The impact of pneumatic tube transport on whole blood coagulation and platelet function assays

Pneumatic tube is an attractive way to transport blood samples from the emergency department to the central laboratory facility. We aimed to investigate the impact of pneumatic tube transportation on blood samples for analysis of whole blood coagulation and platelet function.

We included 21 healthy adult individuals and measured global coagulation assays by rotational thromboelastometry (ROTEM) and platelet aggregation induced by arachidonic acid (AA) and adenosine diphosphate (ADP) using impedance aggregometry (ROTEM Platelet), on samples transported manually or by pneumatic tube transport. Statistical testing was performed with paired tests with post-hoc Bonferroni correction for multiple testing.

Our data revealed no difference in the far majority of ROTEM parameters (P > 0.003), while significantly decreased values were observed for INTEM clotting time (CT) (P = 0.002) and maximum clot firmness (MCF) including the amplitude after 10 min (A10) (P < 0.0001).

No statistically significant difference was observed on impedance aggregometry results when manual transport was compared to pneumatic tube transport (P > 0.003).

This study indicates that only minor and unsystematic differences between manual transport and pneumatic tube transport may be observed in ROTEM analyses, and that there is no influence from pneumatic tube transport on impedance aggregometry analyses using AA and ADP.     

Comparison of in vitro responses to fresh whole blood and reconstituted whole blood after collagen stimulation

Background

In patients who have large bleeds, there is a tendency to transfuse more plasma and platelets than recommended in earlier guidelines, and accordingly many hospitals now provide “transfusion packages” with an intended red cell:platelet:plasma ratio of 1:1:1. The purpose of this study was to investigate in vitro functions of transfusion packs compared with fresh whole blood.

Material and methods

“Reconstituted whole blood” was prepared with the same ratio of red cells, platelets and plasma as used in local transfusion packages. The aggregation and thrombin-antithrombin complex formation responses to collagen stimulation of this reconstituted whole blood were compared with those of fresh whole blood. The storage time of red cells and platelets was varied in a systematic manner, giving nine different compositions of reconstituted whole blood that simulated transfusion packs.

Results

The responses varied significantly between whole blood and reconstituted whole blood -and between the reconstituted whole blood of different compositions. A significant decrease (p<0.005) in collagen-induced platelet count reduction was seen with increasing platelet and red blood cell age. Thrombin-antithrombin complex formation peaked in studies with platelets stored for 5 days. The red cells stored for the longest time induced the greatest thrombin-antithrombin complex formation. Fresh whole blood gives more consistent responses, and the aggregation response to collagen is stronger than in reconstituted whole blood.

Discussion

Our results indicate that in vitro responses of reconstituted whole blood vary substantially according to how long the red cells and platelets are stored for. As the responses obtained by testing whole blood are more consistent and usually stronger, the alternative use fresh whole blood in special conditions should not be excluded without further consideration.     

Post-operative hypercoagulable whole blood profiles in patients undergoing open thoracotomy vs video-assisted thoracoscopic surgery

BackgroundPatients undergoing video-assisted thoracoscopic surgery (VATS) have a lower risk of thrombosis compared to those undergoing open thoracotomy (OT) which may be due to several post-operative factors such as early mobilisation, shorter hospital stays, lower transfusion rates and lower risk of infections. Whether the higher thrombotic risk after OT is also linked to a peri-operative hypercoagulable state is a matter of debate. We therefore conducted a case-control study to compare peri-operative coagulation profiles in patients with primary lung cancer undergoing VATS vs OT.Materials and methodsAll consecutive patients undergoing VATS or OT for primary lung cancer at the Department of Thoracic Surgery of Padua University Hospital, Italy, between February and June 2018 were enrolled. Each patient provided a venous blood sample at least 30 min prior to surgical incision (T0) and 4±1 days after surgery (T1). Peri-operative coagulation profiles were assessed via traditional, viscoelastic whole blood (ROTEM^® [Instrumentation Laboratory-Werfen]) and impedance aggregometry (Multiplate^® Analyser [Roche Diagnostics]) tests.ResultsWe enrolled 65 patients (males 43, females 22; mean age 65±13 years) of whom 35 (54%) underwent VATS and 30 (46%) underwent OT. Compared to healthy controls, the surgical group (VATS and OT patients) had a significantly shorter clot formation time and higher alpha angle and maximum clot firmness values, as well as increased mean platelet function. In the post-operative period, patients who underwent OT had a significantly shorter clot formation time, higher alpha angle and maximum clot firmness values and higher mean platelet function vs VATS patients.DiscussionWhole blood ROTEM^® profiles and Multiplate^® aggregometry identified a more hypercoagulable post-operative state in patients who underwent OT than in those who underwent VATS. Larger studies are warranted to confirm our results and ascertain whether the observed hypercoagulability might promote post-operative thrombosis.     

