Preweaned heifer management on US dairy operations: Part V. Factors associated with morbidity and mortality in preweaned dairy heifer calves.

The objective of this study was to evaluate morbidity and mortality in preweaned dairy heifer calves based on different health, feeding, and management practices, as well as environmental factors. This study was conducted as part of the calf component of the National Animal Health Monitoring System's Dairy 2014 study, which included 104 dairy operations in 13 states. The calf component was an 18-mo longitudinal study focused on dairy heifer calves from birth to weaning; data were collected on 2,545 calves. The percentage morbidity for all calves enrolled in the study was 33.9%. Backward elimination model selection was used after univariate screening to determine which management practices and environmental factors significantly affected morbidity and mortality. The final morbidity model included birth weight, serum IgG concentration, ventilation type, and average temperature-humidity index (THI) during the preweaning period. After controlling for other independent variables in the model, calves born at a higher birth weight had a lower predicted risk of morbidity than calves with a lower birth weight. An increase in serum IgG concentration was associated with decreased morbidity. Calves housed in positive- or cross-ventilated systems had a 2.2 times higher odds of developing disease compared with calves housed in natural ventilation systems. Average THI during the preweaning period was inversely correlated with morbidity; as THI increased, the predicted morbidity risk decreased. The percent mortality for all calves enrolled in the study was 5.0%. The final mortality model included birth weight, serum IgG concentration, amount of fat/day in the liquid diet, and morbidity. After controlling for other independent variables in the model, calves born at a higher birth weight had a lower risk of mortality. An increase in serum IgG concentration decreased the risk of mortality. The odds of mortality were 3.1 times higher in calves fed ≤0.15 kg of fat/d in the liquid diet compared with calves fed ≥0.22 kg of fat/d. The odds of mortality were 4.7 times higher in calves that experienced any disease throughout the preweaning period than in calves with no disease. In summary, morbidity and mortality were both associated with birth weight and serum IgG concentration. Additionally, morbidity was associated with ventilation type and average monthly THI, and mortality was associated with amount of fat per day in the liquid diet and morbidity.

Preweaned heifer management on US dairy operations: Part II. Factors associated with colostrum quality and passive transfer status of dairy heifer calves.

Passive transfer of immunity is essential for the short- and long-term health of dairy calves. The objective of this study was to evaluate factors associated with colostrum quality and passive transfer status of US heifer calves. This study included 104 operations in 13 states that participated in the calf component of the National Animal Health Monitoring System's Dairy 2014 study. This 18-mo longitudinal study included 1,972 Holstein heifer calves from birth to weaning. Multivariable mixed linear regression models were selected using backward elimination model selection after univariate screening to determine which factors were associated with colostrum IgG and serum IgG concentrations. The mean colostrum IgG concentration was 74.4 g/L with 77.4% of colostrum samples having IgG concentrations >50 g/L. The final model for colostrum IgG included colostrum source and a categorized temperature-humidity index value (cTHI) for the month before calving. Mean colostrum IgG concentrations were highest for dams in third and higher lactations (84.7 g/L) and lowest for commercial colostrum replacers (40.3 g/L). Colostrum IgG concentrations were highest for cTHI ≥70 (72.6 g/L) and lowest for cTHI <40 (64.2 g/L). The mean serum IgG concentration was 21.6 g/L, with 73.3% of calves having serum IgG concentrations >15 g/L. The final model for serum IgG concentration included region, heat treatment of colostrum, colostrum source, timing to first feeding, volume of colostrum fed in the first 24 h, age of the calf at blood sampling, and colostrum IgG concentration. Mean serum IgG concentrations were highest for calves that received colostrum from first-lactation dams (25.7 g/L) and lowest for calves fed commercial colostrum replacer (16.6 g/L). Serum IgG concentrations were higher for calves fed heat-treated colostrum (24.4 g/L) than for calves fed untreated colostrum (20.5 g/L). Serum IgG concentration was positively associated with the volume of colostrum fed in the first 24 h and colostrum IgG concentration, and negatively associated with the number of hours from birth to colostrum feeding and age (days) at blood collection. Dairy producers should be encouraged to measure the quality of colostrum before administering it to calves and to measure serum IgG or a proxy such as serum total protein or Brix to evaluate passive immunity and colostrum management programs.

