Serotonin Release Assay Testing Research Articles

3Ts Score, an Attempt for Less Testing in Thrombocytopenia Patients with Suspected Heparin Induced Thrombocytopenia (HIT): Retrospective Analysis in a University Hospital Setting

Background: Heparin-induced thrombocytopenia (HIT) is a prothrombotic and potentially fatal adverse drug reaction mediated by platelet-activating antibodies against multimolecular complexes of platelet factor 4 (PF4) and heparin. The 4Ts is a pretest scoring system for HIT that was developed to improve and standardize clinical diagnosis. The strength of the 4Ts score lies in the negative predictive value of a low pretest probability score. Those with a 4T score ≥4 is considered to have moderate to high probability of HIT and should undergo confirmatory laboratory test. Delays in treatment are associated with risk of thrombosis. Immediate cessation of heparin and initiation of a non-heparin anticoagulant is therefore standard of care. Method: This is a single institutional retrospective study. We reviewed the charts of 219 patients with thrombocytopenia with suspected HIT to evaluate the appropriateness of ordering the HIT testing based on the pretest probability of the 4Ts scoring system. All patients had enzyme linked immunosorbent assay (ELISA) test for HIT antibodies (the screening test). We studied tests that were sent from 03\16\2021 through 07\03\2023. The serotonin release assay (SRA) which is the confirmatory test for HIT was only sent for patients who had a positive ELISA test. 4T scores were calculated retrospectively using chart's information available to clinicians at the time of the HIT tests were sent. Results: Out of the 219 patients, 80(36.5%), 120(54.7%) and 19(8.6%) patients had a low, intermediate and high pretest probability respectively. The HIT ELISA test was positive in 33 (15.7%) patients and 196 (84.9%) patients had negative results. HIT ELISA had 100% and 93.5% sensitivity and specificity respectively while the positive and negative predicative values were 60% and 100% respectively. A positive SRA test was reported in 23 patients (10.5% of the total study population). Seventy-nine patients (36% of our total sample) scored 4 points which represents intermediate pretest probability. Out of these 79 patients only 6 (11.3%) patients had a positive ELISA test, and 5 (6.3%) patients had a positive SRA test. Aiming to minimize the HIT over testing we examined the increase in the cutoff point of the intermediate pretest probability from 4 points to 5 points which resulted in significant increase in the sensitivity and positive predictive value of the intermediate pretest probability (75.4% and 21% versus 39% and 12.9% respectively). However, the sensitivity dropped from 90% to 65%. The 4th part of 4Ts score gives points based on the probability of HIT versus other pathology as a cause of the thrombocytopenia. Most of the patients with suspected HIT will score one point out of the 4th part of 4Ts score which makes this part of the score is less helpful in distinguishing between patients with suspected HIT and will result in higher overall pretest probability and more testing. We studied the 3Ts score (the 4Ts scoring system after elimination of the 4th part) effect on the intermediate pretest probability. The cutoff points for the 3Ts score are 1-3, 4-5 and 6 for low, intermediate and high pretest probability respectively. The 3Ts score model resulted in significant increase in the specificity and the positive predictive value (76% and 21.7% versus 39.2% and 12.9% respectively) however the sensitivity dropped from 90% to 65%. The average turnaround time for HIT ELISA test and the SRA test was 2.3 days and 4.4 days respectively. Fifteen patients (21% out the patient had low pretest probability on the 4Ts score), 41 patients (59% of the patient who had an intermediate pretest probability on the 4Ts score) and 13 patients (18.8% of the patients who had high pretest probability on the 4Ts score) started on non-heparinoid anticoagulation meanwhile awaiting on the HIT testing result. Conclusion: Over testing of HIT is common secondary to underuse of 4Ts scoring system. Multicomponent educational interventions to increase the awareness and utility of the 4Ts score in addition to implantation of clinical decision support based on HIT computerized score will help to reduce unnecessary diagnostic testing of HIT in patients with low pretest probability. Increasing the cutoff point of the intermediate pretest probability or applying the 3Ts score model resulted in significant increase in the specificity and the positive predictive value of the 4Ts scoring system on expense of the scoring system sensitivity.

Performance evaluation of heparin-induced platelet aggregation vs serotonin release assay.

Heparin-induced thrombocytopenia (HIT) is a rare but life-threatening condition that requires rapid diagnosis for proper management. Laboratory testing should only be performed on patients with intermediate- or high-risk pretest probability. The platelet factor 4 (PF4) enzyme-linked immunosorbent assay (ELISA) is the screening test that should be confirmed by higher specificity testing such as the heparin-induced platelet aggregation (HIPA) or the serotonin release assay (SRA). This study aims to evaluate the performance of the HIPA in comparison to the SRA, establish cutoffs of the PF4 ELISA to predict positivity for HIPA/SRA, and study the mortality rate between patients with suspected HIT confirmed as HIT positive vs negative. All positive PF4 ELISA cases with confirmatory HIPA and SRA testing were included. As the SRA was considered the gold standard, the HIPA performance was evaluated in comparison to SRA before and after the implementation of a new standardized interpretation guide in 2022. The mortality of these cases was also documented by chart reviews. In total, 232 cases with positive or indeterminate anti-PF4 IgG ELISA had confirmatory testing with HIPA and SRA. The sensitivity of the HIPA improved from 55.4% in 2018 to 2021 to 73.8% in 2022. The specificity remained similarly high in 2018 to 2021 vs 2022 (94.9% vs 87.5%). The negative predictive value (NPV) improved in 2022. The PF4 optical density cutoff to predict the positivity of SRA was 0.85 vs 1.47 to predict the positivity of HIPA but decreased to 0.83 when combining HIPA and/or SRA. There was no significant difference in mortality between patients with suspected HIT confirmed positive vs negative. Although the HIPA has a lower sensitivity than the SRA, the new standardized interpretation guide improved its sensitivity and NPV in 2022. Future improvements are needed to use the HIPA as a stand-alone confirmatory test with the goal to shorten hospital length of stay and expedite proper anticoagulation management.

Indeterminate serotonin release assays are associated with a high mortality rate

BackgroundThe serotonin release assay (SRA) is considered the gold standard for diagnosis of heparin‐induced thrombocytopenia (HIT). Although the SRA holds high sensitivity and specificity when results are definitive, up to 10% of samples from patients with suspected HIT yield “indeterminate” results. ObjectivesWe aimed to study the clinical course of patients with indeterminate results. MethodsWe conducted a cohort analysis of 2056 patients that underwent SRA testing. ResultsOf 2056 total patients, 152 (7.4%) had indeterminate assays. The prevalence of thrombocytopenia <50,000 × 106 was higher in patients with an indeterminate or positive SRA, compared with a negative SRA (39.5% and 40.0% vs. 27.5%, p < 4.0 × 10–4). Patients with an indeterminate SRA were more likely to have been treated in the intensive care unit than patients with a positive SRA (93.3% vs. 73.7%, p = 0.03). The mean thrombocytopenia, timing of platelet count fall, thrombosis or other sequelae, and other causes for thrombocytopenia score in patients with indeterminate SRA was 2.9, corresponding to a HIT probability of <5%. Of 152 patients, 128 (78.9%) had heparin‐PF4 optical densities (ODs) below 0.60 OD, whereas four patients (2.6%) had ODs above 2.00 OD. Inpatient mortality was significant in patients with indeterminate SRAs compared with positive or negative SRA (49.3% vs. 21.1% and 27.2%, p < 2.4 × 10–10). ConclusionsOur data suggest that an indeterminate SRA may signal an in vivo platelet activation process that is not related to heparin but is associated with increased mortality.