Plasma and whole blood viscosity in treated primary polycythaemia

Whole blood viscosity at a range of shear rates (230-0.77 s^_1) and plasma viscosity have been measured in 39 patients with treated primary polycythaemia (idiopathic erythrocytosis and primary proliferative polycythaemia) and 30 age-matched normal individuals. There was a wider range of plasma viscosity and whole blood viscosity values at the same haematocrit (0.46), particularly at the lower shear rates, in the‘polycythaemic’group than the normal group. Hypochromic microcytic red cell changes present in 14 patients in the‘polycythaemic’group did not have any noticeable influence on whole blood viscosity at a given haematocrit value, since plasma protein factors override any possible effect of these red cell changes. The range of observed whole blood viscosity results in the‘polycythaemic’patients at the same haematocrit (0.46) was equivalent to the effect on whole blood viscosity of a rise in haematocrit from 0.41 to 0.51. Since there is such a range of whole blood viscosity at the same haematocrit, the haematocrit alone does not necessarily give a precise assessment of the viscosity of a whole blood sample. Since there is evidence from other publications that blood flow in vivo may be critically influenced by whole blood viscosity, this lack of precision should be considered when treating patients at risk of vascular occlusive episodes.     

Characteristics of leucocyte adhesion directly observed in flowing whole blood in vitro

Use of whole blood in adhesion assays allows analysis of the rheological and haematological factors that may influence adhesion, and avoids the need for isolation procedures that may modify the properties of leucocytes. We have adapted an in vitro flow model to allow videomicroscopy of leucocytes fluorescently labelled with rhodamine 6G (20 microg/ml) in anticoagulated whole blood. Blood was perfused at a range of wall shear rates (35-280/s) through a vertical glass capillary with a rectangular cross-section (microslide) that had been coated with P-selectin (10 microg/ml). Nearly all adherent cells were rolling in blood that had been anticoagulated with buffered citrate, but 40-50% became immobilized when heparin or thrombin inhibitor (PPACK) were used. The efficiency of leucocyte adhesion decreased steadily during 1-4 h of blood storage. Smaller fluorescent cells (lymphocytes) adhered less efficiently than larger cells (granulocytes) and rolled faster. Adhesion decreased monotonically with increasing wall shear rate or stress, but the velocity of rolling varied little. Among healthy volunteer donors, adhesion correlated with blood leucocyte count, but did not vary significantly with natural variation in haematocrit, blood viscosity or red cell aggregation. In conclusion, we have characterized adhesion of leucocytes in flowing whole blood, identified key experimental variables and demonstrated that physical environmental factors can markedly influence adhesive behaviour.     