Preweaned heifer management on US dairy operations: Part I. Descriptive characteristics of preweaned heifer raising practices.

The objective of this study was to describe preweaned dairy heifer calf management practices on dairy operations across the United States that were used to analyze factors associated with colostrum quality and passive transfer, Cryptosporidium and Giardia, morbidity and mortality, and average daily gain. This study included 104 dairy operations in 13 states that participated in the National Animal Health Monitoring System's Dairy 2014 calf component study. This 18-mo longitudinal study focused on dairy heifer calves from birth to weaning, and data were collected on 2,545 heifer calves. Descriptive statistics were generated regarding colostrum feeding, preweaning housing, milk feeding and consumption, growth, morbidity and mortality, and weaning practices. The majority of calves enrolled were Holsteins (89.4%). Over half the calves (63.2%) enrolled in the study received the majority of their colostrum via bottle; however, 22.1% of calves from 51.0% of operations received colostrum via suckling from their dams. For all calves, the mean time to the first colostrum feeding was 2.8 h, and the average amount of colostrum at the first feeding was 2.9 L, with 4.5 L provided in the first 24 h. The mean serum IgG of all calves was 21.7 g/L; however, 76.0% of operations had at least 1 calf with failure of passive transfer of immunity with a serum IgG below 10 g/L. The majority of calves in the study were housed individually (86.6%). Nonetheless, 20.2% of operations housed some calves in groups, representing 13.4% of all calves. Approximately one-half of the calves in the study (52.3%) were dehorned or disbudded during the preweaning period, with only 27.8% of these calves receiving analgesics or anesthetics during the procedure. Whole or waste milk was the liquid diet type fed to 40.1% of calves, and milk replacer was fed to 34.8% of calves. A combination of milk and milk replacer was fed to 25.1% of calves. Calves, on average, were fed 2.6 L per feeding and fed 2.6 times/d, resulting in a total of 5.6 L of liquid diet fed per day. The mean average daily gain for all calves enrolled in the study was 0.7 kg/d. Fecal samples were collected and almost all operations had at least 1 calf positive for Cryptosporidium (94.2%) or Giardia (99.0%), and 84.6% of operations had calves that tested positive for both Cryptosporidium and Giardia. Over one-third of calves (38.1%) had at least one morbidity event during the preweaning period and the mortality rate was 5.0%. The mean age at weaning was 65.7 d. This study provides an update on dairy heifer raising practices in the United States.

Preweaned heifer management on US dairy operations: Part VI. Factors associated with average daily gain in preweaned dairy heifer calves.

The study objective was to evaluate average daily gain (ADG) in dairy heifer calves based on health, feeding, management practices, and environmental factors. This study included 102 operations in 13 states that participated in the calf component of the National Animal Health Monitoring System's Dairy 2014 study. This 18-mo longitudinal study included 1,410 Holstein heifer calves monitored from birth to weaning. The mean ADG from birth to final weight was 0.74 kg/d. Backward elimination model selection in Proc Mixed after univariate screening determined factors that significantly affected ADG. The final model included dam lactation number, singleton versus twin birth, bedding type, Giardia and Cryptosporidium fecal shedding, disease events, a categorized average temperature-humidity index for the preweaning period (pTHI), amount of protein in the liquid diet (kg/d), milk pasteurization, direct-fed microbials, and the interaction between milk pasteurization and direct-fed microbials. After controlling for other independent variables in the model, calves born to first-lactation dams gained less (0.60 kg/d) than calves from second- (0.65 kg/d) or third- or greater-lactation (0.64 kg/d) dams. Singleton calves gained 0.07 kg/d more than twins. Calves bedded with sand or no bedding gained less (0.49 kg/d) than calves on all other bedding types. Calves negative for Cryptosporidium or Giardia at the time of sampling gained 0.03 or 0.02 kg/d more, respectively, than calves that were positive for Cryptosporidium or Giardia. Calves with no disease events gained 0.07 kg/d more than calves with one or more disease events. Calves experiencing an average pTHI <50 gained more (0.67 kg/d) than calves experiencing an average pTHI from 50 to 69 (0.62 kg/d), or ≥70 (0.59 kg/d). Within the range of observed kilograms of protein fed per day in the liquid diet, every additional 0.1 kg of protein fed per day equated to 0.02 kg/d of gain. Calves fed milk replacer with a direct-fed microbial gained less (0.44 kg/d) than calves fed milk replacer without a direct-fed microbial (0.60 kg/d) and calves fed pasteurized or unpasteurized milk regardless of direct-fed microbial use. These results highlight the importance of feeding a quantity and quality of a liquid diet to achieve optimal growth, keeping calves free of disease, the type or status of bedding, and mitigating the effects of temperature and humidity on preweaning ADG.

