Emergency assessment of hemostasis in the bleeding patient.

Emergency hemostasis testing is typically used to determine which blood products are needed to correct hemostatic defects associated with bleeding. Rapid assessment of hemostasis can be performed using standard or viscoelastic hemostasis tests in the clinical laboratory, satellite laboratory, or using point-of-care methods. The major drawback of standard coagulation testing is that most clinical laboratories are focused more on accuracy than on turnaround time. Improving turnaround times may require revision of the entire approach to performing the testing in the clinical laboratory including specimen processing, testing, and reporting. An advantage to rapid central laboratory testing is the availability to all areas of the hospital, not just the emergency department or operating rooms. Use of point-of-care assays can improve turnaround times, but point-of-care tests typically show more variation and lower precision. Viscoelastic global hemostasis tests can be used for rapid assessment as well, but care needs to be taken in the interpretation of the results. Viscoelastic testing is currently the only method that can detect severe fibrinolysis. Platelet function testing is useful for detection of hereditary platelet function problems and some antiplatelet medication monitoring, but is less useful for preoperative bleeding risk assessment or diagnosing the cause of acquire bleeding syndromes. This review will highlight different approaches to the rapid assessment of hemostasis.

A new microparticle size calibration standard for use in measuring smaller microparticles using a new flow cytometer.

BACKGROUND: Microparticle size measurements are often calibrated on flow cytometers using polystyrene microspheres that forward scatter more light vs. particle diameter than cellular microparticles. METHODS: We compared theoretical with measured forward angle light scattering on the LSRII, FC500 and Apogee A40 using polystyrene and silica microspheres vs. synthetic lipid vesicles and platelets, then compared plasma microparticle counts using different calibration strategies. RESULTS: Polystyrene and silica microspheres with higher refractive indices forward scattered more light with a wavelength of 488 nm for a given size microparticle than did lipid vesicles or platelets. The LSRII and FC500 did not count many, and were unable to separate by size, polystyrene microspheres <0.5 µm in diameter. On the Apogee A40, polystyrene microspheres could be separated by size down to 0.2 µm, and a polystyrene microsphere 0.4 µm in diameter produced the same forward scatter relative intensity as a 1-µm lipid or cellular microparticle. Using the new calibrator, the Apogee A40 found 80 000-4 000 000 µL(-1) total microparticles, 11 000-350 000 µL(-1) annexin V positive microparticles and 6000-350 000 µL(-1) platelet microparticles <1 µm in plasma samples. CONCLUSIONS: The Apogee A40 was able to resolve size differences in polystyrene microspheres down to 0.2 µm and microparticles down to 0.4 µm. On the Apogee A40 we propose using a 0.4-µm polystyrene microsphere as equivalent to a 1-µm cellular microparticle for size calibration. Using this calibrator, the Apogee A40 detected higher numbers of total, platelet and annexin V positive microparticles than were found using a Megamix gate.

Postmenopausal oral estrogen therapy affects hemostatic factors, but does not account for reduction in the progression of subclinical atherosclerosis.

BACKGROUND: Hemostatic factors influenced by postmenopausal hormone therapy may contribute to atherosclerosis. The Estrogen in the Prevention of Atherosclerosis Trial (EPAT), a 2-year, randomized, double-blind, placebo-controlled trial, demonstrated reduced subclinical atherosclerosis progression measured by change in common carotid artery intima-media thickness (CIMT) with unopposed oral 17beta-estradiol. OBJECTIVES: To assess the effect of postmenopausal hormone therapy on the levels of several hemostatic factors, and the relationship between these factors and the progression of subclinical atherosclerosis. PATIENTS AND METHODS: We measured tissue plasminogen activator (t-PA) antigen, factor (F) VII, D-dimer and albumin longitudinally, and plasminogen activator inhibitor type 1 (PAI-1) and fibrinogen at trial-end, in 186 postmenopausal women. RESULTS: Estradiol vs. placebo was associated with greater FVII and lower t-PA, albumin, PAI-1 and fibrinogen (all P < or = 0.001), with no estradiol effect on D-dimer (P = 0.42). Only mean on-trial t-PA was positively associated with the absolute level of CIMT on-trial (r = 0.29, P < 0.0001), but this was attenuated with age and body mass index adjustment. No longitudinally measured hemostatic factor was associated with CIMT progression. However, higher CIMT during the trial was significantly related to increases in t-PA. CONCLUSIONS: These results confirm previous findings regarding estrogen's effect on hemostatic factors and show that albumin is negatively associated with estrogen therapy. These hemostatic factors did not account for the reduction of CIMT progression with 17beta-estradiol seen in EPAT. Atherosclerosis itself may affect levels of hemostatic factors (reverse causality), with subsequent involvement in atherosclerosis-associated thrombosis.

