Serotonin-release assay-positive but platelet factor 4-dependent enzyme-immunoassay negative: HIT or not HIT?

IgG-specific and polyspecific PF4-dependent enzyme-immunoassays (EIAs) have exceptionally high sensitivity (≥99%) for diagnosis of heparin-induced thrombocytopenia (HIT), a drug reaction caused by platelet-activating antibodies detectable by serotonin-release assay (SRA). The IgG-specific EIAs are recommended for screening, as their high sensitivity is accompanied by relatively high specificity vis-à-vis polyspecific EIAs. We investigated the frequency of SRA-positive/EIA-negative (SRA+/EIA-) HIT, prompted by referral to our reference HIT laboratory of serial blood samples from a patient ("index case") with false-negative IgG-specific EIAs. Despite initial clinical suspicion for HIT, repeat negative IgG-specific EIAs prompted heparin resumption, which triggered recurrent thrombocytopenia and near-fatal cardiac arrest, indicating likely post-heparin HIT-associated anaphylactoid reaction. Further investigations revealed a strong-positive SRA, whether performed with heparin alone, PF4 alone, or PF4/heparin, with inhibition by Fc receptor-blocking monoclonal antibody (indicating IgG-mediated platelet activation); however, five different IgG-specific immunoassays yielded primarily negative (or weak-positive) results. To investigate the frequency of SRA+/EIA- HIT, we reviewed the laboratory and clinical features of patients with this serological profile during a 6-year period in which our reference laboratory investigated for HIT using both SRA and IgG-specific EIA. Although ~0.2% of 8546 patients had an SRA+/EIA- profile, further review of 15 such cases indicated clerical/laboratory misclassification or false-positive SRA in all, with no SRA+/EIA- HIT case identified. We conclude that while SRA+/EIA- HIT is possible-as shown by our index case-this clinical picture is exceptionally uncommon. Moreover, the requirement for a positive EIA is a useful quality control maneuver that reduces risk of reporting a false-positive SRA result.

Serotonin-release assay-negative heparin-induced thrombocytopenia.

Heparin-induced thrombocytopenia (HIT) is a prothrombotic drug reaction caused by platelet-activating anti-platelet factor 4 (PF4)/heparin antibodies. Pathogenic HIT antibodies can be detected by the serotonin-release assay (SRA), a platelet activation test. We have regarded the SRA performed in our medical community ("McMaster" SRA) as having high sensitivity and specificity. Recently, the concept of "SRA-negative HIT" has been proposed for enzyme-immunoassay (EIA)-positive/SRA-negative patients with a HIT-compatible clinical picture, who test positive in a PF4-enhanced platelet activation assay. After identifying an index case of SRA-negative HIT, we estimated the frequency of this condition by performing the "PF4-SRA" (modified SRA using high concentrations of added PF4 rather than heparin) in EIA-positive patients from a cohort study evaluating clinical and laboratory diagnosis of HIT. We defined SRA-negative HIT as patients meeting three criteria: clinical picture compatible with HIT (4Ts ≥ 4 points); EIA-positive (≥1.00 units); and PF4-SRA-positive. Among 430 patients, 35 were EIA-positive/SRA-positive and 27 were EIA-positive/SRA-negative. Among these 27 SRA-negative patients, three were found to have subthreshold levels of platelet-activating antibodies by PF4-SRA, of whom one met clinical criteria for SRA-negative HIT. Thus, based on identifying one patient with SRA-negative HIT within a cohort study that found 35 SRA-positive HIT patients, we estimate the sensitivity of the McMaster SRA for diagnosis of HIT to be 35/36 (97.2%; 95% CI, 85.8-99.9%). Although the McMaster SRA is highly sensitive for HIT, occasional SRA-negative but EIA-positive patients strongly suspected of having HIT can have this diagnosis supported by a PF4-enhanced activation assay such as the PF4-SRA.

Fondaparinux cross-reactivity in heparin-induced thrombocytopenia successfully treated with high-dose intravenous immunoglobulin and rivaroxaban.

