome, 6 coagulation inhibitors, 17 samples with hereditary elevated APTT, and 24 FVIII/IX/XI/XII and 17 FII/V/X artificial single deficiency plasmas were used., Results: Thirty-three samples out of 105 (31%) were LA-positive in Cephen LS/Cephen. The total percent positivity in Cephen LS/Cephen and LA1/LA2 pairs was 89.1% against samples with the routine dRVVT/dAPTT double positive. The percent corrections of Cephen LS/Cephen in the routine dAPTT/dRVVT positive group were significantly higher than those in all other groups., Conclusions: The percent correction of the APTT reagent pair showed higher values in LA-positive samples. The combination will be useful with respect to differentiating LA from other abnormal samples and is effective in LA detection when paired with dRVVT., (© 2018 John Wiley & Sons Ltd.)

Background Dilute Russell's viper venom time (dRVVT) is indispensible in lupus anticoagulant (LA) detection yet commercial reagents from different suppliers perform variably, no gold standard assays exist and therapeutic anticoagulation interference is problematic. Objective The objective of this study was to compare a new formulation dRVVT with two currently available dRVVTs. Materials and methods Life Diagnostics (LD) dRVVT and Stago PTT-LA were routinely used for lupus anticoagulant detection, plus Taipan snake venom time/ecarin time (TSVT/ET) for patients on warfarin or rivaroxaban. Siemens dRVVT and the new HYPHEN BioMed (HBM) dRVVT were tested with 193 patient samples. Group 1, 59 non-anticoagulated patients (NAPs) LA-positive in LD dRVVT; Group 2, 15 PTT-LA-positive/dRVVT-negative NAPs; Group 3, 24 LA-positive warfarinized patients; Group 4, 13 patients on rivaroxaban; Group 5, 62 LA-negative thrombotic NAPs; Group 6, 20 warfarinized, non-antiphospholipid syndrome patients. Results Accepting that the Life Diagnostics reagents were acting as a pseudo-gold standard, Siemens dRVVT detected 56/59, (95%) Group 1 LA and HBM dRVVT 46/59, (76%), one each from Group 2, and Siemens dRVVT detected one in Group 5. The lower HBM dRVVT detection rate mainly concerned weaker LA, where between-reagent concordance is problematic. All Group 3 patients appeared LA-positive in undiluted plasma with Siemens dRVVT, as did 16/24 (67%) with HBM dRVVT but the fewer LA-positives in mixing tests better mapped to clear LA-positives with LD dRVVT. LD and Siemens dRVVTs exhibited 87% and 95% false-positivity for Group 6 whilst HBM dRVVT had none. Increasing the cut-off improved accuracy. Applying higher cut-offs improved accuracy in Group 4 patients. Conclusion HBM dRVVT exhibited improved specificity, mainly due to less interference by anticoagulation, but reduced sensitivity, compared to the other dRVVTs employed.

Background: Lupus anticoagulant (LA) is classified in the antibody family that is recognized as antiphospholipid antibodies. Guidelines for LA detection recommend mixing test interpretation with either a mixing test specific cut-off (MTC) or index of circulating anticoagulant (ICA). We previously evidenced that MTC was superior to ICA in detecting the in vitro inhibition of LA with a single dilute APTT (activated partial thromboplastin time) and dRVVT (diluted Russell's viper venom time) pairing., Objectives: The objective in the present study was to compare the LA diagnostic effectiveness of MTC and ICA by multiple APTT and dRVVT reagents., Methods: One hundred-five samples from non-anticoagulated patients positive for LA in the dilute APTT (dAPTT) and dRVVT reagent pairing employed for diagnostic examination were performed by undiluted and in a 1:1 mix with normal pooled plasma with four additional APTT reagents and another dRVVT reagent (dRVVT B)., Results: Frequencies of MTC and ICA positivity were determined from samples LA positive in undiluted plasma. MTC positivity in mixing test were 63%, 77%, 80%, 84%, 46%, 81%, and 72% in 4 APTT, dAPTT and 2 dRVVT, respectively. ICA positivity were 47%, 67%, 58%, 54%, 42%, 47%, and 29%, respectively. There were no samples of ICA-positive/MTC-negative with any reagent., Conclusions: The data indicate that MTC is superior to ICA for LA detection in mixing tests in multiple reagents and reagent types. Although mixing tests may make weak LA samples appear negative, the efficacy of LA detection can be improved by the method to interpret the results.