Coagulation factor VII gene haplotypes, obesity-related traits, and cardiovascular risk in young women.

BACKGROUND: Associations between common F7 haplotypes, plasma factor VII (FVII) levels, and cardiovascular risk have recently been reported in population studies involving predominantly European men. METHODS: We assessed associations between F7 haplotypes and cardiovascular risk in two US population-based studies: a case-control study of these alleles related to a decreased risk of arterial thrombotic outcomes such as myocardial infarction (MI) in young-to-middle-aged women (n = 671), and a cohort study of cardiovascular disease risk factors in young women (n = 1040). RESULTS: The high-expression F7 haplotype B (containing the promoter variant allele -402A) was associated with an increased FVII level among controls, but not with MI risk. Women carrying a> or =1 copy of the low FVII expression level haplotype C (containing the -401T/-323del/-122C and Gln353 alleles) had decreased FVII levels and decreased risk of MI (odds ratio 0.54, 95% CI 0.31-0.93) compared with women homozygous for the most common haplotype A. Haplotype C was also associated with a decreased body mass index (BMI) and an increased high-density lipoprotein (HDL) cholesterol level, but not with MI risk after adjustment for these metabolic risk factors. In a cohort study composed of young US women, individuals homozygous for haplotype C had a lower BMI and lower systolic blood pressure, but the association between the F7 haplotype and HDL cholesterol was not confirmed. CONCLUSION: Common FVII haplotypes may contribute to the risk of MI in women, but the mechanisms appear complex. The association between F7 haplotypes and MI susceptibility may be mediated in part through an influence on atherogenic risk factors such as BMI.

Heritability of C-reactive protein and association with apolipoprotein E genotypes in Japanese Americans.

Numerous studies have demonstrated that increased C-reactive protein (CRP) levels predict coronary heart disease, stroke, peripheral vascular disease, and diabetes, and are associated with features of the metabolic syndrome. Only three previous studies have investigated the heritability of CRP levels, primarily in samples of Caucasian families. The purpose of the present study was to estimate the magnitude of genetic influences on CRP levels, and to examine potential associations between variation in the APOE gene and CRP levels, using a sample of 562 individual Japanese Americans from 68 extended kindreds. In general, correlation coefficients between first-degree relatives for CRP were approximately 0.2, and spouse correlations did not differ from zero, consistent with genetic influences. Heritability estimates were approximately 0.3 (p < 0.01), even with adjustment for factors known to influence CRP levels. A significant relationship was seen between unadjusted CRP levels and APOE genotypes (p = 0.02), with the highest mean CRP level among epsilon2 carriers (1.20 mg/L), and nearly the same mean levels among epsilon3/epsilon3 subjects and epsilon4 carriers (0.72 and 0.74 mg/L, respectively). However, this relationship was diminished with adjustment for covariates (p = 0.07). These results demonstrate the presence of both genetic and environmental effects on CRP levels among Asian Americans, and additional studies are needed to determine if the APOE gene contributes to these genetic influences.

Activation, aggregation and adhesion of platelets exposed to high-intensity focused ultrasound.

Using platelet-rich plasma, we investigated the effect of 1.1-MHz continuous wave high-intensity focused ultrasound (HIFU) on platelet activation, aggregation and adhesion to a collagen-coated surface. Platelets were exposed for durations of 10-500 s at spatial average intensities of up to 4860 W/cm(2). To avoid heating effects, the average temperature in the HIFU tank was maintained at 33.8 +/- 4.0 degrees C during platelet experiments. Flow cytometry, laser aggregometry, environmental scanning electron microscopy and passive cavitation detection were used to observe and to quantify platelet activation, aggregation, adhesion to a collagen-coated surface and associated cavitation. It was determined that HIFU can activate platelets, stimulate them to aggregate and promote their adherence to a collagen-coated surface. In principle, HIFU can stimulate primary, or platelet-related, hemostasis. Cavitation was monitored by a passive cavitation detector during aggregation trials and was quantified to provide a relative measure of the amount of cavitation that occurred in each aggregation trial. Regression analysis shows a weak correlation (r(2) = 0.11) between aggregation and ultrasound intensity, but a substantial correlation (r(2) = 0.76) between aggregation and cavitation occurrence.