HIT, a prothrombotic disorder caused by heparin-dependent antibodies, is often treated with fondaparinux, usually with good outcomes. A 70-year-old female developed severe HIT (platelet count, 25 × 109/L) post-glioblastoma resection during heparin thromboprophylaxis, complicated by disseminated intravascular coagulation (DIC) and symptomatic lower-limb deep-vein thrombosis (DVT). Despite therapeutic-dose fondaparinux, thrombocytopenia/hypofibrinogenemia persisted, with new symptomatic catheter-associated upper-extremity DVT. This clinical picture could be explained by autoimmune HIT (aHIT) refractory to fondaparinux or by fondaparinux cross-reactivity, so high-dose intravenous immunoglobulin (IVIG) was given (to treat possible aHIT) and fondaparinux switched to rivaroxaban, with subsequent clinical recovery. In vitro studies revealed strong fondaparinux cross-reactivity, without aHIT antibodies. Moreover, the patient's serotonin-release assay became negative post-IVIG, suggesting in-vivo inhibition of HIT antibody-induced platelet activation. Our case illustrates fondaparinux cross-reactivity in HIT manifesting as persisting thrombocytopenia, new thrombosis, and DIC, with successful rivaroxaban treatment, adding to emerging data that oral factor Xa inhibitors are efficacious for treating HIT.

Autoimmune heparin-induced thrombocytopenia and venous limb gangrene after aortic dissection repair: in vitro and in vivo effects of intravenous immunoglobulin.

BACKGROUND: Heparin-induced thrombocytopenia (HIT) is a prothrombotic disorder characterized by heparin-dependent antibodies that activate platelets (PLTs) via PLT FcγIIa receptors. "Autoimmune" HIT (aHIT) indicates a HIT subset where thrombocytopenia progresses or persists despite stopping heparin; aHIT sera activate PLTs strongly even in the absence of heparin (heparin-independent PLT-activating properties). Affected patients are at risk of severe complications, including dual macro- and microvascular thrombosis leading to venous limb gangrene. High-dose intravenous immunoglobulin (IVIG) offers an approach to interrupt heparin-independent PLT-activating effects of aHIT antibodies. CASE REPORT: A 78-year-old male who underwent cardiopulmonary bypass for aortic dissection developed aHIT, disseminated intravascular coagulation, and deep vein thrombosis; progression to venous limb gangrene occurred during partial thromboplastin time (PTT)-adjusted bivalirudin infusion (underdosing from "PTT confounding"). Thrombocytopenia recovered with high-dose IVIG, although the PLT count increase began only after the third dose of a 5-day IVIG regimen (0.4 g/kg/day × 5 days). We reviewed case reports and case series of IVIG for treating HIT, focusing on various IVIG dosing regimens used. RESULTS: Patient serum-induced PLT activation was inhibited in vitro by IVIG in a dose-dependent fashion; inhibition of PLT activation by IVIG was much more marked in the absence of heparin versus the presence of heparin (0.2 U/mL). Our literature review indicated 1 g/kg × 2 IVIG dosing as most common for treating HIT, usually associated with rapid PLT count recovery. CONCLUSION: Our clinical and laboratory observations support dose-dependent efficacy of IVIG for decreasing PLT activation and thus correcting thrombocytopenia in aHIT. Our case experience and literature review suggests dosing of 1 g/kg IVIG × 2 for patients with severe aHIT.

Plasma exchange to remove HIT antibodies: dissociation between enzyme-immunoassay and platelet activation test reactivities.

Repeated therapeutic plasma exchange (TPE) has been advocated to remove heparin-induced thrombocytopenia (HIT) IgG antibodies before cardiac/vascular surgery in patients who have serologically-confirmed acute or subacute HIT; for this situation, a negative platelet activation assay (eg, platelet serotonin-release assay [SRA]) has been recommended as the target serological end point to permit safe surgery. We compared reactivities in the SRA and an anti-PF4/heparin IgG-specific enzyme immunoassay (EIA), testing serial serum samples in a patient with recent (subacute) HIT who underwent serial TPE precardiac surgery, as well as for 15 other serially-diluted HIT sera. We observed that post-TPE/diluted HIT sera-when first testing SRA-negative-continue to test strongly positive by EIA-IgG. This dissociation between the platelet activation assay and a PF4-dependent immunoassay for HIT antibodies indicates that patients with subacute HIT undergoing repeated TPE before heparin reexposure should be tested by serial platelet activation assays even when their EIAs remain strongly positive.