An institutional report of heparin induced thrombocytopenia type II in aneurysmal subarachnoid hemorrhage patients.

Heparin induced thrombocytopenia Type II (HIT-II) is a dangerous thromboembolic complication of heparin therapy. The current literature on incidence and outcomes of HIT-II in aneurysmal subarachnoid hemorrhage (aSAH) patients remains sparse. We report our institution's incidence and outcomes of HIT-II in aSAH patients. We performed a retrospective cohort study at an academic medical center between June 2014 and July 2018. All patients had aSAH confirmed by digital subtraction angiography. Diagnosis of HIT-II was determined by positive results on both heparin PF4-platelet antibody ELISA (anti-PF4) and serotonin release assay (SRA). 204 patients met inclusion criteria. Seven patients (7/204, 3.5%) underwent laboratory testing, three of whom met clinical criteria. HIT-II incidence was confirmed in two of these seven patients (2/204, 0.98%), who had high BMI and T4 scores. Our institution's report of HIT-II incidence in aSAH patients is lower than previously reported in this population and more closely parallels HIT-II incidence in the general and surgical ICU setting. Widely-accepted American College of Chest Physicians (ACCP) clinical diagnostic criteria in conjunction with anti-PF4 and SRA testing is the gold standard of clinical diagnosis of HIT-II in aSAH patients.

Heparin‐induced thrombocytopenia in end‐stage renal disease: Reliability of the PF4‐heparin ELISA

BackgroundDiagnosing heparin‐induced thrombocytopenia (HIT) in patients with end‐stage renal disease (ESRD) can be difficult, as they are frequently exposed to heparin and have multiple etiologies for thrombocytopenia. ObjectiveTo correlate 4T scores, IgG heparin–platelet factor 4 (PF4‐heparin) ELISA results, and serotonin release assay (SRA) results in patients with ESRD. MethodsWe performed a retrospective review of patients with ESRD (creatinine clearance < 15 mL/min or on renal replacement therapy [RRT]) who underwent PF4‐heparin ELISA testing from October 2015 to September 2019. True‐positive PF4s required an intermediate to high 4T score (≥4), a positive SRA, and receipt of treatment for a HIT diagnosis. False‐positive PF4s were defined as a positive PF4 with a negative SRA, low 4T score (<4), or lack of treatment for HIT. Indeterminant cases were classified on the basis of clinical assessment by the treating team (eg, hematology or vascular medicine). ResultsOf 254 patients with ESRD (92% on RRT), 29 patients (11.4%) had a positive PF4. Eleven (37.9%) had a confirmed diagnosis of HIT: 10 patients who met all of the above criteria, and one who met the 4T criteria and was treated for HIT but did not have SRA testing due to high clinical suspicion and a positive PF4 test. False‐positive PF4 values occurred in 8 patients (27.5%). Of 10 (34.5%) indeterminant cases of patients with a negative SRA but intermediate to high 4T and positive PF4, only 3 patients were treated for HIT, whereas the other 7 were judged not to have HIT as assessed by the treating clinician. In patients with an intermediate to high 4T score and PF4 optical density > 0.4 but negative SRA, who were not treated for HIT, there were no adverse outcomes documented such as new or progressive thrombosis. ConclusionIn our ESRD population, 4T scores and PF4 testing were not predictive of a clinical diagnosis of HIT.

Recognizing the Existence of Heparin Induced Thrombocytopenia and Thrombosis in the Setting of High-Dose Melphalan and Autologous Stem Cell Transplantation: A Single Institution Case Series

Background: Heparin induced thrombocytopenia and thrombosis (HIT/HITT) is a disorder characterized by transient hypercoagulability, with increased risk of venous or arterial thrombosis, related to the development of antibodies directed against the heparin-PF4 complex. HIT can occur in any patient receiving heparin but has rarely been described in the setting of high dose melphalan and autologous stem cell transplantation (HDM/SCT) so the true incidence in this population is unknown. Clinical suspicion of HIT in this setting is often clouded by the increased frequency of thrombocytopenia due to stem cell collection or high dose chemotherapy, as well as the increased risk of thrombosis in hematologic malignancies. We report five such cases, with clinically apparent and laboratory confirmed HIT that were diagnosed while undergoing treatment with high dose melphalan and autologous stem cell transplantation.Results: Five patients pursuing treatment with HDM/SCT were diagnosed with HIT during a five year period at our institution. Four patients were being treated for AL amyloidosis and one for multiple myeloma. The average patient age was 64 years (range, 59 to 69), including four males and 1 female. At the time of HIT diagnosis one patient was receiving prophylactic doses of enoxaparin due to a history of line-associated deep vein thrombosis. The other four patients had only been exposed to heparin as part of standard practice for central line placement, line flushes, or stem cell collection. Suspicion of HIT arose in all cases when the patient had delayed platelet recovery during the days following stem cell collection. Only one patient had proceeded with melphalan treatment before HIT was diagnosed. The average platelet nadir was 29, with a range of 6 to 68 x 109/L (Table 1). In two patients the development of thrombosis prompted the work-up for HIT: one with an internal jugular thrombosis and the other with limb threatening aortoiliac thrombosis requiring aortoiliac and femoral artery embolectomy with fasciotomy. All five patients had confirmation of diagnosis with heparin-PF4 antibody (optical density range, 0.5 to 3.3) and a positive serotonin release assay (SRA). Four patients were started on anticoagulation with fondaparinux or argatroban with appropriate platelet recovery in all patients. Three patients proceeded with HDM/SCT, while the remaining two patients received alternative therapies because the previously collected stem cells had been processed with heparin and could not be given to the patients.Conclusions: Herein we have describe five cases of HIT confirmed by the presence of a heparin-PF4 antibody and SRA testing. In all five cases the cause of thrombocytopenia was initially thought to be related to stem cell collection, which is known to cause a decline in platelets. It is notable that the platelet nadir in these patients, was frequently much lower than the nadir that is typically expected in HIT (50 to 80 x 109/L). Despite the development of thrombosis in two patients and the known increase in mortality in patients with HIT, the long term outcome in each of these patients was favorable. This is potentially attributable to rapid diagnosis, cessation of heparin, and initiation of alternative anticoagulation. In reaction to the initial cases, our institution has changed practices to eliminate heparin in the processing of stem cells so that all patients are able to receive their collected stem cells even in the setting of HIT. These cases highlight the existence of HIT during the process of HDM/SCT and the need for increased diligence in monitoring for unexpected thrombocytopenia or sluggish platelet recovery, in particular immediately after stem cell collection. Clinicians should have a low threshold to send diagnostic testing, begin alternative anticoagulation and postpone high dose chemotherapy in order to prevent life threatening complications. [Display omitted] DisclosuresSloan:Molecular Templates, Inc.: Consultancy; Stemline Pharmaceuticals: Consultancy.