An all leucocyte whole blood lyse method for multiparameter flow-cytometry

Summary A newly designed lyse-reagent for processing of whole blood samples for multiparameter flow-cytometry (MP-FCM) allows for flow-cytometrical determinations without any centrifugation during processing of samples. Comparing sample-processing by an FDA-approved lyse-reagent and by the newly designed lyse-reagent, two major advantages of the new lyse-reagent turned out: i) substantially higher recovery (about 100%) of leucocytes and ii) no selective loss of lymphocytes. Encouraged by the finding to have for determinations by MP-FCM almost all leucocytes as present in blood, MP-FCM-determinations of a particular small subset of leucocytes, namely of basophil granulocytes, have been compared to determinations of basophils, as performed by a routine haematological analyser (NE-7000). Along with a fairly good correlation (correlation coefficient about 0.8), there was no statistical significant difference in the percentages as determined by both technologies. The present data indicated that the new lyse-reagent for processing of whole blood for MP-FCM, due to the fact that leucocytes as present in blood are present for determinations by MP-FCM, may facilitate to establish MP-FCM-defined leucocyte-differential as a reference-method for the leucocyte-differential.     

Extended storage of whole blood before the preparation of blood components: in vitro effects on red blood cells in Erythro-Sol environment

Background and Objectives  Routine procedures for extended storage of whole blood (WB) before the preparation of blood components are of interest primarily for logistical reasons. We stored red cell units in either Erythro-Sol 2 (E-Sol 2, test units, 150 ml added) or in saline-adenine-glucose–mannitol (SAG-M) (reference units, 100 ml added) that were prepared after storage of WB at room temperature for 8, 12, 16 or 19 h after blood collection.
Study Design and Methods  Red blood cells were stored for 42 days. We measured pH, glucose, lactate, haemolysis, red blood cell adenosine triphosphate and 2,3-diphosphoglycerate on days 1, 7, 14, 21, 28, 35 and 42.
Results  Haematocrits were significantly lower in E-Sol 2 than in SAG-M due to the higher volume of E-Sol 2 added compared to SAG-M. Significantly reduced levels were found in E-Sol 2 of extracellular pH (throughout storage after 8-h hold and initially after 12-, 16- or 19-h hold), of lactate (initially after 8-h hold and throughout storage after 12-, 16- or 19-h hold), and of haemolysis from day 35 in the 8-h and on day 42 in the 12-h hold group. Sigificantly increased levels of adenosine triphosphate were seen in E-Sol 2 after 8-h hold (from day 14) and after 12-h hold (at days 21, 35 and 42) compared to SAG-M. Significantly higher concentrations of 2,3-diphosphoglycerate were noticed primarily after 8-h hold of WB.
Conclusion  The use of E-Sol 2 as a replacement for SAG-M does not significantly improve in vitro data after extended storage of WB at room temperature before preparation of blood components. However, after 8-h hold in vitro characteristics similar to or better than in fresh blood will be maintained for several weeks in E-Sol 2, a situation that makes E-Sol 2 superior to SAG-M when storage of WB is limited to 8 h. Some improvement was noted after 12-h hold as well.     

The impact of two whole blood inline filters on markers of coagulation, complement and cell activation

BACKGROUND AND OBJECTIVES: There exists a current lack of information about the impact of different inline filters, used for the leucoreduction of whole blood (WB), on the levels of clotting factors and markers of coagulation, complement and cell activation in plasma. Only a few small comparisons of different types of WB inline filters have been published to date. MATERIALS AND METHODS: This study compared two plasma types of 200 units each. Both study groups were derived from WB, inline-filtered and held for 2 h at 20 degrees between donation and filtration. Then, 200 units (Group A) were filtered using a positively charged polyester filter (Baxter RZ2000) and the other 200 units (Group B) were filtered using an uncharged polyester filter (Fresenius). After filtration, both groups were analysed for fibrinogen, factors V and VIII:C (FV and FVIII:C, respectively), immunoglobulin G (IgG), residual leucocytes and platelets, and markers of coagulation, complement and cell activation. Predonation plasma samples from CPDA1-anticoagulated blood were obtained from 100 different individuals and served as controls. RESULTS: WB inline filtration did not influence fibrinogen, FV, FVIII:C or IgG levels. Neither filter induced thrombin or fibrin formation. The charged filter caused substantial complement activation and neutrophil elastase and platelet factor 4 release. In contrast, the plasma filtered through the uncharged filter showed markedly lower levels of C3a-desArg, C5a, neutrophil elastase and platelet factor 4, and moderately reduced levels of prothrombin fragments 1+2 and D-dimer, compared with controls. CONCLUSIONS: Filter type has a significant impact on the quality of plasma derived from WB filtered through inline filtration systems. Some filters produce substantial coagulation and complement activation and cell release, while others appear to reduce the plasma levels of activation markers. The clinical significance of these findings remains to be determined.     