Serological investigation of patients with a previous history of heparin-induced thrombocytopenia who are reexposed to heparin.

Heparin reexposure despite a history of previous heparin-induced thrombocytopenia (HIT) can be appropriate if platelet-activating antibodies are no longer detectable. We determined the frequency, timing, and magnitude of the antiplatelet factor 4 (anti-PF4)/heparin immune response (by serotonin-release assay [SRA] and enzyme-immunoassay [EIA]), and the frequency of recurrent HIT in 20 patients with previous HIT reexposed to heparin 4.4 years (mean) post-HIT; 17 patients were given heparin intraoperatively (without postoperative heparin) for cardiac/vascular surgery. One patient developed recurrent HIT beginning 7 days after cardiac surgery, with newly regenerated HIT antibodies exhibiting strong heparin-independent platelet-activating properties. Intraoperative heparin induced EIA seroconversion in 11/17 (65%) patients (immunoglobulin G [IgG]>IgA>IgM) and SRA seroconversion in 8/17 (47%), whereas none of 3 medical patients reexposed to heparin developed seroconversion. Anti-PF4/heparin IgG became detectable at day 7 (median), ie, no sooner than observed in typical-onset HIT. The high proportion of SRA positivity among EIA-seroconverting patients (8/11 [73%]) suggests that patients with previous HIT may be especially predisposed to forming recurrent antibodies with platelet-activating properties. We conclude that among patients with a previous history of HIT who are reexposed to intraoperative (but not postoperative) heparin, the risk of recurrent HIT appears to be low, but is possible if antibodies with strong heparin-independent platelet-activating properties are formed.

Investigation of anti-PF4 versus anti-PF4/heparin reactivity using fluid-phase enzyme immunoassay for 4 anti-PF4 disorders: classic heparin-induced thrombocytopenia (HIT), autoimmune HIT, vaccine-induced immune thrombotic thrombocytopenia, and spontaneous HIT.

BACKGROUND: Four platelet-activating anti-platelet factor 4 (PF4) disorders have been recognized: classic heparin-induced thrombocytopenia (cHIT), autoimmune heparin-induced thrombocytopenia (aHIT), spontaneous heparin-induced thrombocytopenia (SpHIT), and vaccine-induced immune thrombotic thrombocytopenia (VITT). All test immunoglobulin G (IgG) positive using solid-phase enzyme immunoassay (solid-EIA) against PF4/heparin (PF4/H) and/or PF4 alone. Fluid-phase EIA (fluid-EIA) should better discriminate between anti-PF4 and anti-PF4/H antibodies since conformationally altered PF4 bound to solid phase is avoided. OBJECTIVES: To compare anti-PF4 vs anti-PF4/H antibody profiles for anti-PF4 disorders using solid- and fluid-EIA. METHODS: We developed a novel fluid-EIA to measure anti-PF4 vs anti-PF4/H antibodies. RESULTS: Using fluid-EIA, 27 of 27 (100%) cHIT sera tested IgG positive with PF4/H, but only 4 of 27 (14.8%) tested positive against PF4 alone; all 27 exhibited heparin-enhanced binding. In contrast, 17 of 17 (100%) VITT sera tested IgG positive against PF4 alone, with markedly reduced binding against PF4/H; this distinct VITT antibody profile was not evident using solid-EIA. All 15 aHIT sera and all 11 SpHIT sera tested IgG positive against PF4 alone, with variable reactivity in PF4/H-EIA (heparin-enhanced binding in 14 of 15 and 10 of 11 aHIT and SpHIT sera, respectively). Remarkably, 1 SpHIT patient with a VITT-mimicking fluid-EIA profile (PF4 >> PF4/H) also clinically resembled patients with VITT (postviral cerebral vein/sinus thrombosis), with anti-PF4 reactivity correlating inversely with platelet count recovery; moreover, the single aHIT patient with a VITT-mimicking fluid-EIA profile also developed postviral cerebral vein/sinus thrombosis. CONCLUSION: cHIT and VITT sera showed opposite fluid-EIA profiles (cHIT: PF4/H >> PF4, with most testing negative against PF4 alone; VITT: PF4 >> PF4/H, with most testing negative against PF4/H). In contrast, all aHIT and SpHIT sera reacted against PF4 alone but with variable (usually enhanced) reactivity against PF4/H. VITT-mimicking clinical/serologic profiles occurred in only a minority of patients with SpHIT and aHIT.