Background: Gold standard lupus anticoagulant (LA) assays and reference plasmas do not exist and detection is based on inference in a medley of coagulation assays, creating potential for interpretive discrepancies when applying different algorithms., Objectives: To investigate discrepancies from applying different algorithms to a common data set., Methods: Diagnostic data on 311 non-anticoagulated patients LA-positive by dilute Russell's viper venom time (dRVVT) and/or dilute activated partial thromboplastin time (dAPTT) assays were employed to compare algorithms. Routine testing applied interpretive criteria from guidelines endorsing classification as LA-positive despite negative mixing tests, after exclusion of other clotting abnormalities. Integrated testing without mixing tests, and the classical algorithm where negative mixing tests preclude confirm tests, were then retrospectively applied to those data., Results: Initial testing showed 92/311 (29.6%) were LA-positive by dRVVT only, 156/311 (50.1%) by dAPTT only, and 63/311 (20.3%) by both assays. All dAPTT-positive plasmas remained positive with integrated testing but eight dRVVT-positives became negative. Other data suggested they were false-negatives. The classical algorithm altered 52/155 (33.5%) dRVVT and 111/219 (50.7%) dAPTT interpretations to LA-negative because of normal mixing tests, most of which were apparently weak LA in undiluted plasma., Conclusions: The classical algorithm improves diagnostic specificity and confidence but risks missing some genuine LA due to false-negative mixing tests. Integrated testing can be diagnostically accurate and logistically efficient but oversimplifies complex cases. Performing mix and confirm in response to an elevated screen with their interpretation based on clinical data, coagulation screens and the LA-assay design offers a potentially valuable option.

Background: Autologous platelet-rich plasma (PRP) is gaining increasing use as a wound healing promoter in a variety of clinical settings, including dentistry. Fresh PRP is often used, necessitating daily draws. The present study investigates the possibility of using stored PRP without having to freeze it by storing PRP under variable conditions and assessing growth factor release as a surrogate marker of continued viability., Methods: Freshly drawn PRP was stored in oxygen permeable and non-oxygen permeable containers under conditions of constant agitation with or without added prostaglandin, intermittent agitation and no agitation, over an 8-day period. Serial platelet counts, mean platelet volume, platelet distribution width and platelet-large cell ratio, and collagen-induced aggregometry were undertaken. Once collagen-induced aggregation had gone to completion, the plasma was centrifuged to pellet platelet material and the supernatants separated and frozen for batched analysis of released platelet-derived growth factor-BB (PDGF-BB)., Results: As would be anticipated, platelet counts, percentage aggregation and PDGF-BB levels all reduced over time. Platelet parameters suggested that platelets were more stable in the non-oxygen permeable containers, possibly due to pH drift and a degree of microaggregate formation in the oxygen permeable containers., Conclusion: Although platelet integrity and PDGF-BB fell over time, the intermittently agitated non-oxygen permeable container appeared to retain better platelet integrity and function, and PDGF-BB release, than other storage conditions, with potential for clinical use for 5-8 days.