Identification and Characterization of Isolates of Pythium and Phytophthora spp. from Snap Beans with Cottony Leak.

Cottony leak is an important disease of snap bean in Oklahoma and nearby states. Oomycete pathogens isolated from diseased pods collected from commercial fields and research plots consisted of both Pythium spp. (n = 131) and Phytophthora spp. (n = 46). Isolates were identified to species by morphological characteristics and by sequencing a portion of the internal transcribed spacer region of representative isolates. The most common Pythium spp. were Pythium ultimum var. ultimum; Pythium 'group HS', a self-sterile form of P. ultimum that produces hyphal swellings in lieu of sporangia (n = 74); and P. aphanidermatum (n = 50). Phytophthora spp. included Phytophthora drechsleri (n = 41) and P. nicotianae (n = 5). Nearly all of the isolates (95%) and all species were pathogenic on detached pods but Pythium ultimum var. ultimum and Pythium 'group HS' were most aggressive. Phytophthora drechsleri was most aggressive on seedlings, causing preemergence damping off and seed rot. Pythium ultimum var. ultimum, Pythium 'group HS', and P. aphanidermatum were intermediate in virulence to seedlings, causing root rot, stunting, and limited postemergence damping off. Phytophthora nicotianae and Pythium diclinum (n = 4) were not pathogenic on seedlings. Most (87%) isolates were sensitive to metalaxyl-M (concentration that caused a 50% reduction in mycelial growth [EC50] < 1 µg/ml) and the rest were intermediate in sensitivity (EC50 > 1 to < 100 µg/ml). Phytophthora drechsleri was the most sensitive species (EC50 = 0.06 µg/ml) compared with Pythium aphanidermatum, which was least sensitive (EC50 = 1.3 µg/ml). Cottony leak is a disease complex caused by several oomycete species that should include Phytophthora drechsleri, a newly reported pathogen of snap bean in the United States.

Reproductive risk factors for culling and productive life in large dairy herds in the eastern United States between 2001 and 2006.

Knowledge of reproductive risk factors for culling is useful in making insemination and culling decisions and helps motivate efforts to reduce or eliminate risk factors. The objective of this study was to describe survival and reproductive risk factors for culling in Holstein dairy herds with at least 200 cows. Results were calculated from 2,345,015 DHI lactation records from 727 herds with at least 200 cows from 2001 to 2006. Herds were located in 36 states primarily located east of the Mississippi River. Kaplan-Meier survival curves were obtained and daily hazards of culling were calculated with the actuarial method. Cox regression was performed with the GLIMMIX procedure in SAS (SAS Institute Inc., Cary, NC). The hazard of culling increased with parity number. Cows in their sixth parity had 3 times greater hazards than cows in their first parity. Medium remaining productive life for cows calving in parity 1 to 6 were 907, 697, 553, 469, 423, and 399 d, respectively. Daily hazards of culling first peaked approximately 30 d after calving and then again later in lactation, after 280 d, for older cows. Hazards for first-parity cows peaked earlier, around d 10 after calving, and the first-parity cows had lower risks of culling later in lactation than older cows. Pregnant cows had 3 to 7 times lower hazards of culling than open cows. Hazards of culling increased for cows that had greater calving difficulty, gave birth to males or twins, were in herds with shorter days to first insemination, or had longer days to conception. The possible to likely use of a synchronized breeding program increased from 21.9% in 2001 to 41.4% in 2006. Cows in herds that did not use a synchronized breeding program had slightly lower risks of culling than those in herds that at least possibly used a synchronized breeding program.

Fetal hemorrhage and platelet dysfunction in SLP-76-deficient mice.