HIT Diagnosis: A Proposed New Definition for Clinical Research

Introduction: Heparin-induced thrombocytopenia (HIT) is a rarely diagnosed but often suspected complication of heparin therapy. Initial diagnosis is generally made based upon a combination of pretest probability (such as the 4Ts score) and the results of readily available immunoassays. However, confirmation requires the result of a functional assay such as the serotonin release assay (SRA), which is currently considered to be the gold standard. Because the SRA is technically challenging, it is offered only in a few reference laboratories, resulting in a turnaround time of several days to a week. In addition, recent data suggest that the accuracy of the SRA may not be as high as previously thought. Thus, new laboratory assays that are both highly sensitive and specific for HIT antibodies are urgently needed. Here we propose a new set of criteria, independent of the SRA, that may be used to define HIT in the evaluation of novel assays.Methods: We conducted a retrospective cohort study of adult inpatients undergoing laboratory testing for suspicion of acute HIT at the University of Washington Medical center and/or Harborview Medical Center between May 2016 and July 2017. All patients underwent enzyme-linked immunosorbent assay (ELISA) testing for antiPF4/heparin antibodies (IgG-specific Zymutest EIA, Hyphen Biomed). According to institutional policy, those samples with optical density (OD) ≥ 0.300 were reflexively sent to a reference laboratory for SRA testing. Charts were examined by trained reviewers for calculation of the 4Ts score, results of ELISA and SRA testing (when available) and outcomes at 30 days including clinical diagnosis/management of HIT and mortality.Results: We identified 155 patients undergoing evaluation for HIT. Each patient was placed into one of three categories: ‘negative’, ‘indeterminate’, or ‘positive’, based upon a combination of 4Ts score and the optical density (OD) resulting from ELISA testing. Any patient with an OD ≤0.300 was considered to be ‘negative’ regardless of the 4Ts score. Patients with ODs ranging from 0.301-0.999 were defined as ‘negative’ unless the 4Ts score was high (6-8), in which case they were defined as ‘indeterminate’. Patients with an OD of 1.000-1.999 were defined as ‘positive’ if the 4Ts was high (6-8), indeterminate if the 4Ts was intermediate (4-5) and negative if the 4Ts was low (0-3). Patients with an OD ≥2.000 were defined as ‘positive’ unless the 4Ts score was low (0-3) in which case they were defined as ‘indeterminate’. The number of patients that fell into each category as well as the number of subjects with a clinically confirmed diagnosis of HIT are shown in Table 1.The median age of all subjects was 60 and was this was roughly similar across ‘negative’, ‘indeterminate’, and ‘positive’ categories (60, 62, and 66, respectively). Males represented 62% of all subjects and 71% of patients categorized as HIT positive. Seventy percent of all subjects received heparin for venous thromboembolism prophylaxis. Eighty-nine percent of ‘negative’ and 100% of ‘indeterminate’ subjects were exposed to unfractionated heparin (UFH). Six of 7 ‘positive’ patients were exposed to UFH and 1 to low molecular weight heparin (LMWH). All ‘positive’ patients were clinically managed for HIT by stopping all heparin products and/or beginning patients on direct thrombin inhibitor therapy, with 57% mortality within 30 days of ELISA testing. One (0.8%) ‘negative’ and 7 (32%) ‘indeterminate’ patients were clinically managed for HIT. Thirty-day mortality was 25% and 4.5% in ‘negative’ and ‘indeterminate’ groups respectively.Discussion: Our study proposes a framework for the use of clinical pretest probability scores and rapidly available HIT ELISA testing to define HIT in research studies. Such a strategy allows for assessment of accuracy of newly developed assays. The inclusion of an ‘indeterminate’ category adds statistical complexity but is necessary to avoid misclassification of patients who may or may not have true HIT. Mortality in HIT positive patients defined in this manner was high, although small numbers limit definitive conclusions and unfractionated heparin was implicated in most cases of HIT, as has been observed previously. [Display omitted] DisclosuresPadmanabhan:BloodCenter of Wisconsin: Patents & Royalties: A patent application has been filed related to the PF4-dependent P-selectin expression assay (Method of Detecting Platelet- Activating Antibodies That Cause Heparin-Induced Thrombocytopenia/Thrombosis; PCT/US14/62591).; Retham Technologies LLC: Equity Ownership.

Impact of Systems-Based Hematologists in Maintaining a HIT Program Using Continuous Clinical Audit and Education in an Academic Medical Center

Systems-based hematologists (SBH) play a vital role in navigating the health care system in the use of diagnostic tests that crosses multiple disciplines by implementing evidence-based strategies to improve patient safety and outcomes. In 2013, we demonstrated that an Anticoagulation and Bleeding Management Service (ABS) led by SBH played a major role in developing safe and cost-effective approaches for the diagnosis of patients with heparin-induced thrombocytopenia (HIT) via implementation of a centralized hospital-wide physician paper order protocol (Vaughn et al. Blood. Vol. 124, Abstract 4848). Platelet factor 4 (PF4) ELISA testing decreased from 600 in 2011 to 375 in 2013 (37.5% decrease), and serotonin release assay (SRA) testing decreased from 202 in 2011 to 29 in 2013 (85% decrease). In addition, 100% of patients with positive PF4 ELISA received an automatic consult by the ABS team to determine management of patients with suspected HIT. This led to a reduction in the use of direct thrombin inhibitors (DTI) by 78%. Despite this initial success, we were unsure if these improvements were sustainable moving forward.The purpose of this study was to provide quantitative data demonstrating the continuous impact that SBH have on health care delivery via annual re-auditing on PF4 ELISA testing to identify shortfalls and implement improvements as needed. In 2014, our institution switched to a new electronic medical record (EMR) system using EPIC in the inpatient environment. The 4T's Scoring Calculator physician paper order became obsolete. An electronic order set for PF4 ELISA test was in discussion but due to the complexity of switching to a new EMR system, this discussion was stalled. Now, physicians could order a PF4 ELISA test electronically without a 4T score. As a result, PF4 ELISA testing increased to 402 in 2014 and 632 in 2015. Ordering of SRA test, which was restricted to the ABS team, remained constant at 31 in 2014 and 36 in 2015.To overcome this, throughout 2015-2016, the ABS team gave specific feedback to the ordering physician when a false-positive PF4 ELISA test was ordered inappropriately. This allowed for individual discussion on each patient case to improve physicians' understanding and awareness of HIT testing and management. In addition, departmental-wide lectures on HIT/4T score to attending physicians and residents/fellows were scheduled.Despite these departmental-wide lectures, the ABS team found that ordering physicians remained unclear about calculation of the 4T score. Ordering physicians were calculating higher 4T scores and ordering PF4 ELISA based on their calculations. As a result, specific interactive lectures based on real-life case scenarios where the audience (internal medicine residents) calculated the 4T score for each scenario were given. This allowed for case-based discussions and opportunity for clarifying any uncertainty on the 4T score. Feedback from the residents was overwhelmingly positive. These efforts led to a reduction in PF4 ELISA test by 49% to 323 in 2016. A re-audit revealed that the number of PF4 ELISA test remained steady at 321 in 2017, which demonstrates the success of our interventions (Figure 1).In conclusion, this study highlights the important of re-auditing to maintain the improvements made. Despite initial success, annual re-auditing allowed for early detection of challenges, which then allowed appropriate early intervention. Administrative and financial support from the health care system for SBH allowed for the initiation, implementation and monitoring of this HIT program, and should be encouraged nationwide as a rewarding career opportunity for hematologists interested in hospital quality improvement and patient safety. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Heparin Induced Thrombocytopenia: A Laboratory Comparison of the IgG Specific EIA Vs Polyspecific EIA