Thrombogenic alleles, Escherichia coli O157:H7 infections, and hemolytic uremic syndrome.

Hemolytic uremic syndrome (HUS) of childhood most commonly follows gastrointestinal infection with Escherichia coli O157:H7. This pathogen elaborates Shiga toxins that are believed to cause microvascular injury and to trigger a thrombogenic response. The exact mechanisms leading to variable disease manifestations are unknown. Allelic variation in genes encoding selected coagulation factors and inhibitors of fibrinolysis were examined to determine whether or not a causal relationship exists between hypercoagulability and the development of HUS. No correlation between the thrombogenic factor V (G1691A), factor II (G20210A), methylenetetrahydrofolate reductase (C677T), or the plasminogen activator inhibitor (PAI)-1 promotor (4G/5G) genotypes and the risk of infection with E. coli O157:H7, or the risk of development of HUS among infected patients, was found. Serum PAI-1 levels did not correlate with the PAI-1 genotype. We conclude that the alleles studied are not major risk factors for the acquisition of E. coli O157:H7 infection, or of E. coli O157:H7-related HUS.

von Willebrand factor and von Willebrand factor-cleaving metalloprotease activity in Escherichia coli O157:H7-associated hemolytic uremic syndrome.

Hemolytic uremic syndrome (HUS) usually occurs after infection with Shiga toxin-producing bacteria. Thrombotic thrombocytopenic purpura, a disorder with similar clinical manifestations, is associated with deficient activity of a circulating metalloprotease that cleaves von Willebrand factor at the Tyr842-Met843 peptide bond in a shear stress-dependent manner. We analyzed von Willebrand factor-cleaving metalloprotease activity and the status of von Willebrand factor in 16 children who developed HUS after Escherichia coli O157:H7 infection and in 29 infected children who did not develop this complication. Von Willebrand factor-cleaving metalloprotease activity was normal in all subjects, but von Willebrand factor size was decreased in the plasma of each of 16 patients with HUS. The decrease in circulating von Willebrand factor size correlated with the severity of thrombocytopenia and was proportional to an increase in von Willebrand factor proteolytic fragments in plasma. Immunohistochemical studies of the kidneys in four additional patients who died of HUS demonstrated glomerular thrombi in three patients, and arterial and arteriolar thrombi in one patient. The glomerular thrombi contained fibrin but little or no von Willebrand factor. A decrease in large von Willebrand factor multimers, presumably caused by enhanced proteolysis from abnormal shear stress in the microcirculation, is common in HUS.

Factor XIIIA and clot strength after cardiopulmonary bypass.

Reduced factor XIIIA levels and decreased clot strength have been associated with increased bleeding after cardiopulmonary bypass (CPB). The purpose of this study was to evaluate the relationship between hemostatic factors, including factor XIIIA, and clot strength before, during and after CPB. Factor XIIIA antigen, platelet counts, fibrinogen, factor V activity, tissue plasminogen activator and clot strength (by thromboelastograph) were measured at baseline, after 45 min of CPB, at the end of CPB and 4 h post-operatively in 34 patients. Baseline factor XIIIA antigen was 5.2 +/- 1.4 mg/l. On average, factor XIIIA levels dropped to 64% and clot strength to 77% of baseline values after 45 min on CPB and remained below baseline during the immediate post-operative period. Clot strength was significantly correlated (r = 0.81) with platelet count and fibrinogen but not plasma factor XIIIA levels. Addition of 10 mg/l recombinant factor XIII[a2] significantly increased clot strength. Postoperative bleeding at 2 h was inversely correlated with platelet count, factor XIIIA antigen and clot strength measured at the end of CPB. Maintenance of adequate platelet counts and factor XIIIA levels at the end of CPB may play a role in maintaining clot strength and reducing blood loss.

Comparison of the lipoprotein, carbohydrate, and hemostatic effects of phasic oral contraceptives containing desogestrel or levonorgestrel.