The adapter protein SLP-76 is expressed in T lymphocytes and hematopoietic cells of the myeloid lineage, and is known to be a substrate of the protein tyrosine kinases that are activated after ligation of the T-cell antigen receptor. Transient overexpression of SLP-76 in a T-cell line potentiates transcriptional activation after T-cell receptor ligation, while loss of SLP-76 expression abrogates several T-cell receptor-dependent signaling pathways. Mutant mice that lack SLP-76 manifest a severe block at an early stage of thymocyte development, implicating SLP-76 in signaling events that promote thymocyte maturation. While it is clear that SLP-76 plays a key role in development and activation of T lymphocytes, relatively little is understood regarding its role in transducing signals initiated after receptor ligation in other hematopoietic cell types. In this report, we describe fetal hemorrhage and perinatal mortality in SLP-76-deficient mice. Although megakaryocyte and platelet development proceeds normally in the absence of SLP-76, collagen-induced platelet aggregation and granule release is markedly impaired. Furthermore, treatment of SLP-76-deficient platelets with collagen fails to elicit tyrosine phosphorylation of phospholipase C-gamma2 (PLC-gamma2), suggesting that SLP-76 functions upstream of PLC-gamma2 activation. These data provide one potential mechanism for the fetal hemorrhage observed in SLP-76-deficient mice and reveal that SLP-76 expression is required for optimal receptor-mediated signal transduction in platelets as well as T lymphocytes.

In situ site-specific specimen preparation for atom probe tomography.

Techniques for the rapid preparation of atom-probe samples extracted directly from a Si wafer are presented and discussed. A systematic mounting process to a standardized microtip array allows approximately 12 samples to be extracted from a near-surface region and mounted for subsequent focused-ion-beam sharpening in a short period of time, about 2h. In addition, site-specific annular mill extraction techniques are demonstrated that allow specific devices or structures to be removed from a Si wafer and analyzed in the atom-probe. The challenges presented by Ga-induced implantation and damage, particularly at a standard ion-beam accelerating voltage of 30 keV, are shown and discussed. A significant reduction in the extent of the damaged regions through the application of a low-energy "clean-up" ion beam is confirmed by atom-probe analysis of the damaged regions. The Ga+ penetration depth into {100} Si at 30 keV is approximately 40 nm. Clean-up with either a 5 or 2 keV beam reduces the depth of damaged Si to approximately 5 nm and <1 nm, respectively. Finally, a NiSi sample was extracted from a Si wafer, mounted to a microtip array, sharpened, cleaned up with a 5 keV beam and analyzed in the atom probe. The current results demonstrate that specific regions of interest can be accessed and preserved throughout the sample-preparation process and that this preparation method leads to high-quality atom probe analysis of such nano-structures.

Hyperspectral data processing improves PpIX contrast during fluorescence guided surgery of human brain tumors.

Fluorescence guided surgery (FGS) using aminolevulinic-acid (ALA) induced protoporphyrin IX (PpIX) provides intraoperative visual contrast between normal and malignant tissue during resection of high grade gliomas. However, maps of the PpIX biodistribution within the surgical field based on either visual perception or the raw fluorescence emissions can be masked by background signals or distorted by variations in tissue optical properties. This study evaluates the impact of algorithmic processing of hyperspectral imaging acquisitions on the sensitivity and contrast of PpIX maps. Measurements in tissue-simulating phantoms showed that (I) spectral fitting enhanced PpIX sensitivity compared with visible or integrated fluorescence, (II) confidence-filtering automatically determined the lower limit of detection based on the strength of the PpIX spectral signature in the collected emission spectrum (0.014-0.041 µg/ml in phantoms), and (III) optical-property corrected PpIX estimates were more highly correlated with independent probe measurements (r = 0.98) than with spectral fitting alone (r = 0.91) or integrated fluorescence (r = 0.82). Application to in vivo case examples from clinical neurosurgeries revealed changes to the localization and contrast of PpIX maps, making concentrations accessible that were not visually apparent. Adoption of these methods has the potential to maintain sensitive and accurate visualization of PpIX contrast over the course of surgery.

Indium-111 platelet scintigraphy and two-dimensional echocardiography for detection of left ventricular thrombus: influence of clot size and age.