Introduction:Heparin-induced Thrombocytopenia (HIT) is a potentially life/limb threatening immune mediated pro-thrombotic syndrome triggered by heparin/platelet factor 4 (PF4) complexes. Based on clinical probability, if indicated, initial laboratory testing for HIT is typically based on enzyme immunoassay (EIA) to detect anti-PF4/heparin(H) antibodies with or without supplemental functional assays, the serotonin release assay (SRA) is considered to be the gold standard. EIAs detect either polyspecific (IgG, IgA, IgM) or IgG specific antibodies, the latter are recommended by ISTH/SSC, and should have optimal performance characteristics (sensitivity/specificity/negative and positive predictive values (NPV/PPV)). Each performing laboratory, however, needs to establish in-house performance characteristics of the assay.Objective:To compare laboratory performance characteristics of IgG specific and polyspecific PF4/H EIAs for a positive SRA.Materials and methods: Between 2013 to 2017, 284 serum samples submitted to our laboratory for clinical HIT EIA testing were studied. IgG specific and polyspecific (Immucor GTI Diagnostics, Inc. Waukesha, WI) assays were performed per the manufacturer's instructions with the exception of performance on a liquid handling instrument rather than manually. The manufacturer assigned optical density (OD) cutoff of ≥0.400 was defined as positive. The SRA, ordered separately by clinicians, were sent out to reference laboratories; criteria for a positive result was ≥20% serotonin release with low dose heparin and <20% release with high dose of heparin. The data were entered into Excel spreadsheet, de-identified and analyzed by Analyse-it (Analyse-it Software, LTD, Leeds, U.K.). We also correlated the SRA positivity with the EIA ODs.Results:Of the 284 serum samples, 112 were positive with IgG specific EIA, 68 (60.7%) of those also had a positive SRA. 172 were negative with IgG specific EIA, 2 (1.2%) of those had a positive SRA. Of the 148 were positive with polyspecific EIA, 69 (46.6%) also had a positive SRA. 136 were negative with polyspecific EIA, 1 (0.7%) had a positive SRA. The performance characteristics are shown in table 1. Compared with the polyspecific EIA, the IgG specific EIA had superior specificity, PPV and positive likelihood ratio (LR); the NPVs and negative LR of both assays were similar. The percent of patients having a positive SRA at different OD cutoffs is illustrated in Table 2.Conclusion: In aggregate the data suggest that the IgG specific EIA had superior performance characteristics in detecting HIT antibodies. Given the high likelihood of a positive SRA for OD >2.0, in the appropriate clinical context, supplemental SRA testing may not be essential. Since the data were de-identified, assay performance for the clinical diagnosis of HIT was not possible. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Indeterminate Serotonin Release Assays Are Associated with a High Mortality Rate

INTRODUCTION: The serotonin release assay (SRA) is considered the gold standard for diagnosis of heparin-induced thrombocytopenia (HIT). While the SRA holds high sensitivity and specificity when results are definitive, up to 10% of samples from patients with suspected HIT yield "indeterminate" results. In classic HIT, the SRA typically reveals high levels of serotonin release when serum is mixed with low concentrations of heparin and low levels of serotonin release when serum is mixed with high concentrations of heparin. Results are considered "indeterminate" when high levels of serotonin release are seen at both low and high concentrations of heparin, indicating a failure of high-dose heparin to saturate the heparin binding sites of PF4 molecules and inhibit platelet activation. Explanations for indeterminate assays include the presence of heparin-binding proteins that interfere with the assay, high titers of HLA class I alloantibodies, or immune complexes. Since the diagnosis of HIT carries such significance and has so many ramifications, an indeterminate SRA may leave therapeutic indecision. The etiologies, platelet trends, clinical course and outcomes of patients that receive indeterminate SRA results are not well-understood. We conducted a retrospective review of 2,056 patients that underwent SRA testing as part of their evaluation for HIT. METHODS: Using the electronic medical record data extraction software Clinical Looking Glass, we identified patients that underwent SRA testing between 1/1/2014 and 12/31/2018. SRA results were considered "indeterminate" when serotonin release exceeded 19% at all heparin dilutions (0.1 U/mL, 0.5 U/mL and 10 U/mL). We conducted a retrospective chart review to study the clinical course among patients who had "indeterminate" SRA results, the trends in platelet count, the timing of platelet drops and the physician response to this result. Statistical analysis was performed using chi-square testing for categorical variables. RESULTS: We identified 2,056 patients that underwent SRA testing between 2014 and 2019. Of these, 90 patients (4.4%) had "positive" SRA results and 1,814 patients (88.2%) had "negative" SRA results. Among the 152 patients (7.4%) with "indeterminate" SRA results, the mean 4T score was just 2.9, corresponding to a HIT probability of &amp;lt;5%. One-hundred and twenty of these 152 patients (78.9%) had heparin-PF4 antibody testing with optical densities below 0.60 OD, while only 4 of 152 patients (2.6%) had optical densities above 2.00 OD. Seventy eight of 152 patients (51.3%) either continued or were re-exposed to heparin after the indeterminate SRA result, and in 71 of 78 cases (91.0%), the platelet count stabilized or improved despite heparin exposure. In the remaining 7 cases, no acute VTE were found and no diagnosis of HIT was made. Four of the 71 patients (5.6%) that continued or were re-exposed to heparin were noted to have an acute VTE. A significantly higher portion of patients with an indeterminate SRA died during admission compared to those with a positive or negative SRA (49.3% vs. 21.1% and 27.2%, p &amp;lt;2.4 x10-10). The prevalence of thrombocytopenia &amp;lt;50,000 was substantially higher in patients with an indeterminate or positive SRA, compared to those with a negative SRA (39.5% and 40.0% vs. 27.5%, p &amp;lt;4.0 x 10-4). Re-exposure or continuation of heparin did not affect mortality; patients that were given heparin after an indeterminate SRA had a 47.4% mortality, compared to 51.4% in those given no more heparin (p = 0.63). Table 1 summarizes these findings. CONCLUSIONS: This is the largest study to date looking at patients with indeterminate SRA testing results, in which platelets exhibit high levels of serotonin release at both low and high heparin concentrations. Our data suggest 1) the majority of patients with indeterminate SRAs likely did not have HIT, 2) as evidenced by the low 4T scores and heparin-PF4 antibody levels, an indeterminate SRA suggests thrombocytopenia related to in-vivo platelet activation, and 3) an indeterminate SRA is associated with increased mortality. SRA results warrant a case-by-case assessment of the clinical picture in order to avoid unnecessary cessation of heparin products. Further studies exploring the mechanism leading to high serotonin release at both low and high heparin concentrations, causing an "indeterminate" SRA result, are warranted. Disclosures No relevant conflicts of interest to declare.