Acquired haemophilia: dynamic whole blood coagulation utilized to guide haemostatic therapy

Acquired haemophilia is a rare bleeding disorder caused by autoimmune antibodies interacting with factor VIII (FVIII) or factor IX. Anticipating a high degree of heterogeneity amongst cases, we recently initiated systematic recording of whole blood (WB) coagulation dynamic profiles using our recently developed thrombelastographic method employing very small amounts of tissue factor for activation. Six newly diagnosed patients with acquired haemophilia A in our University Hospital were investigated with the purpose to characterize the WB clotting phenotypes in each patient, as well as inspecting the ex vivo and in vivo response to supplementation with various haemostatic agents. Our results show a striking heterogeneity in patients WB clotting profiles, each patient having a particular pattern and an individual type of response to bypassing agents. Profiles in some of patients resembled severe haemophilia A, even if there was a measurable residual FVIII:C activity while others were more similar moderate-to-mild haemophilia. In one case the profile was very close to normal. Each patient seemed to respond individually to bypassing agents. WB clotting profiles assisted us in selecting an optimal treatment modality in each case and whenever possible, we compared the clinical effects of the treatment selected with the appearance of the WB clotting pattern. In one patient, the ex vivo response to FVIII looked promising, and a approximately 200 IU kg-1 per 24 h high-dose programme nearly normalized the clotting profile in 2-week time. Our preliminary small series of data should be concluded with caution. However, it seems that WB clotting profile studies at baseline, with ex vivo addition of haemostasis promoting agents, and during treatment may hold the potential to predict the success of treatment.     

全血γ干扰素测定试验在承德地区结核病诊断价值研究

目的探讨全血γ干扰素测定试验(TB-IGRA)在承德地区结核分枝杆菌感染的临床价值。方法选取自2014年5月1日至2015年5月1日间,来中国人民解放军第二六六医院就诊和住院治疗的142例结核病患者、85例非活动性结核患者,均行全血特异性γ干扰素(IFN-γ)含量测定,同时与抗酸染色和血结核抗体检测进行平行比较分析。结果结核病患者体内γ干扰素(IFN-γ)水平显著高于非结核病患者,差异有统计学意义(P0.05);全血γ干扰素测定试验对结核分枝杆菌(MTB)感染者的敏感性为88.73%(126/142),特异性为88.24%(75/85);142例结核病患者中,抗酸染色和血结核抗体的阳性率分别为:41.55%(59/142)和65.49%(93/142),全血γ干扰素测定试验与抗酸染色、结核抗体阳性率差异有统计学意义(P0.01、P0.05)。结论全血γ干扰素测定试验可以快速、有效的诊断结核分枝杆菌感染患者,是一种适合本地区结核病诊断的试验方法。     

Automated processing of whole blood units: operational value and in vitro quality of final blood components

Background

The Community Transfusion Centre in Madrid currently processes whole blood using a conventional procedure (Compomat, Fresenius) followed by automated processing of buffy coats with the OrbiSac system (CaridianBCT). The Atreus 3C system (CaridianBCT) automates the production of red blood cells, plasma and an interim platelet unit from a whole blood unit. Interim platelet unit are pooled to produce a transfusable platelet unit. In this study the Atreus 3C system was evaluated and compared to the routine method with regards to product quality and operational value.