Two-dimensional echocardiography and indium-111 platelet scintigraphy were performed on 50 dogs to determine the influence of clot age and size on the detection of experimentally induced left ventricular mural thrombus. Thrombus was induced by apical infarction and injection of a sclerosing agent and thrombin. The animals were classified into four groups according to the time of indium-111 platelet injection after thrombus induction: Group I (17 dogs, 1/2 hour after induction; 3 dogs, before induction), Group II (12 dogs, 24 hours after induction) and Group III (12 dogs, 1 week after induction). In Group IV (six control dogs) apical infarction was produced, but thrombin was not injected; indium-111 platelets were injected 1/2 to 1 hour after infarction. The dogs were studied by indium-111 platelet scintigraphy and by two-dimensional echocardiography 1/2 to 5 hours (Group I) and 1 to 5 and up to 72 hours (Groups II to IV) after platelet administration and before death was induced. Two-dimensional echocardiography showed the best overall sensitivity for detection of acute thrombus (97%; 29 of 30). The sensitivity of indium-111 platelet scintigraphy was 86% (18 of 21) for clots greater than or equal to 0.08 ml in size, and 67% (20 of 30) for detection of all clots. Thrombus did not form in 14 dogs of Groups I to III and in 6 of 6 control dogs. The specificity of scintigraphy was 100% (20 of 20) compared with 80% (16 of 20) for echocardiography. Echocardiography was more sensitive than scintigraphy for detecting very small clots in this experimental model.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunochemical detection using the light-addressable potentiometric sensor.

Rapid, sensitive and flexible assay systems are needed for immunoassays, receptor-ligand binding studies and DNA probe assays. Filtration capture and sensor detection offer several advantages to these areas. Although dependent on the affinity of the specific binders employed, the sensitivity of these techniques can be in the order of 10(-12) M, and total assay time can be less than 15 minutes.

Long-term outcome of low-grade oligodendroglioma and mixed glioma.

BACKGROUND: Low-grade oligodendrogliomas and mixed gliomas can be indolent and remain unchanged for years. Optimal timing and effectiveness of initial treatment is uncertain and therapy can be associated with toxicity. METHODS: Retrospective review of patients diagnosed between 1979 and 1997 with low-grade oligodendroglioma or mixed glioma. Time to progression, survival, prognostic factors, and treatment toxicities were evaluated. RESULTS: A total of 106 patients (77 oligodendroglioma, 29 mixed glioma) were identified; median age was 36.7 years. Initial presenting symptoms were seizures in 76 (72%) and headache in 11 (10%); tumor was diagnosed as an incidental finding in five patients. Tumor progression was diagnosed in 72 patients (68%). Overall median time to progression (MTTP) was 5.0 years (range 0.5 to 14.2). Median overall survival (OS) was 16.7 years. No prognostic factors reached statistical significance. MTTP and OS were not significantly affected by treatment. Of 62 patients who received radiation therapy, 9 (15%) developed radiation necrosis and 13 developed radiation therapy-related cognitive changes, requiring ventriculoperitoneal shunting in six. Significant myelosuppression was seen in 35 of 76 (46%) patients treated with chemotherapy. CONCLUSIONS: Low-grade oligodendroglioma and mixed glioma have a long median overall survival. There were no apparent differences in either immediate versus deferred treatment or choice of initial therapy on disease-free or overall survival. Chemotherapy was associated with significant acute toxicity in almost one half of patients; radiation therapy produced late neurotoxicity in one third, justifying deferred treatment until clinically necessary.

A silicon sensor-based filtration immunoassay using biotin-mediated capture.

A sensitive sandwich immunoassay for human chorionic gonadotropin (hCG) was developed with biotin-mediated filtration capture and silicon sensor detection. A high density of biotin on the membrane assured efficient capture of complexes containing streptavidin and analyte. Capture efficiency was not affected over a wide range of filtration flow rates or biotin concentrations. The assay utilized the pH sensing ability of the light addressable potentiometric sensor (LAPS) for the detection of urease-antibody conjugates. A LAPS reader was constructed which allowed the enzyme conjugate to be detected in approximately 1 microliter volumes. Effects from variations in detection volume were studied. 10 pg of hCG could be detected in an assay time of 20 min with four standard deviations separation from background. Comparison to a commercial RIA was made.

The spectrum of von Willebrand's disease revisited.

We have examined nine patients with presumed von Willebrand's disease who present the spectrum of that disorder. Two had findings that would be accepted generally as diagnostic of von Willebrand's disease, and seven had variations of the usual pattern. The commonest variation was the combination of borderline and variable levels of coagulant Factor VIII, commensurate levels of Factor VIII-related antigen, and low levels of ristocetin-Willebrand factor.