Laboratory and demographic predictors of functional assay positive status in suspected heparin-induced thrombocytopenia: A multicenter retrospective cohort study.

Heparin-induced thrombocytopenia (HIT) is an antibody-mediated immune response against platelet factor 4 (PF4) bound to heparin anticoagulants. A priori identification of patients at-risk for HIT remains elusive and a number of risk factors have been identified, but these associations and their effect sizes have limited validation in large cohorts of suspected HIT patients. The aim of this study was to investigate existing anti-PF4/heparin antibody thresholds and model the relationship of demographic variables and anti-PF4/heparin antibody levels with functional assay positivity across multiple institutions in the absence of detailed clinical data. In a large collection of suspected HIT patients (n = 8904), we tested for associations between laboratory and demographic variables and functional assay positive status as well as anti-PF4/heparin antibody levels. We also tested for correlation between IgG-specific and polyspecific (IgG/IgA/IgM) anti-PF4/heparin antibody values and their ability to predict functional assay positive status using area under the receiver operating characteristic (AUROC). Logistic regression identified increasing anti-PF4/heparin antibody OD levels (OR = 51.84 [37.27-74.34], p < 2.0 × 10-16) and female sex (OR = 1.47 [1.19-1.82], p = 3.5 × 10-4) as risk factors for positive functional assay in the largest cohort with consistent effect sizes in two other cohorts. In a subset of 1175 patients, polyspecific and IgG-specific anti-PF4/heparin antibody values were heterogeneous (mean coefficient of variation = 31.9 %), but strongly correlated (rho = 0.878; p < 2 × 10-16) with similar prediction of functional assay positivity (polyspecific AUROC = 0.976 and IgG-specific AUROC = 0.980). Thus, we recapitulate previously identified risk factors of functional assay positivity, providing precise effect sizes in a large observational population of suspected HIT patients. Our data reinforce the necessity of functional assay confirmation and suggest that, despite heterogeneity, polyspecific and IgG-specific anti-PF4/heparin antibody assays predict functional assay positive status similarly, even in the absence of 4Ts scores and detailed clinical data.

Temporal presentations of heparin-induced thrombocytopenia following cardiac surgery: A single-center, retrospective cohort study.

BACKGROUND: Heparin-induced thrombocytopenia (HIT) is an important adverse drug reaction that can occur postcardiac surgery. Preoperative exposure to unfractionated heparin (UFH) is common, raising the issue of how frequently cardiac surgery-associated HIT occurs after immunizing preoperative exposure to heparin. OBJECTIVE: To determine the frequency and clinical picture of HIT occurring within 4 days of cardiac surgery (early presentation) versus later presentations (typical, delayed). METHODS: We identified patients with laboratory-confirmed HIT following cardiac surgery over 30 years in a single cardiac surgery center. Three different clinical presentations of HIT were identified: typical (HIT-related platelet count fall beginning between postoperative days [PODs] 5-10), delayed (patients with falls after POD10 or who presented following hospital discharge), and early (established before POD5, including during cardiac surgery [acute intraoperative HIT]). RESULTS: Of 129 patients identified with HIT complicating cardiac surgery, 100 had typical and 16 had delayed presentation of HIT; only 13 patients (10.1%) presented with early HIT, all of whom had received exposure to UFH during the 10 days before cardiac surgery. No patient was identified in whom remote preoperative UFH exposure was implicated in explaining early HIT. Notably, five patients appeared to have had acute intraoperative HIT, without immediate adverse consequences. CONCLUSIONS: Approximately 90% of patients with HIT after cardiac surgery appear to develop this complication due to immunization triggered by cardiac surgery; however, in approximately 10% of patients, early presentation during the first four PODs (or intraoperatively) can be explained by recent immunizing exposure to heparin.