AORTIC THROMBOSIS AFTER CARDIAC SURGERY FROM HEPARIN-INDUCED THROMBOCYTOPENIA

SESSION TITLE: Fellows Critical Care Posters SESSION TYPE: Fellow Case Report Posters PRESENTED ON: October 18-21, 2020 INTRODUCTION: Heparin-induced thrombocytopenia (HIT) is a complication caused by exposure to heparin. Although incidence after cardiac surgery is of 0.3%, there is a 50% increase in mortality when HIT occurs [1]. CASE PRESENTATION: A 65-year-old male with history of diabetes mellitus presented with chest pain. Two days prior to coronary artery bypass grafting (CABG), coronary angiography revealed triple vessel disease. He received 3000 units of heparin for the procedure. He underwent on-pump CABG, where 19,500 units of heparin were given. After surgery, platelet count decreased with nadir of 105×103 /μL (40% decrease) on postoperative day (POD) 4. He complained of abdominal and lower extremities’ pain on POD 5. Computed tomography (CT) showed a thrombosed infrarenal aorta (Fig 1). Vascular team performed axillary and femoral artery bypass surgery. He complained of abdominal pain and CT showed dilated loops of bowel with pneumatosis intestinalis on POD 7. Laparotomy was performed and necrosed small bowel was resected. HIT was suspected due to thromboembolic events (4Ts score was 6; high risk of HIT) [2]. Particle Immuno-Filtration Assay (PIFA®) and Serotonin Release Assay (SRA) were ordered and results came negative. Assays were done for second time on POD 10 and PIFA® resulted negative. Echocardiogram revealed small thrombus in left ventricle apex on POD 11. Although SRA result was pending, heparin was discontinued and argatroban was administered. SRA test came positive on POD 12. Platelet count began recovering and there was no more thromboembolic event. Platelet count returned to normal at 5-month follow-up (Fig 2). DISCUSSION: There are two requirements to determine presence of HIT: a) clinical diagnosis and b) detection of heparin antibodies activating platelet (through positive SRA). Clinical diagnosis can be completed by use of 4Ts score [2] (Fig 3). If 4Ts score is more than 4 (intermediate risk), the serological tests should be performed. There are two types of tests for HIT: one is the antigen assay PIFA® and the other is the functional assay SRA. Both have high sensitivity of 90%. PIFA® is used for screening, has a specificity ranging from 50% to 89% and its result can be available within 15 minutes. SRA has a higher specificity of more than 90%. To confirm the diagnosis, SRA result should be positive, although several days may take to obtain result. CONCLUSIONS: If the serological test shows negative and HIT is still suspected clinically, the test should be repeated. High clinical suspicion should lead to termination of heparin and initiation of argatroban. Reference #1: Seigerman M., Cavallaro P., Itagaki S., Chung I., Chikwe J. Incidence and outcomes of heparin-induced thrombocytopenia in patients undergoing cardiac surgery in North America: an analysis of the nationwide inpatient sample. J Cardiothorac Vasc Anesth. 2014;28:98-102. Reference #2: East M.J., Cserti-Gazdewich M.C., Granton T.J. Heparin-induced thrombocytopenia in the critically ill patient. Chest. 2018; 154:678-690. Reference #3: Ortel T.L. Heparin-induced thrombocytopenia: when a low platelet count is a mandate for anticoagulation. Hematology Am Soc Hematol Educ Program. 2009;225-232. Fig 1; Contrast computed tomography (CT) showed total occlusion of the infrarenal abdominal artery and bilateral iliac arteries. DISCLOSURES: No relevant relationships by Youdelman Benjamin, source=Admin input No relevant relationships by Shinichiro Ikeda, source=Web Response No relevant relationships by Ronald Kaleya, source=Web Response No relevant relationships by Robert Rhee, source=Web Response

Utility and Pitfalls of Stepwise Laboratory Testing for Heparin Induced Thrombocytopenia: Retrospective Review from an Academic Medical Center

Introduction: Heparin induced thrombocytopenia (HIT) occurs in up to 5% of adults exposed to heparin due to formation of heparin-dependent antibodies to the heparin/platelet factor 4 (PF4) complex. Patients develop thrombocytopenia and are at risk for severe thrombotic complications. Mortality rates can be as high as 20%, but are often much lower with prompt recognition, cessation of heparin, and treatment with alternative anticoagulants. However, these alternative medications are often both labor-intensive drips and expensive, highlighting the need for quick decision making in such challenging cases. The classic workup of HIT involves assessing clinical suspicion (often using the "4T score" on a scale of 0-8), followed by lab testing for detection of the heparin-dependent antibody and then a functional assay to confirm pathogenicity of the antibody. Our institution uses an in-house rapid Particle ImmunoFiltration Assay (PIFA), which is a same-day test and reported as "positive" or "negative." Other testing available includes send-out tests for Enzyme-Linked Immunoassay (ELISA) to IgG, A, and M of the PF4 antibody as well as the C-14 Serotonin Release Assay (SRA) functional assay. The goal of this retrospective review is to analyze the utility of an algorithmic approach to laboratory testing in aiding the rapidity of diagnosis of HIT without missing possible critical cases. Methods: This was a single institutional study at a large urban academic medical center. We reviewed inpatient charts from 2015-2018 of patients who had any lab testing for HIT. As per institutional guidelines, first-line testing is recommended by using the in-house same-day testing PIFA. If positive, a sample is automatically reflexed and sent-out for PF4 ELISA testing and SRA. Furthermore, clinicians are able to directly order ELISA and SRA testing at their medical discretion. Our analysis looked at those patients with at least two different "HIT-related" laboratory tests to best analyze the concordance and discordance rates of the above testing to assess for sensitivity, specificity, and overall accuracy of using a stepwise testing approach. We used a cutoff value of &gt;0.4 optical density (OD) for ELISA testing, and &gt;20% release at low-dose heparin concentration for SRA testing. Results: There were 118 patients who had at least two different HIT-related laboratory tests sent. 91 patients had both PIFA and ELISA testing, with 37/79 (47%) positive concordance rate and 6/12 (50%) negative concordance rate, for a sensitivity of 86% and specificity of 13%. 3 patients with positive PIFA also had positive SRA, and there were 2 patients with negative PIFA with positive SRA testing (see attached Table). When comparing ELISA testing to SRA, 4/41 (10%) had concordant positive testing, while no patient with a negative ELISA test had a positive SRA (28 concordant negative cases). Overall, of 94 SRA tests run, 5 were positive, of which 2/5 had negative PIFA and 0/4 had negative ELISA testing. Conclusions: While PIFA testing had a high sensitivity compared to ELISA, the overall accuracy compared to ELISA was low, while ELISA testing was 100% sensitive in this analysis. Furthermore, there was still a risk of missing cases of HIT using PIFA testing alone. In both cases of positive SRA with a negative PIFA, patients had a high 4T score of &gt;6, consistent with a high clinical suspicion for HIT. We conclude that PIFA testing is not equivalent to ELISA testing, and that use of a laboratory "algorithm only" approach would be inappropriate in the diagnosis of HIT. Our results highlight the importance of using both clinical scoring systems and appropriate lab testing together in the workup and diagnosis of HIT. Disclosures No relevant conflicts of interest to declare.