Antiphospholipid syndrome is an autoimmune, acquired thrombophilia diagnosed when vascular thrombosis or pregnancy morbidity are accompanied by persistent antiphospholipid antibodies. Lupus anticoagulants (LA) are one of the criteria antibodies but calibration plasmas are unavailable and they are detected by inference based on antibody behaviour in a medley of coagulation-based assays. Elevated screening tests suggest the presence of a LA, which is confirmed with mixing tests to evidence inhibition and confirmatory tests to demonstrate phospholipid-dependence. At least two screening tests of different principle must be used to account for antibody heterogeneity and controversy exists on whether assays, in addition to dilute Russell's viper venom time and activated partial thromboplastin time, should be employed. A variety of approaches to raw data manipulation and interpretation attract debate, as does inclusion or exclusion of mixing studies in circumstances where the presence of a LA is already evident from other results. Therapeutic anticoagulation compromises coagulation-based assays but careful data interpretation and use of alternative reagents can detect or exclude LA in specific circumstances, and this aspect of LA detection continues to evolve. This review focuses on the main areas of debate in LA detection.

Introduction: Recent guidelines for lupus anticoagulant (LA) detection recommend mixing test interpretation with either a mixing test-specific cut-off (MTC) or index of circulating anticoagulant (ICA). Few studies directly compare efficacy of these approaches. We retrospectively applied MTC and ICA assessment to raw data of 350 LA-positive plasmas from non-anticoagulated patients to compare detection rates of inhibition., Materials and Methods: Screen and confirm dRVVT and dilute APTT assays were performed on undiluted plasma and 1:1 mixtures with normal pooled plasma. Samples were considered LA-positive if one or both screening test ratios were elevated and corrected by ≥10% with the confirmatory test. Mixing tests were assessed against locally derived cut-offs for MTC (dRVVT >1.13, dAPTT >1.15) and ICA (dRVVT >11.9%, dAPTT >13.2%)., Results: 105 of 350 (30%) were positive in dRVVT and dAPTT, 109/350 (31.1%) were dRVVT positive only and 136/350 were dAPTT positive only (38.9%), from undiluted plasma results. Of the 214 dRVVT positive plasmas, 53 (24.8%) were negative for inhibition by MTC and 65 (30.4%) negative by ICA. Of the 241 dAPTT positive plasmas, 48 (19.2%) were negative by MTC and 97 (40.2%) negative by ICA., Conclusion: Whilst integrated testing often detects LA without mixing tests they are diagnostically useful in certain circumstances. Thus, it is valuable to maximise mixing test interpretation as the dilution can lead to false-negative results. These data on a large cohort of LA-positive plasmas reveal that, with the reagents and equipment employed, MTC is superior to ICA in detecting the in vitro inhibition of LA., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Introduction: Activated protein C resistance (APC-R) due to FV Leiden is the most common hereditary thrombophilia. Rarer FV mutations can also confer APC-R, and acquired APC-R is encountered in a number of conditions. APC-R screening with clotting tests is common, yet they are prone to interferences and elevated baseline clotting times can invalidate testing., Methods: APTT-based classic APC-R (CAPC-R) screening, and modified screening (MAPC-R) employing dilution in FV-deficient plasma were performed on an automated analyser. Baseline clotting times and APC-R ratios of 1340 patients being screened for hereditary and acquired thrombophilia were assessed for analytical and diagnostic validity., Results: Most patients (1117/1340) had normal baseline clotting times, and in 270 of these cases, this was despite the presence of a lupus anticoagulant (LA). FV Leiden was genetically confirmed in all patients with reduced CAPC-R and MAPC-R ratios. A subgroup with normal CAPC-R but reduced MAPC-R also identified FV Leiden, but also other patients with minimally reduced MAPC-R ratios not due to FV Leiden. Reduced CAPC-R and normal modified APC-R identified possible acquired APC-R in 49 patients. LA-positive patients with elevated baseline clotting times did not affect distinction between APC-R and normality, although therapeutic anticoagulation did invalidate CAPC-R, and occasionally MAPC-R too., Conclusions: Many departments only screen with MAPC-R to detect just FV mutations. Concurrent performance of CAPC-R and MAPC-R increases diagnostic capability by detecting acquired APC-R. Elevated baseline clotting times can invalidate APC-R ratios, although prolongation by LA alone may not., (© 2015 John Wiley & Sons Ltd.)