Materials and methods

Over a 5-week period 810 whole blood units were processed using the Atreus 3C system. The attributes of the automated process were compared to those of the routine method by assessing productivity, space, equipment and staffing requirements. The data obtained were evaluated in order to estimate the impact of implementing the Atreus 3C system in the routine setting of the blood centre. Yield and in vitro quality of the final blood components processed with the two systems were evaluated and compared.

Results

The Atreus 3C system enabled higher throughput while requiring less space and employee time by decreasing the amount of equipment and processing time per unit of whole blood processed. Whole blood units processed on the Atreus 3C system gave a higher platelet yield, a similar amount of red blood cells and a smaller volume of plasma.

Discussion

These results support the conclusion that the Atreus 3C system produces blood components meeting quality requirements while providing a high operational efficiency. Implementation of the Atreus 3C system could result in a large organisational improvement.     

No difference of accuracy between capillary and venous blood in rapid whole blood test for diagnosis of Helicobacter pylori infection

The aims of the study were to evaluate the diagnostic accuracy of a rapid whole blood test (one-step BM-Test Helicobacter pylori) in different blood collections and compare it with a quantitative ELISA test (HEL-p TEST II). One hundred four dyspeptic patients were studied, and 53 (51%) were H. pylori-positive as determined by [¹³C] urea breath test. The area under the ROC curve was higher for HEL-pTEST II compared to the one-step BM-Test with either fingerstick or venipuncture (0.972 vs 0.721 and 0.714, P = 0.000). The most appropriate cutoff reading time was 5 min for the one-step BM-Test. The sensitivity, specificity, and accuracy were 52.8%, 88.2%, 70.2% for fingerstick; 62.3%, 80.4%, 71.2% for venipuncture; and 90.6%, 92.2%, 91.3% for HEL-pTEST II. We conclude that there is no difference in diagnostic performance between capillary and venous blood for rapid whole blood test, and extending the reading time beyond the manufacturer's suggestion for the one step BM-Test does not improve its accuracy.     

用微量全血彗星试验观察砷中毒病区人群DNA损伤

目的 将微量全血彗星实验方法及微量血样保存方法在地方性砷中毒病区进行实践，以验证该方法的可行性。方法 采集病区人群末梢微量血样及相应的水样及尿样，观察人群DNA损伤程度与总砷摄入量及尿砷之间的相关关系以及不同人群DNA损伤程度之间的差别。结果 没有发现总砷摄入量及尿砷与DNA损伤程度之间的相关关系，但砷中毒病人的DNA损伤程度明显高于非病人及可疑病人。结论 将微量血样保存方法及微量全血彗星实验用于地方性砷中毒病区的人群监测可行。     

Monokine production by peripheral whole blood in chronic hepatitis C patients treated with interferon

Using our scoring system, we studied the production of monokines (interleukin-1, interleukin-1, tumor necrosis factor-, and interleukin-6) by lipopolysaccharide-stimulated peripheral whole blood in 35 patients with chronic hepatitis C during the interferon-/ therapy. It decreased in 25.7% (9/35, group A), fluctuated in 60.0% (21/35, group B), and increased in 14.3% (5/35, group C). The patients in group A were younger than those in group B (P<0.05). The histological grade of injury was milder in group A than in group B or C. The rate of sustained response was 66.7% (6/9) in group A, 19.0% (4/21) in group B, and 40.0% (2/5) in group C (P=0.0184, group A versus group B). In summary, monokine production by peripheral whole blood varied during interferon therapy for chronic hepatitis C patients. No significant change was noted in 60% of the patients. However, patients with decreased monokine production were younger, with a mild histological grade, and likely to respond to the interferon therapy.     