Effective Implementation of a Structured Protocol for Facilitation of the Judicious Use of Antibody Test for Diagnosis of Heparin Induced Thrombocytopenia

Introduction: Heparin induced thrombocytopenia (HIT) is a serious prothrombotic condition, usually triggered by exposure to heparin products with formation of antibodies to platelet factor 4/ heparin polyanion complexes. Diagnostic algorithm for HIT combines clinical scoring (4T score) with time sensitive screening for HIT antibodies (HIT-ab), while serotonin release assay (SRA) is remains the gold standard for confirmatory diagnosis. The rate of utilization of 4T score was low in our institution, resulting in inappropriate orders for HIT-Ab test and subsequent administration of unnecessary alternative anticoagulation (AC) in patients with false positive results. In this project, we designed a structured HIT diagnostic workflow incorporating 4T score calculation in our electronic medical record (EMR) and replaced particle immunofiltration assay (PIFA) with latex immunoturbidometric assay (LIA) in our laboratory for HIT-Ab screening, with an aim to improve the rate of 4T utilization and accuracy of HIT diagnosis in a cost-efficient manner. Methods: In phase I, we performed a retrospective chart review of all patients with PIFA ordered between March 2017-March 2018. Two investigators independently calculated 4T, collected data on results of HIT-Ab, confirmatory SRA tests, and the duration of alternative AC from each record. Any variations in 4T score were resolved by a senior investigator. In phase II, we implemented a new workflow in the EMR incorporating mandatory calculation of 4T score with every order for HIT-Ab test. Our lab started using LIA in place of PIFA. Charts were reviewed on patients with HIT-Ab orders (LIA) from January-June of 2019. Results: On review of data from phase I, we noted that 4T score was documented in only 5 (0.02%) of 170 patients in whom a PIFA test was ordered. Per investigators assessment, 113 (66.4%) patients had low probability (4T ≤ 3), 47 (27.6%) had intermediate probability (4T 4 or 5), and 10 (5.8%) had a high probability (4T ≥ 6) for a diagnosis of HIT. SRA was ordered in 32 patients, although 17 of them had low probability per investigator assessment. PIFA test came back positive in 26 patients, of whom 16 had corresponding SRA results, and three samples were positive for SRA. PIFA was negative in two patients with confirmed HIT (SRA positive). A total of 19 patients received alternate AC in the first phase, 7 of them had low 4T score per our assessment. In phase II, 69 records were found with available LIA results, showing a relative decrease in HIT-Ab orders compared to earlier phase at the six months mark. Documentation of 4T score has been 100% by ordering physicians, a certain improvement from phase I. Investigator calculated 4T score showed low probability in 33 (47.8%) patients, intermediate probability in 31 (44.9%) patients, high probability in 5 (0.07%) patients. LIA was positive in 7 of the 69 ordered tests, 6 of whom scored high/intermediate in the 4T score. HIT diagnosis was confirmed in 3 of these 7 patients with a positive SRA result. During this period, all the 7 of the eight patients who received alternate AC had a high or intermediate probability for HIT as per 4T. Conclusion: Our study demonstrated that the successful implementation of a structured protocol for HIT diagnosis ensured 100% adherence to the calculation and documentation of 4T score by clinicians, and significantly reduced the number of inappropriate HIT-Ab test orders in our institution. Use of alternate AC was also more consistent with the level of probability for HIT. Table Disclosures No relevant conflicts of interest to declare.

Reduction in False-Positive PF4/Heparin Antibody Testing Following Implementation of a Latex Immunoturbidimetric Assay

Abstract Background Heparin-induced thrombocytopenia (HIT) is a life-threatening complication of exposure to heparin that is caused by autoantibodies against heparin-PF4 complexes. We recently changed our in-house HIT screening platform from a manual, daily batched ELISA (Stago-Asserochrom HPIA Immunoassay) to an automated, on-demand latex immunoturbidimetric assay (LIA, HemosIL HIT-Ab) and have also implemented a reflex from a positive LIA result to the confirmatory serotonin release assay (SRA). We compared the two methods in terms of utilization, test performance, and turnaround time. Methods Data were collected retrospectively from a 7-month period before (June-December 2017) and after (June-December 2018) implementation of the HemosIL LIA in the clinical laboratory at a large academic institution. This study includes consecutive test results from adults (median age: 64 years, range: 19-98 years) seen at our 1,300-bed main hospital. Test utilization, turnaround time (sample receipt to verification), and test performance characteristics were compared between the two methods. Repeat testing was excluded from the analysis. Samples with a positive result on the HemosIL LIA were reflexed to a serotonin release assay (SRA), performed at a large reference laboratory, whereas samples tested with the earlier ELISA assay were referred for SRA testing based upon clinical judgment. When performed, SRA was considered the gold standard for diagnosis of HIT. Results During the 7 months before and after switching methods, there were 109 of 594 (18.4%) positive ELISA results and 45 of 523 (8.6%) positive LIA results. Only 90 of 109 (82%) of the positive results from the ELISA HIT Ab test were sent out by clinicians for SRA testing, whereas 45 of 45 (100%) of the positive results from LIA testing were reflexed to SRA per protocol. Although fewer LIA tests were sent out for SRA testing, there were an equal number of SRA-confirmed cases of HIT with the ELISA (PPV: 16/90 [17.8%]) and LIA methods (PPV: 16/45 [35.6%]), resulting in a high positive predictive value (PPV) with the newly implemented method. Not only was the PPV higher with the LIA test, but it had a significantly shorter mean turnaround time of 96 minutes compared to the ELISA TAT of 1,234 minutes (P &lt; .0001). Conclusions With the new testing protocol, patients received results faster (average 96-minute TAT) and had fewer false-positive results (74/594 pre vs 29/523 post), with no apparent reduction in detection of true-positive cases of HIT (16/594 pre vs 16/523 post).

Multi-Year Study of Unfractionated Heparin Serotonin Release Testing at a Large U.S. Reference Laboratory

Introduction: The serotonin release assay (SRA) is a functional assay for the detection of heparin-induced thrombocytopenia (HIT). Laboratory practice suggests that it is important to test several low doses of unfractionated heparin (UFH), followed by suppression at a high dose (100 U/mL) of UFH to demonstrate the specificity for HIT. However, little is known of the demographic associations with UFH SRA testing at a large reference laboratory.Purpose: To characterize the population characteristics of UFH SRA testing at a large US reference laboratoryMethods: We performed a multi-year retrospective review of positivity rates among adults (n=63,651) and children (<18 yrs, n=233) who underwent SRA testing; we examined positivity rates at 0.1 and/or 0.5 U/mL of UFH. A positive SRA test was defined by ≥20% release at either low dose (0.1 or 0.5 U/mL) of UFH and >50% suppression of release at high dose (100 U/mL) of UFH. Indeterminant results were defined by ≥20% release at either low dose of UFH, but without >50% suppression at the high dose of UFH. A negative result was defined as <20% release at both low doses of UFH. All testing was performed at Quest Diagnostics, Nichols Institute in Chantilly, VA. Demographic and laboratory information collected included age, gender, date of collection, and SRA results at 0.1 and 0.5 U/mL UFH. Descriptive statistics were performed as well as non-parametric statistical analyses using the Mann-Whitney U test (two-tailed) with significance defined at p <0.05.Results: Over the study period, SRA testing included 63,051 adults and 233 children who encompassed 99.5% and 0.5% of the total population tested, respectively. Over the years studied, the positivity rate increased steadily from 4.08% to 7.50%, which was seen in adults but not in children. For positive SRA results, the ratio of male to female was nearly identical to the overall male to female ratio (55:45) in the tested population. In additional, no seasonal variation was noted. Yearly median percent positive, indeterminant, and negative SRA results were 5.42%, 3.32%, and 89.73%, respectively, for adults, and 1.96%, 6.52%, and 93.48%, respectively, for children, with children having significantly lower SRA positivity than adults (p<0.037).Among positive tests in adults, we examined the results by level of low dose UFH: the median positivity rates were 3.46% for 0.1 U/mL only, 1.03%, for 0.5 U/mL only, and 95.89% for both 0.1 and 0.5 U/mL UFH. In contrast, children demonstrated positive tests only when both 0.1 and 0.5 U/mL UFH results were positive. These children (n=6, median age 14 yrs with range 3 to 15 yrs) demonstrated median serotonin release of 81% and 77% at 0.1 and 0.5 U/mL of UFH, respectively, which was not significantly different from each other and similar to the corresponding adult positive (both 0.1 and 0.5 U/mL UFH) SRA population.Conclusions: Adults have a higher prevalence of SRA testing compared to children and have median positivity rates that are significantly higher (2-3 fold).The consistent increase in yearly percentage of positive tests, suggests improved clinical screening and/or diagnostic pathways for HIT. Testing was not influenced by gender or seasonal variation. To our knowledge, these findings provide, for the first time, a detailed picture of SRA testing from a large US reference laboratory. DisclosuresWong:Quest Diagnostics: Employment, Other: stock . Sahud:Quest Diagnostics: Consultancy. Noh:Quest Diagnostics: Employment. Aboueyada:Quest Diagnostics: Employment. Frock:Quest Diagnostics: Employment. Haley:Quest Diagnostics: Employment. Wu:Quest Diagnostics: Employment. Dlott:Quest Diagnostics: Employment, Other: stock.