Prospective multicentre cohort study of heparin-induced thrombocytopenia in acute ischaemic stroke patients.

Acute ischaemic stroke patients sometimes receive heparin for treatment and/or prophylaxis of thromboembolic complications. This study was designed to elucidate the incidence and clinical features of heparin-induced thrombocytopenia (HIT) in acute stroke patients treated with heparin. We conducted a prospective multicentre cohort study of 267 patients who were admitted to three stroke centres within 7 d after stroke onset. We examined clinical data until discharge and collected blood samples on days 1 and 14 of hospitalization to test anti-platelet factor 4/heparin antibodies (anti-PF4/H Abs) using an enzyme-linked immunosorbent assay (ELISA); platelet-activating antibodies were identified by serotonin-release assay (SRA). Patients with a 4Ts score ≥4 points, positive-ELISA, and positive-SRA were diagnosed as definite HIT. Heparin was administered to 172 patients (64·4%: heparin group). Anti-PF4/H Abs were detected by ELISA in 22 cases (12·8%) in the heparin group. Seven patients had 4Ts ≥ 4 points. Among them, three patients (1·7% overall) were also positive by both ELISA and SRA. National Institutes of Health Stroke Scale score on admission was high (range, 16-23) and in-hospital mortality was very high (66·7%) in definite HIT patients. In this study, the incidence of definite HIT in acute ischaemic stroke patients treated with heparin was 1·7% (95% confidence interval: 0·4-5·0). The clinical severity and outcome of definite HIT were unfavourable.

The temporal profile of the anti-PF4/heparin immune response.

The immune response in heparin-induced thrombocytopenia (HIT) is puzzling: heparin-naive patients can develop IgG antibodies and clinical HIT as early as day 5, and evidence for an anamnestic response on heparin reexposure is lacking. We assessed daily serum samples by anti-PF4/heparin enzyme-immunoassay (EIA) in patients receiving heparin thromboprophylaxis. Of 435 patients, 56.1% showed an increase in EIA optical density (OD) of more than or equal to 15%, with more than 90% starting between days 4 and 14. After reaching maximum reactivity by days 10 to 12, ODs declined despite heparin continuation, including in 2 patients with clinical HIT. Individual IgG/A/M classes showed identical time of onset (median, day 6). Most (58.7%) antibody-positive patients developed all 3 Ig classes; only 11.3% lacked IgG response. IgG/A/M increase usually occurred simultaneously (+/- 1 day) with no general tendency for IgM precedence. Consistent with the transient immune response, none of the IgG-EIA-positive (OD > 0.5) patients at discharge developed clinically evident thrombosis during extended low-molecular-weight heparin thromboprophylaxis. The rapid onset of the anti-PF4/heparin immune response, its transience, and the simultaneous appearance of antibodies of different classes with no IgM precedence suggest short-term activation of B cells that have previously undergone Ig-class switching even without previous pharmacologic heparin exposure.

Studies of the immune response in heparin-induced thrombocytopenia.