Cytokine and complement levels in patients undergoing cardiopulmonary bypass.

Patients undergoing cardiopulmonary bypass are known to develop whole body inflammation that often results in a characteristic syndrome early postoperatively. This phenomenon has been attributed to complement activation caused by exposure of blood to the foreign surfaces of the cardiopulmonary bypass circuit. It has been unknown if cytokines are involved. Plasma levels of complement activation products (C3a, C4a, C5a, and C5b-9), interleukins (IL-1 beta, IL-2, IL-4, and IL-6), and tumor necrosis factor-alpha were measured at multiple time points before, during, and after cardiopulmonary bypass in 29 patients. No significant increase over preinduction levels was seen in the cytokines except for IL-6, which was significantly increased during cardiopulmonary bypass (p < 0.001), reaching a maximum 3 hours after cardiopulmonary bypass. C3a, C4a, and C5b-9 levels were significantly elevated during cardiopulmonary bypass (p < 0.001), with maximum C5b-9 levels preceding the IL-6 elevation. Heparin coating of the cardiopulmonary bypass circuit was not demonstrated to have an effect on activation of complement or cytokine production. There was no statistically significant correlation among hemodynamic variables or pulmonary function and complement, interleukin, or tumor necrosis factor-alpha levels. These results confirm the presence of complement activation and demonstrate the production of IL-6 after the generation of C5b-9 in patients undergoing cardiopulmonary bypass. IL-6 may contribute to adverse systemic reactions associated with cardiopulmonary bypass.

Value of transesophageal echocardiography as an adjunct to transthoracic echocardiography in evaluation of native and prosthetic valve endocarditis.

To determine if transesophageal echocardiography provides better visualization of valvular vegetations than transthoracic echocardiography, we used both methods to evaluate 24 consecutive patients (mean age, 54 years; 15 female patients and nine male patients) referred for symptoms suggestive of infectious endocarditis. Ten of the 24 patients had one or more valvular prostheses. Echocardiograms were classified as positive or negative based on visualization of valvular vegetations or abscesses. Of ten patients with a final diagnosis of infectious endocarditis on extended follow-up, transthoracic echocardiography was positive in five patients. Transesophageal echocardiography not only yielded abnormal findings in all ten of these patients, but also revealed additional information in four of the five patients with abnormal transthoracic echocardiographic examinations. Among the 14 patients who, on subsequent follow-up, were found not to have infectious endocarditis, transthoracic echocardiography was normal in 13 and falsely abnormal in one. Transesophageal echocardiography revealed no evidence of infectious endocarditis in any of these patients. The ten patients who were determined to have infectious endocarditis all had positive blood cultures and no alternative cause for their clinical presentation; in seven patients in this group who underwent operative or postmortem evaluation, infectious endocarditis was confirmed. All patients without infectious endocarditis were demonstrated to have other causes for their clinical presentation. We conclude that transesophageal echocardiography is a highly valuable test in the work-up of patients with suspected infectious endocarditis, especially those patients with inconclusive or normal transthoracic echocardiograms. In addition, transesophageal echocardiography may be of benefit to patients with previously documented infectious endocarditis and a complicated clinical course in whom additional cardiac lesions are suspected but not demonstrated by transthoracic echocardiography.

Effect of varying fibrinogen and hematocrit concentrations on magnetic resonance relaxation times of thrombus.

OBJECTIVES: The authors assessed the effects of varying one extracellular component (fibrinogen concentration) and one cellular component (hematocrit) on magnetic resonance (MR) T1 and T2 relaxation times of in vitro blood clots. METHODS: Blood from six male subjects was collected into sodium citrate anticoagulant (3.8%) and the whole blood was separated into platelet-rich plasma and packed erythrocytes. Subsequently, in vitro blood clots were made from varying concentrations of fibrinogen (1, 10, and 100 microM) in Tyrode's solution and washed, packed erythrocytes (hematocrit levels: 0%, 10%, 40%, and 80%). T1 and T2 measurements were completed at 20 MHz within 8 hours of initiating clotting. RESULTS: Significant shortening of MR relaxation times occurred with increasing fibrinogen concentration for hematocrit values of 0% and 10%. Extracellular fibrinogen concentration did not contribute significantly to variation in relaxation times at hematocrit values of 40% and 80%. For any given fibrinogen level, significant shortening occurred in T1 and T2 values for each successive increase in hematocrit values. CONCLUSIONS: Both extracellular (fibrinogen) and cellular (erythrocyte concentration) factors are significant determinants of thrombus T1 and T2 relaxation times.