Desogestrel (DSG) is a less-androgenic progestogen than levonorgestrel (LNG). This difference in androgenicity may be responsible for observed differences in metabolic effects between oral contraceptive (OC) formulations containing almost equivalent estrogen doses but with either DSG or LNG as a progestogen. To test the hypothesis, a prospective 9-month randomized comparison of plasma lipids, glucose, insulin, hemostasis, and sex hormone binding globulin (SHBG) was conducted in 66 healthy women using phasic formulations of OCs containing either DSG (DSG-OC) or LNG (LNG-OC). The study results showed that SHBG increased 3-fold with DSG-OC and 2-fold with LNG-OC. DSG-OC increased HDL-C, HDL(2)-C and HDL(3)-C; LDL-C decreased transiently. LNG-OC decreased HDL(2)-C and increased HDL(3)-C; HDL-C was unchanged and LDL-C decreased transiently. Both formulations increased VLDL-C and triglycerides, more with DSG-OC, but apolipoprotein B levels increased equally. Apo A-I and A-II increased more with DSG-OC than with LNG-OC. Neither formulation altered Lp(a) or fasting glucose and insulin levels. Postprandially, both formulations decreased glucose and increased insulin responses, but to an equivalent degree. Both OCs slightly enhanced procoagulant and profibrinolytic parameters to the same extent except for internally compensating decreases in Factor V and protein S with DSG-OC. In summary, at almost equivalent estrogen doses, a phasic OC containing DSG compared with LNG has a less androgenic effect on lipoproteins and SHBG, similar effects on hemostatic parameters with lower protein S and factor V activity and equivalent effects on carbohydrate metabolism. The lipoprotein, SHBG, and protein S and factor V differences are likely due to the lesser androgenicity of DSG allowing for a greater expression of the dose of estrogen.

A second-generation blood substitute (Perfluorodichlorooctane emulsion) generates spurious elevations in platelet counts from automated hematology analyzers.

UNLABELLED: Perfluorocarbon emulsions (PFEs) appear as platelets in automated cell counters, which may affect samples from thrombocytopenic patients (less than 100,000/microL). Therefore, we mixed clinically relevant concentrations of perfluorodichlorooctane (Oxyfluor(R); Hemagen, Inc., St. Louis, MO) in vitro with whole blood samples ranging from 0 to 150,000 platelets/microL and compared a new counter that uses optical platelet recognition (Abbott CellDyn 3200; Santa Clara, CA) with conventional electroimpedance-based counters (Abbott CellDyn 3500 and CellDyn 1700). We found that emulsion particles appear as small-sized platelets either in diluent or in blood. The emulsion results in a reproducible overestimate of the platelet counts, of greater importance as PFE concentration increases, and as the actual platelet count of the blood samples decreases. The new optical technology yields smaller overestimates but, even at low PFE concentrations, gives an unacceptable relative error at platelet counts near the transfusion thresholds recommended by the American Society of Anesthesiologists guidelines for blood component therapy. Unexpected interference in the leukocyte and erythrocyte channels is also reported. Experimental limitations preclude extrapolation of these findings to other automated cell counters, because differences in technology or software may affect their capacity to separate PFE particles from platelets. IMPLICATIONS: Perfluorocarbons are being investigated under conditions in which thrombocytopenia is likely to occur. In this in vitro study, we demonstrate significant overestimates in platelet counts from automated cell counters at clinically relevant perfluorocarbon concentrations in thrombocytopenic blood samples.

A kinetic model of the circulatory regulation of tissue plasminogen activator during orthotopic liver transplantation.

To better understand the regulation of tissue plasminogen activator (t-PA) and plasminogen activator inhibitor 1 (PAI-1) during liver transplantation, we used a computer model of the human circulatory system to simultaneously evaluate the effect of t-PA secretion, t-PA inhibition by PAI-1, hepatic clearance of t-PA, blood loss, transfusion and hemodynamics on t-PA and PAI-1 levels during liver transplantation in three patients that differed in severity of liver disease, blood loss and anhepatic changes in t-PA. Higher preoperative t-PA levels were primarily related to underlying liver disease and reduced hepatic clearance. During the anhepatic stage, when hepatic t-PA clearance was eliminated: (1) the expected rise in t-PA was modulated by the extent of bleeding, which acted as an alternate t-PA clearance mechanism; and (2) the ratio of t-PA:PAI-1 was increased due both to lower t-PA clearance and reduced PAI-1 secretion. Recirculation of the new liver was associated with renewed clearance of t-PA, an acute phase increase in PAI-1 and a drop in the t-PA:PAI-1 ratio. Understanding fibrinolytic regulation required simultaneous analysis of t-PA secretion, inhibition and clearance. Anhepatic t-PA levels could be predicted based on preoperative liver function and surgical blood loss, which acted as an alternate t-PA clearance mechanism.