Confirmation of the Utility of the 4T Scoring System for HIT

Background: Heparin-induced thrombocytopenia (HIT) is a complication of heparin-based anticoagulation (AC) resulting in thrombocytopenia and thrombosis. Laboratory testing can often be avoided as the 4T score (4TS) has a negative predictive value (NPV) of 0.998 for low risk patients. Despite this scoring system, which has been validated since 2006, physicians continue to send inappropriate studies despite a low probability of HIT. We sought to evaluate our academic institution's compliance, perform a cost analysis and determine if appropriate AC was initiated. By analyzing our data, we sought to educate our staff and implement measures to improve cost efficiency and quality of care.Methods: We performed a retrospective chart review of patients admitted to Hahnemann University Hospital (HUH) between November 1, 2016 and April 30, 2017 who had HIT antibodies (HITAb) and serotonin release assay (SRA) studies. These laboratory tests were performed at Quest Diagnostics. This data was compiled from the EMR at HUH. According to the 4TS, patients were assigned a score of 0-8: 0-3 for low, 4-5 for intermediate, 6-8 for high probability respectively. Laboratory results of HITAb and SRA were then compared to the calculated 4TS. We then investigated whether appropriate AC was initiated. Data on the cost associated with the inappropriate management of suspected HIT was compiled.Results: 72 patients had HITAb sent during the interval studied. Table 1 shows the 4TS and results of HITAb and SRA testing. Table 2 lists the AC used based on the 4TS.The NPV of not having HIT in the low probability group was 100%. The positive predictive value (PPV) of having HIT in the high probability group was 100%. At our institution, HITAb with reflex SRA costs $503. Expenditure due to inappropriate testing was estimated to be around $23,000 dollars over the study's time course. Inappropriately switching to argatroban cost up to $1,000 per day or fondaparinux $500 per day of overspending on anticoagulation per patient.Discussion: We found the majority of HITAb and SRA testing was unnecessary based on the 4TS. Our data showed a low 4TS had a very high NPV confirming the scoring system's utility. HIT testing was often overutilized as part of a general workup for thrombocytopenic patients who were often septic, on marrow suppressive medications and had multiple comorbidities such as hepatitis and HIV infections which confounded their clinical picture. Furthermore, this scoring system had a very high PPV in the high probability group. This study confirmed that HIT laboratory studies rarely change patient management in these scenarios. With the turnaround time of laboratory studies taking up to 4 days, there is a significant increase to the cost of patient care when solely relying on HITAb and SRA due to the use of expensive anticoagulants. In contrast, it remains unknown if HITAb and SRA could be useful in patients with an intermediate 4TS as our data is limited with no SRA results for patients with intermediate scores and a positive HITAb.To prevent unnecessary testing in the future and to improve the management of HIT, we propose to implement the following at our institution: 1. create a hard stop in our EMR which would prevent studies from being sent off inappropriately; 2. add a 4TS to the calculator section of the EMR and encourage collaboration with the hematology department if additional questions remain after calculating a 4TS; 4. start resident based educational sessions on the importance of calculating a 4TS and its significance prior to sending laboratory studies. In conclusion, the 4TS remains a useful tool to prevent unnecessary diagnostic testing and use of expensive therapeutic anticoagulants in patients with suspected HIT. DisclosuresNo relevant conflicts of interest to declare.

A novel diagnostic algorithm for heparin-induced thrombocytopenia.

While diagnostic algorithm using PF4-heparin enzyme-linked immunosorbent assay (ELISA) optical density (OD), and heparin neutralization assay (HNA), or 4T score have been proposed to replace serotonin-release assay (SRA) for heparin-induced thrombocytopenia (HIT), their performance against SRA is unclear. In this study, we proposed and validated the performance of a new algorithm combining PF4-heparin ELISA optical density (OD), HNA and 4T score against SRA for HIT diagnosis. Heparin neutralization assays were performed on specimens submitted for HIT testing with positive PF4-heparin ELISA from December 2015 to September 2017, which were separated into a "training" and a "validation" data set. 4T scores were calculated for ELISA positive cases. A total of 123 consecutive unique patient samples had positive PF4-heparin ELISA with also HNA data, SRA data, and 4T scores available. Compared to SRA, a "laboratory criteria" (ELISA OD≥1.4 and HNA≥70%) had a sensitivity of 88% (14/16) and specificity of 91% (42/46), and a "combined criteria" (4T score=8, or 4T score=6 or 7 and ELISA OD≥1.0, or 4T score=4 or 5 and ELISA OD≥2.0) had a sensitivity of 75% (12/16) and specificity of 98% (45/46) in the training data set. Laboratory and combined criteria had 90% (56/62) concordance rate. Importantly, for these concordant cases, the diagnostic specificity is 100% (46/46). Based on the data, a novel diagnostic algorithm combining these 2 criteria was proposed and validated prospectively. A novel algorithm has high diagnostic accuracy and potentially could eliminate the need for SRA testing in at least 90% patients with suspected HIT.

Evaluation of a Reflex Testing Algorithm for Suspected Heparin-Induced Thrombocytopenia.

We implemented a policy of reflex serotonin-release assay (SRA) testing for all patients with a positive heparin-induced thrombocytopenia (HIT) immunoassay. We identified all patients who had SRA testing sent as a consequence of a positive HIT immunoassay test. We reviewed charts of patients to calculate the 4Ts clinical score, determined the effect of testing on clinical management, and documented the change in utilization of direct thrombin inhibitors (DTIs). The likelihood of a positive SRA varied with the optical density (OD) of the immunoassay. The performance of the immunoglobulin G (IgG)-specific and polytypic enzyme-linked immunosorbent assay was not statistically different. Both OD and 4Ts score correlated with the likelihood of a positive SRA but demonstrated poor specificity. Discontinuation of DTIs in patients with negative SRAs resulted in decreased drug utilization. The IgG-specific HIT immunoassay OD correlates with the likelihood of a positive SRA but does not achieve high specificity. The reflex testing algorithm allows for definitive classification of patients, and the cost of such a reflex testing program may be offset by decreased utilization of DTIs.

Overtreatment of Heparin-Induced Thrombocytopenia in the Surgical ICU.