Heparin-induced thrombocytopenia (HIT) is caused by platelet-activating antibodies that recognize PF4/heparin complexes. Uncertainties remain regarding HIT immunobiology, including the temporal relation of antibody formation to onset of thrombocytopenia, and whether immunoglobulin class switching occurs. Using serial plasma samples from 2 heparin thromboprophylaxis trials, we determined the time of onset, antibody levels, and immunoglobulin class distributions (IgG, IgA, IgM) for 12 patients with HIT and 36 patients who formed anti-PF4/heparin antibodies, but did not develop HIT ("seropositive non-HIT controls"). In patients with HIT, anti-PF4/heparin antibodies became detectable 4 days (median) after starting heparin; antibody detection preceded the platelet count decline by 2 days (median). Patients with HIT produced higher levels of IgG antibodies, but similar IgA and IgM levels, compared with seropositive non-HIT controls. Among all 48 seroconverting patients, the first day of a positive antibody test (median, day 6) did not differ among the immunoglobulin classes. Thus, the HIT immune response does not exhibit the classic paradigm of IgM class precedence/immunoglobulin class switching; rather, relatively rapid formation of IgG antibodies is observed, sometimes with concomitant IgA and IgM formation. Compared with seropositive non-HIT controls, HIT patients develop significantly higher anti-PF4/heparin IgG levels that are detectable before the onset of thrombocytopenia.

High-dose IVIG plus cangrelor platelet "anesthesia" during urgent heparin-CPB in a patient with recent SRA-negative HIT-thrombosis with persisting platelet-activating antibodies.

In a high-risk patient with subacute heparin-induced thrombocytopenia (HIT) type A (platelet count recovery following acute HIT but with persisting platelet-activating antibodies), in whom urgent cardiac surgery was required, a key clinical question arose: could intraoperative heparin be given safely with "platelet anesthesia" provided with high-dose intravenous immunoglobulin (IVIG) plus cangrelor (ultra-short-acting antiplatelet agent)? This approach proved successful, without unexpected postoperative thrombocytopenia or thromboembolism. In vitro studies confirmed that both IVIG and cangrelor contributed to perioperative inhibition of HIT antibody-induced platelet activation. Interestingly, despite the patient testing strongly positive in 4 HIT immunoassays (latex immunoturbidimetric assay and 3 enzyme-immunoassays), the serotonin-release assay (SRA) was consistently negative. Nevertheless, platelet-activating HIT antibodies were detectable using modified (platelet factor 4-enhanced) SRA. Our protocol of heparin rechallenge following IVIG/cangrelor provides both intraoperative and early postoperative inhibition of HIT antibody-induced platelet activation and is applicable to patients with circulating functional HIT antibodies requiring urgent heart surgery, including those with "SRA-negative HIT."

Combination of two complementary automated rapid assays for diagnosis of heparin-induced thrombocytopenia (HIT).

BACKGROUND: HIT diagnosis typically uses complementary diagnostic assays (eg, a PF4-dependent enzyme-immunoassay [EIA] and a platelet activation assay such as the serotonin-release assay [SRA]). OBJECTIVES: To determine whether the combination of two automated assays-a latex immunoturbidimetric assay (LIA) that evaluates competitive inhibition of a HIT-like monoclonal antibody and a chemiluminescence immunoassay (CLIA) for detecting anti-PF4/heparin IgG-optimizes diagnostic sensitivity while also yielding good specificity, particularly at high assay reactivities. PATIENTS/METHODS: We determined operating characteristics using combined LIA/CLIA results from a HIT observational trial (n = 430; derivation cohort) and 147 consecutive patients with HIT (n = 147; supplementary derivation cohort). We also evaluated 678 consecutive samples referred for HIT testing (replication cohort). LIA/CLIA reactivities were scored individually as "negative" (<1.00 U/mL, 0 points), "weak" (1.00-4.99 U/mL, 1 point), "moderate" (5.00-15.99 U/mL, 2 points) and "strong" (≥16.00 U/mL, 3 points), thus contributing up to 6 points (maximum) when LIA/CLIA results were combined. We also examined whether higher LIA/CLIA scores predicted presence of platelet-activating antibodies by conventional and modified (PF4- or PF4/heparin-enhanced) SRA. RESULTS: Combined LIA/CLIA testing yielded high diagnostic sensitivity (~99%) similar to EIA. Interpretation of LIA/CLIA results using the 6-point scale indicated progressively greater likelihood for the presence of platelet-activating antibodies with increasing scores (semi-quantitative reactivity). A LIA/CLIA score ≥ 4 points predicted the presence of platelet-activating antibodies by SRA or PF4-enhanced SRA with high probability (~98%). CONCLUSION: Combined LIA/CLIA testing optimizes diagnostic sensitivity, with progressively greater probability of detecting platelet-activating antibodies with higher assay reactivity that reaches 98% when both automated assays yield moderate or strong results.