Measurement of different forms of tissue plasminogen activator in plasma.

BACKGROUND: We evaluated assays to measure both total tissue plasminogen activator (tPA) and the three principle forms of tPA in plasma: active tPA, tPA complexed with plasminogen activator inhibitor type 1 (PAI-1), and tPA complexed with C1-inhibitor. METHODS: Active tPA was measured by use of an indirect amidolytic assay and immunofunctional assays. tPA/PAI-1, tPA/C1-inhibitor, and total tPA antigen were measured by use of microtiter plates coated with anti-tPA antibodies and, respectively, anti-PAI-1, anti-C1-inhibitor, and anti-tPA antibodies conjugated to peroxidase. RESULTS: The immunofunctional tPA assay detected 1 U/L (0.001 U/mL) tPA and recovered 108% +/- 12% of active tPA added to samples containing high (mean, 60 000 IU/L) PAI-1 activities vs a detection limit of 10 U/L (0.01 U/mL) and 13% +/- 25% recovery for the indirect amidolytic tPA activity assay. For measurement of tPA/PAI-1 complex, polyclonal anti-PAI-1 conjugates recovered 112% +/- 20% of the expected tPA/PAI-1 vs recovery of only 38% +/- 16% when monoclonal anti-PAI-1 conjugates were used. Of three methods tested, two total tPA antigen assays correlated well (r(2) = 0.85) and showed recoveries near 100%, whereas the third method showed lower correlations, higher intercepts, and falsely high recovery. A single anti-tPA capture antibody that performed the best in the individual assay evaluations was used to measure the different forms of tPA in 22 samples with a range of tPA and PAI-1 values. The sum of the molar concentrations of active tPA, tPA/PAI-1, and tPA/C1-inhibitor using the optimized methods was equal to 94% +/- 7% of measured total tPA. CONCLUSION: Optimized assays based on a single anti-tPA capture antibody can be used to accurately measure the major forms of tPA in plasma.

Formation, inhibition and clearance of plasmin in vivo.

A 5 microg/kg bolus of tissue plasminogen activator (t-PA) was infused into 11 healthy subjects followed by measurement of t-PA activity and antigen, PAI-1 activity and antigen, t-PA/PAI-1 complex, plasmin/antiplasmin (PAP) complex and D-dimer over 4 h. Infusion of t-PA resulted in a rise in PAP levels in all subjects from a baseline of 2.4 +/- 1.1 nmol/l to a peak of 5.1 +/- 2.3 nmol/l, but had no effect on plasminogen, antiplasmin or D-dimer levels. Using a kinetic model of plasminogen activation in vivo, the second-order rate constant for t-PA binding to plasminogen was estimated to be 3,100 +/- 1,300 mol(-1)l x s(-1), 200-500 times slower than t-PA accelerated by fibrin. The half-life of PAP was 4.5 +/- 1.6 h. In healthy subjects, the PAP level in plasma represents the average rate of plasminogen activation over the last 2-8 h.

Cerebrospinal fluid creatine kinase-BB isoenzyme activity and outcome after subarachnoid hemorrhage.

BACKGROUND: The brain is rich in creatine kinase-BB isoenzyme activity (CK-BB), which is not normally present in cerebrospinal fluid (CSF). Results of previous studies have shown that CK-BB can be detected in the CSF of patients with aneurysmal subarachnoid hemorrhage (SAH), but whether CK-BB levels correlate with patients' neurologic outcomes is unknown. OBJECTIVE: To evaluate the relationship between CSF CK-BB level and outcome after SAH. DESIGN: Prospective observational cohort. SETTING: University-affiliated tertiary care center. PATIENTS: Convenience sample of 30 patients seen for cerebral aneurysm clipping. INTERVENTIONS: We sampled and assayed CSF for CK isoenzymes a median of 3 days after SAH in 27 patients, and at the time of unruptured aneurysm clipping in 3 patients. MAIN OUTCOME MEASURES: Without knowledge of CK results, we assigned the Glasgow Outcome Scale score early (approximately 1 week) and late (approximately 2 months) after surgery. RESULTS: Higher CSF CK-BB levels were associated with higher Hunt and Hess grades at hospital admission (Spearman rank correlation, p = 0.69; P<.001), lower Glasgow Coma Scale scores at hospital admission (p = -0.72; P<.001), and worse early outcomes on the Glasgow Outcome Scale (p = -0.64; P<.001). For patients with a favorable early outcome (Glasgow Outcome Scale score, 3-5), all CK-BB levels were less than 40 U/L. With a cutoff value of 40 U/L, CK-BB had a sensitivity of 70% and a specificity of 100% for predicting unfavorable early outcome (Glasgow Outcome Scale score, 1-2). Having a CK-BB level greater than 40 U/L increased the chance of an unfavorable early outcome, from 33% (previous probability) to 100%, whereas a CK-BB level of 40 U/L or less decreased it to 13%. Similar findings were obtained when considering late outcomes. CONCLUSION: The level of CSF CK-BB may help predict neurologic outcome after SAH.