Recent studies reveal a high occurrence of overdiagnosis of heparin-induced thrombocytopenia in surgical patients with critical illness. The optimal criteria for diagnosis of heparin-induced thrombocytopenia remain unclear, contributing to unnecessary treatment. We reviewed patients who were admitted to surgical ICUs and were suspected of heparin-induced thrombocytopenia to identify how often patients were correctly treated. In this clinical prospective study, data were collected including age, sex, antiplatelet factor 4/heparin enzyme-linked immunosorbent assay, serotonin release assay, and Warkentin 4Ts scores. Heparin-induced thrombocytopenia-positive patients were defined as those with both positive antiplatelet factor 4/heparin enzyme-linked immunosorbent assay (optical density, ≥ 0.40) and positive serotonin release assay results. Urban tertiary medical center. Patients admitted to the surgical and cardiac ICU who were presumed to have heparin-induced thrombocytopenia and underwent antiplatelet factor 4/heparin enzyme-linked immunosorbent assay and serotonin release assay testing between January 1, 2011, and August 1, 2014. None. A total of 135 patients had 4Ts, antiplatelet factor 4/heparin enzyme-linked immunosorbent assay, and serotonin release assay scores. A total of 11 patients (8.1%) had positive serotonin release assay and 80 patients had positive antiplatelet factor 4/heparin enzyme-linked immunosorbent assay; 10 patients were identified as heparin-induced thrombocytopenia positive. Positive serotonin release assay was noted in nine of 11 patients (81.8%) with antiplatelet factor 4/heparin enzyme-linked immunosorbent assay optical density greater than or equal to 2.0, compared with one of 22 patients (4.5%) with optical density values of 0.85-1.99, and one of 102 patients (1.0%) with optical density values of 0-0.84. Out of 135 patients, 29 patients (21.5%) received treatment with argatroban, lepirudin, or fondaparinux: 10 of 10 heparin-induced thrombocytopenia-positive patients (100%) compared with 19 of 125 heparin-induced thrombocytopenia-negative patients (15%). Overtreatment of heparin-induced thrombocytopenia in the surgical ICU continues even with recent increased caution encouraging a higher antiplatelet factor 4/heparin enzyme-linked immunosorbent assay optical density threshold before initiating treatment. More stringent criteria should be used to determine when to order serologic testing and when the results of such testing should prompt a change in anticoagulant treatment. If antiplatelet factor 4/heparin enzyme-linked immunosorbent assay is used to consider immediate treatment, an optical density greater than or equal to 2.0 may be a more appropriate threshold.

A Novel PF4-Dependent P-Selectin Expression Assay (PEA) Facilitates Early Detection of Pathogenic HIT Antibodies: Implications for Diagnosis and Treatment of HIT

The PF4-dependent P-Selectin Expression Assay (PEA) is a novel assay for diagnosis of heparin-induced thrombocytopenia (HIT) which, in preliminary testing, demonstrates higher diagnostic accuracy than the "gold standard" serotonin release assay (SRA) (Padmanabhan et al. Chest, 2016 epub). Here, we describe two cases of severe thrombotic HIT in which the PEA detected pathogenic HIT antibodies (abs) before the SRA tested positive, and in one case, earlier than the highly sensitive PF4 ELISA. These findings have implications for diagnosis and monitoring of the response to therapy in HIT.Patient 1, a 67 yr old woman was admitted for treatment of a Bochdalek hernia and was started on unfractionated heparin (UFH). Her course was complicated by a declining platelet count and on the eighth day of admission PF4 ELISA and SRA tests were strongly positive (Fig 1A). Vascular imaging demonstrated venous thrombosis in both upper and lower extremities. Pre- and post-diagnosis patient samples were tested in the PF4 ELISA, SRA and PEA (Fig 1A). The PEA was positive at 152 hrs (~6 days) post-heparin exposure (69% expression), but the PF4 ELISA was only weakly positive (OD 0.67) and the SRA was negative (0%) at a time when the platelet count was 52% of baseline (Fig 1A). The SRA was still negative (18%) 16 hours later (168 hrs). Thirty hours after the first positive PEA test result, the SRA became positive (100% release). The patient's condition was then further complicated by sepsis-associated multisystem organ failure (MSOF) and therapeutic plasma exchange (TPE) was instituted. TPE treatment led to a rapid reduction in the SRA but only a gradual decline in PEA and ELISA test results (Fig 1B), in line with gradual improvement in platelet counts. Dissociation between SRA and PF4 ELISA test results following TPE has been described previously (Blood. 2015;125:195-8). In our patient, a PEA-SRA dissociation was also observed. Early in this patient's course, PEA-positive/SRA-negative abs were clearly "pathogenic," causing a significant decrease in platelet counts. Thus, it is plausible that PEA-positive/SRA-negative abs detected during the course of TPE were also "pathogenic". PEA results during the course of TPE appeared to correlate better with clinical status in light of the gradual improvement in platelet counts. Unfortunately, the patient expired after the 4th TPE secondary to complications of MSOF.Patient 2, a 57 yr old man was admitted for a creation of a thigh flap in preparation for knee replacement. Postoperatively, the flap became pale with poor pulses and UFH was started. On day 5 platelets dropped 25% from baseline and PF4 ELISA and SRA were both negative (Fig 1C). A strong suspicion of HIT prompted repeat testing on day 6, and yielded positive SRA (79%) and PF4 ELISA (2.8 OD) test results. Notably, the PEA was positive (74%) on Day 5 when both PF4 ELISA and SRA were negative. All tests remained strongly positive through day 12. Thrombocytopenia persisted (Fig 1C) and extensive venous thrombosis of the right upper extremity developed on day 15. The patient was transitioned to Coumadin after platelet recovery and discharged on Day 30.Previous studies indicate that the PEA is at least as accurate as the SRA for detection of pathogenic abs (Padmanabhan et al Thromb Haemost. 2015;114:1322-3, Padmanabhan et al. Chest, 2016 epub). Experience with these two cases suggests that the PEA may in some cases detect pathogenic antibodies before the SRA becomes positive and may be superior to the SRA for monitoring response to treatment. Because of its technical simplicity, the PEA has the potential to be performed on demand, rather than through a referral laboratory, facilitating early diagnosis and treatment. [Display omitted] DisclosuresPadmanabhan:Fenwal (Fresenius Kabi): Research Funding; Schlesinger & Associates: Consultancy, Honoraria; LEK Consulting: Consultancy, Honoraria; Mallinckrodt Pharmaceuticals: Consultancy, Honoraria; Terumo BCT: Consultancy, Honoraria; Bloodcenter of Wisconsin: Patents & Royalties: A patent application has been filed on a Method of detecting platelet activating antibodies that cause heparin-induced thrombocytopenia/thrombosis; PCT/US14/62591. Jones:Bloodcenter of Wisconsin: Patents & Royalties: A patent application has been filed on a Method of detecting platelet activating antibodies that cause heparin-induced thrombocytopenia/thrombosis; PCT/US14/62591. Bougie:Bloodcenter of Wisconsin: Patents & Royalties: A patent application has been filed on a Method of detecting platelet activating antibodies that cause heparin-induced thrombocytopenia/thrombosis; PCT/US14/62591. Aster:Bloodcenter of Wisconsin: Patents & Royalties: A patent application has been filed on a Method of detecting platelet activating antibodies that cause heparin-induced thrombocytopenia/thrombosis; PCT/US14/62591.