Magnetic resonance characterization of blood coagulation in vitro. Effect of platelet depletion.

To optimize magnetic resonance (MR) methods of characterizing thrombi, further studies of biologic determinants of spinlattice (T1) and spin-spin (T2) relaxation times of thrombi are needed. As a step toward evaluating the influence of thrombus cellular composition on MR properties, the authors evaluated the effect of platelet depletion on MR relaxation times of in vitro blood clots. Blood from 13 fasting normal men was collected into sodium citrate (3.8%) and centrifuged ( [10,000 X g] X 5-10 minutes). Platelet-poor specimens (less than 20,000 per mm3) were reconstituted from the plasma and packed erythrocytes to match precentrifugation hematocrit levels. T1 and T2 measurements were made at 20 MHz within three to six hours of initiating clotting. The mean T1 value for platelet-rich (normal) specimens was 1117 +/- 86 mseconds versus 1119 +/- 68 mseconds for the platelet-poor specimens (P greater than .90). The mean T2 value for platelet-rich (normal) specimens was 616 +/- 130 mseconds versus 434 +/- 79 mseconds for the platelet-poor specimens (P less than .001). The mean water content in the platelet-rich (normal) specimens was 79.5% +/- 1.2% versus 80.0% +/- 1.2% in the platelet-poor specimens (P greater than .50). In summary, platelet depletion by buffy coat removal significantly shortens MR T2 values of in vitro clot. These data suggest that thrombus cellular composition, other than erythrocytes, alters MR relaxation times of clotted blood.

Low interinstrument variability of the international normalized ratio with the coagamate X2/Simplastin excel system.

Minimizing the interlaboratory variability of the International Normalized Ratio (INR) for patients receiving coumarin therapy will require local laboratory calibration of individual coagulation instrument/reagent systems. However, some laboratories possess multiple coagulation instruments of the same model, and it is unclear if each instrument would require separate calibration. To address this question, a controlled study was performed that examined the interinstrument variability of the INR on three separate Coagamate X2 coagulometers (Organon Teknika, Durham, NC) using the Simplastin Excel reagent (Organon Teknika) (International Sensitivity Index = 2.14). The interinstrument coefficient of variation (CV) of the INR among the three instruments was 3.4% and 3.5% for patient control plasmas (n = 20) and coumarin plasmas (n = 40, INR range 1.5-4.6), respectively. The number of discordant INRs between paired instruments (one INR within and one INR out of the therapeutic range, plus a difference of at lest 0.4 INR units) was very low (0%). The interinstrument INR CVs for three commercial quality control plasmas were 2.6% (INR = 0.92), 4.1% (INR = 2.1), and 6.3% (INR = 4.5), and correlated well with the low CVs for patient samples. This study showed that the Coagamate X2/Simplastin Excel system is capable of very low interinstrument INR variability, and surpasses the interlaboratory CVs reported in the literature. Formal calibration of a single instrument may be adequate for laboratories possessing multiple instruments of the same model.

Evaluation of a new protamine titration method to assay heparin in whole blood and plasma.

When unfractionated heparin is used for therapeutic anticoagulation, the heparin effect must be monitored to avoid thrombotic or hemorrhagic complications. The ability of a factor Xa inhibition (XaI) assay was compared with that of a low-level heparin protamine titration (LLHPT) assay to measure the concentration of heparin after heparin was added in vitro to specimens of plasma and whole blood. Heparin effect on the activated partial thromboplastin time also was assessed in the same specimens. The XaI and LLHPT assays had comparable precision and provided linear results over a wide range of heparin concentrations. Both assays slightly underestimated the total amount of heparin added to the specimens. The most rapid test was the whole blood LLHPT assay; this test therefore may be useful for bedside monitoring of heparin. A significant disadvantage of the LLHPT assay was the large sample size required to perform it. These results provide in vitro evidence that the XaI and LLHPT assays can provide equally precise monitoring of heparin concentration.