Effect of high-intensity focused ultrasound on whole blood with and without microbubble contrast agent.

Using human whole blood samples with and without contrast agent (CA), we evaluated the effect of exposures to focused, continuous wave (CW) 1.1-MHz ultrasound for durations of 10 ms to 1 s at spatial average intensities of 560 to 2360 W/cm2. Cavitation was monitored with a passive cavitation detector and hemolysis was determined with spectroscopy. In whole blood alone, no significant cavitation, heating or hemolysis was detected at any exposure condition. Conversely, cavitation and hemolysis, but not heating, were detected in whole blood with CA. A CA concentration as low as 0.28 microL CA per mL whole blood at an intensity of 2360 W/cm2 for 1 s resulted in measurable cavitation and a 6-fold increase in hemolysis compared to shams. Cavitation and hemolysis increased proportional to the concentration of CA and duration of exposure. In samples containing 4.2 microL CA per mL whole blood exposed for 1 s, a threshold was seen at 1750 W/cm2 where cavitation and hemolysis increased 10-fold compared to exposures at lower intensities. HIFU exposure of whole blood containing CA leads to significant hemolysis in vitro and may lead to clinically significant hemolysis in vivo.

Venous thrombus stability during acute phase of therapy.

The purpose of this study was to use serial venous duplex scans to document the status of deep venous thrombi during the early phase of therapy for acute, deep-vein thrombosis (DVT). A total of 71 consecutive participants treated for a first episode of acute DVT were monitored for new venous thrombosis using serial venous duplex scans. An average of 4.6 duplex scans were performed per patient (range, three to seven) during the 3-week study period. The cumulative incidence of contiguous/non-contiguous extension of the DVT at 3 weeks was 26% (95% CI = 14% to 38%). Nine of the 15 (60%) occurrences were asymptomatic. None of the classical risk factors for DVT was significantly associated with the development of new thrombi. The fraction of time during which the level of anticoagulation was considered 'adequate' (international normalized ratio > or =2.0 and/or heparin concentration > or =0.2 IU/ml) was inversely associated with the risk of extension/new thrombi (p = 0.01, Cox proportional hazards analysis). It was concluded that: (1) the frequency of contiguous/non-contiguous extension of venous thrombosis detectable during the first 3 weeks of therapy was higher than previously reported; (2) the majority of the occurrences were asymptomatic; and (3) the risk of developing this complication was inversely associated with the level of anticoagulation achieved.

A second-generation blood substitute (perfluorodichlorooctane emulsion) does not activate complement during an ex vivo circulation model of bypass.

OBJECTIVES: To examine whether a second-generation perfluorocarbon (PFC) blood substitute added to the cardiopulmonary bypass (CPB) prime influences complement production. DESIGN: A prospective, randomized, single-blinded, ex vivo model. SETTING: A university hospital, laboratory, and clinics. PARTICIPANTS: Ten healthy adult consented volunteer blood donors (five men, five women). INTERVENTIONS: Ex vivo closed-loop extracorporeal circuit including membrane oxygenator, tubing, and filter primed with crystalloid or crystalloid plus PFC was circulated for 1 hour with the addition of 500 mL of heparinized fresh human whole blood. MEASUREMENTS AND MAIN RESULTS: Laboratory specimens were drawn from the circuit at 10-minute intervals for 1 hour and measured for complement (C3a, Bb fragment) concentrations, blood gases, fibrinogen concentration, platelet count, and hematocrit. In the PFC group, C3a and Bb fragments were equal to or less than those in the group that received crystalloid alone. CONCLUSION: The second-generation PFC added to the prime of a CPB circuit does not independently increase complement production.