The effect of pre-donation hypotension on whole blood donor adverse reactions: a systematic review

Blood services are reliant upon healthy blood donors to provide a safe and adequate supply of blood products. Inappropriate variables contained within blood donor exclusion criteria can defer potentially appropriate donors. The aim of this systematic review was to examine the effect of low pre-donation blood pressure, as compared with normal blood pressure, on adverse events in allogeneic whole blood donors. A systematic review was performed using highly sensitive search strategies within five databases (Cochrane Central Register of Controlled Trials, CINAHL, Embase, MEDLINE, and Web of Science) from inception date until April 12, 2013. Out of 8305 records, 10 observational studies were identified that addressed the question. Five of these studies (with a combined total of 1,482,020 donations and 2903 donors) included either a statistical analysis or an appropriate study design that controlled for possible confounding factors. Based on the currently available evidence, hypotension has not been shown to be an independent predictive factor for donor complications. However, the overall quality of evidence was rather limited and rated ‘low,’ using the GRADE approach. In conclusion there is currently no evidence that hypotensive blood donors have a greater risk for donor adverse events compared with their normotensive counterparts.     

碘营养充足地区新生儿脐带血TSH切点值的研究

目的 探讨新生儿脐带血促甲状腺激素（TSH）测定在临床应用的可行性和在碘营养充足的情况下新生儿脐带血TSH的切点值。方法 选择采取食盐加碘干预措施后当地居民碘营养充足地区的孕妇作为的研究对象,观察指标有孕妇尿碘、新生儿脐带血TSH和新生儿足跟血TSH水平及相关因素。结果 ①碘营养充足地区碘营养充足的孕妇所分娩的新生儿脐带血TSH的切点值为8.3mU/L,高于国际标准5mU/L;②新生儿脐带血TSH与足跟血TSH呈正相关关系。结论 新生儿脐带血TSH测定用于碘缺乏病的监测是可行的,有必要建立适合我国的新生儿脐带血TSH切点值。     

Effects of triflusal and acetylsalicylic acid on platelet aggregation in whole blood of diabetic patients

A study was made on the inhibitory effect of triflusal (600 mg/d X 15) and acetylsalicylic acid (ASA, 400 mg/d X 15) on platelet aggregation in whole blood (WB) and platelet-rich plasma (PRP) induced by ADP (2.5 mumol/l), adrenaline (50 mumol/l), collagen (1 microgram/ml) and arachidonic acid (0.8 mmol/l), in 30 insulin-dependent diabetic patients without vascular complications. Determination was also made of the serum levels of thromboxane B2 (TxB2) and of the plasma levels of 6-keto-PGF1-alpha and of beta-thromboglobulin (B-TG). Both drugs exhibited higher inhibitory effects in WB than in PRP. In WB, a significant difference between triflusal and ASA was observed against ADP-induced aggregation (67% and 46% inhibition respectively, p less than 0.01). Both drugs strongly inhibit the formation of TxB2 in serum (85% and 99%, respectively). Triflusal does not significantly change the plasma levels of 6-keto-PGF1-alpha; ASA, by contrast, causes reduction of over 95% in those plasma levels. The plasma levels of B-TG were not modified by either of the drugs.     

Mild and moderate hypothermia increases platelet aggregation induced by various agonists: a whole blood in vitro study

The mechanisms causing temperature-dependent bleeding, especially in hypothermic patients, warrant clarification. Therefore the aim of this study was to investigate platelet aggregation at the clinically important temperature range of 30–34°C. After obtaining informed consent citrated whole blood was drawn from 12 healthy adult male volunteers, who had not taken any medication in the previous 14 days. After venipuncture blood samples were incubated at 37°C until platelet testing. Platelet aggregation was performed in whole blood using the impedance aggregometer Multiplate® at five different test temperatures between 30°C and 34°C. Aggregation responses at 37°C served as controls. At temperatures of mild and moderate hypothermia (30–34°C), overall platelet aggregation was increased compared to 37°C. Increases were recorded in response to collagen, thrombin receptor activating peptide and ristocetin between 31°C and 34°C and in response to adenosine diphosphate between 30°C and 34°C. Overall platelet aggregation is increased at mild and moderate hypothermia down to 30°C. These results indicate that bleeding complications reported in mildly hypothermic patients are not due to hypothermia-induced platelet inhibition. The pathomechanism of the overall increased platelet aggregation between 30°C and 34°C requires further detailed study.