Immune mechanisms in heparin-induced thrombocytopenia: no evidence for immunoglobulin M anti-idiotype antibodies.

BACKGROUND: Heparin-induced thrombocytopenia (HIT), which is caused by platelet (PLT)-activating immunoglobulin (Ig)G antibodies against platelet factor 4 (PF4)/heparin complexes, differs from other immune responses seen in immunohematology: IgG antibodies are formed as early as 5 days even without previous heparin exposure; antibodies are remarkably transient (<100 days); HIT is more frequent in postsurgery patients compared with medical patients despite administering the same type and dose of heparin; and increasing evidence implicates autoantibody-like reactivity of anti-PF4/heparin antibodies. We hypothesized that these unusual features could be caused by loss of regulatory anti-idiotype IgM antibodies due to disturbance (e.g., by surgery) of an idiotype-anti-idiotype network. STUDY DESIGN AND METHODS: Sera were obtained prospectively before heparin administration and during the immunization phase of HIT and also from patients with previous HIT after waning of antibodies to nondetectable levels. To detect inhibitory IgM anti-idiotype antibodies, we performed serum coincubation experiments and IgG purification by protein G and size filtration to exclude coprecipitating IgM. Sera (n = 3) containing known anti-PF4/heparin IgG or IgM antibodies and normal sera (n = 20) were processed as controls. RESULTS: Fifteen preimmune response sera (seroconverting in the PF4/heparin-IgG enzyme-linked immunosorbent assay only [n = 4] or additionally in a PLT activation assay [n = 5] or in both assays plus thrombosis [n = 6]) and four sera of previously immunized patients were included. Neither did the neat sera inhibit binding of anti-PF4/heparin antibodies nor did the purified IgG fractions show enhanced binding to PF4/heparin complexes. CONCLUSION: The atypical immunologic features of HIT do not appear to be caused by disruption of an idiotype (IgG)-anti-idiotype (IgM) network.

Neutrophil and Monocyte Counts in Heparin-Induced Thrombocytopenia.

BACKGROUND: Heparin-induced thrombocytopenia (HIT) antibodies activate platelets, monocytes and neutrophils. Despite these findings, it is unknown whether white blood cell (WBC) counts, including neutrophils and monocytes, are altered during HIT. MATERIALS AND METHODS: We evaluated changes in total WBC counts (including WBC subsets), in 50 post-cardiac surgery patients with serologically confirmed HIT (30 patients with HIT-associated thrombosis). Daily leukocyte counts were compared with those measured one day prior to HIT onset; WBC increases were classified as mild (20.0-49.9%), moderate (50.0-99.9%) or major (≥ 100% increase). We also compared changes in WBC counts in HIT patients with and without HIT-associated thrombosis, and non-HIT patients with thrombosis. RESULTS: Most (34/50 [68.0%]) patients with HIT developed WBC count increases (mild, 35.3%; moderate, 44.1%; major, 20.6%). The peak WBC count occurred on day 4 (median) of HIT, which corresponded to day 10 (median) post-surgery. Absolute neutrophil counts increased in most patients (38/50 [76.0%]); whereas absolute monocyte counts rose in some patients, the overall tendency was for the monocyte count to decrease during HIT. Unexpectedly, we found that the increase in total WBC counts, as well as in neutrophils, was seen mainly in patients who developed HIT-associated or non-HIT-associated thrombosis; in contrast, no difference in monocyte levels was seen in patients with or without thrombosis. CONCLUSION: Leukocytosis and neutrophilia are commonly observed in patients with HIT, particularly in patients with HIT-associated thrombosis, as well as non-HIT patients with thrombosis. Thus, leukocytosis/neutrophilia should not infer automatically a diagnosis of infection or inflammation, when evaluating thrombocytopenia in heparin-exposed patients.