Ability of whole blood aggregometer to detect platelet hyperaggregability

In this study the reliability of platelet aggregation in whole blood (WB) was investigated in clinical conditions associated with thromboembolic complications. Spontaneous (SPA) and collagen-induced platelet aggregation were evaluated both in whole and diluted blood by the impedance method and in platelet-rich plasma (PRP) by the Born aggregometer in 18 healthy subjects, 15 patients with ischemic heart disease (IHD), and 15 patients with insulin-independent diabetes. SPA occurred more often in WB than in PRP, and in WB the occurrence of SPA was significantly more frequent in the patient groups (4 of 15 patients with IHD and 6 of 15 diabetic patients) than in the controls (1 of 18). WB aggregation induced by collagen was significantly higher in patients with IHD and in diabetic patients than in controls (P less than 0.01), whereas diluted WB and PRP aggregation were not statistically different from controls either in patients with IHD or in diabetic patients. WB aggregation values were found to be related, although not very closely, to megathrombocyte count (r = 0.31, P less than 0.05) whereas not at all to platelet count or hematocrit. No relationship was observed between WB aggregation and disease severity (angiographic lesions and number of ischemic attacks) in patients with IHD and between WB aggregation and HbAlc values in diabetic patients.     

Sub-ambient measurement of platelet function: the requirements for modification of a standard platelet aggregometer

This methodological report documents the electro-mechanical modifications that were applied to a commercially available aggregometer, in order to achieve direct platelet aggregation measurement at sub-ambient temperatures. This development provides a facility, not previously available, for examining certain aspects of platelet physiology at any desired temperature. This innovation was primarily developed for investigating optimum storage parameters for platelets, as they apply to clinical blood transfusion practice. However, a wider scope of application is envisaged, including its use as both a general research tool, and eventually as an additional component of clinical measurement in the routine coagulation laboratory. Some illustrative data is briefly provided.     

Ristocetin-induced platelet aggregate formation and adherence to the probe of an impedance aggregometer

The examination of the interaction between ristocetin and platelets generally is performed in an optical aggregometer where aggregate formation is detected by a change in turbidity. Ristocetin also will induce conductivity changes in an impedance aggregometer with either fresh whole blood or fresh platelet-rich plasma due to aggregate attachment to the electrical probe. Formaldehyde-fixed platelets, however, do not attach, and although fresh platelets agglutinate at extremes of pH, probe adhesion does not occur unless the pH is within a certain range. Incubation of platelet preparations with neuraminidase, colchicine, or cytochalasin-b affects neither aggregate formation nor probe adherence. Incubation of platelet preparations with a fibronectin tetrapeptide inhibitor eliminates aggregate formation in response to collagen but is without effect on ristocetin-induced attachment of aggregates to the impedance probe. It is suggested that the ability of platelet aggregates to attach to an impedance probe in response to ristocetin is dependent on additional aspects of the functional integrity of platelet preparations and that these changes required for probe adherence may have in vivo significance.     

RBC aggregometer head as a warning monitor of flow disturbance in extracorporeal systems

A simple photoelectric apparatus is described, to monitor the flow condition of whole blood in an extracorporeal system. It can raise an immediate alarm on disturbance of flow. The apparatus comprises an RBC aggregometer head, described previously, for measuring the rate of RBC aggregation in whole blood. Its construction was modified for easy attachability and detachability without causing tube damage. In practice, the apparatus was applied by hooking it to the tube of the extracorporeal system in animals and in a clinical case of blood dialysis in a patient with renal failure, in which stoppage of blood flow elicited a dramatic change in the baseline record of light transmission of the mobile blood. The signal of the apparatus was fed to an alarm system via a voltage comparator switch. The apparatus is inexpensive, solid and durable, easy to operate by untrained personnel, and has excellent stability and reproducibility.     

The platelet function profile. A new format for recording platelet function tests.

A platelet function profile that provides a concise record of platelet function tests has been developed. This format can be easily modified to meet local requirements or to include hematologic tests unique to each laboratory.     

A new approach to evaluate platelet function in hemodialysis patients--saponin susceptibility of the platelet

Saponin susceptibility of the platelet was studied in 10 healthy men (ages 24 to 39) and 31 male hemodialysis (HD) patients (ages 24 to 49) in order to assess platelet abnormalities in HD patients. Platelet-rich plasma (PRP) was added to 50 ml of saline containing 5 mM phosphate buffer (pH 7.0) and 0.3% saponin. Platelet mean particle volume (MPV) was observed continuously for 200 seconds by a continuous mean particle analyzer model CC-108 (Sysmex, Kobe, Japan). Initial MPV (Vo, fl) was 7.12 +/- 0.89 and 7.68 +/- 0.56 respectively for the control and HD patients (p less than 0.05), and maximum MPV (V1) was 9.80 +/- 1.06 and 9.78 +/- 0.83, respectively (n.s.). The time of start of swelling (seconds) was 51.2 +/- 14.6 and 63.4 +/- 15.6 (p less than 0.05) and the time of V1 was 110.4 +/- 15.3 and 112.3 +/- 16.1 (n.s.), respectively. The expansion rates (V1/Vo) were 1.38 +/- 0.05 and 1.27 +/- 0.06, respectively (p less than 0.01). The low expansion rate means that the platelet is less responsive to saponin in HD patients, and this may be related to the cholesterol concentration in the platelet membrane and membrane flexibility. This new method offers a useful way of obtaining information about the platelet.     

A laboratory study of spontaneous platelet aggregation

During studies on platelet aggregation using the EEL platelet aggregation meter, 8% of the individuals tested were found to have platelets which aggregated spontaneously when citrated, platelet-rich plasma was stirred at 37 degrees C. The EEL aggregation meter differs from other machines in that it incorporates a vertical stirrer which subjects platelets to greater mechanical force. When using this machine it is suggested that spontaneous platelet aggregation is related to increased mechanical fragility of the platelets and low levels of plasma ADP-inhibitor.     

A stochastic model of pulmonary platelet production

Normally, cells reproduce by mitosis. In mammals, a system has evolved that is unique to cell biology. The circulating blood cells called platelets are produced from parent cells called megakaryocytes, but the mechanism of platelet production is not mitosis. At present, both the site and mechanism of production are still debated. This article describes a production mechanism based on a sequence of random binary divisions of the megakaryocyte cytoplasm as it traverses the pulmonary microcirculation. Using the measured megakaryocyte cytoplasmic volumes circulating in the blood of three different mammals, rat, rabbit, and man, a computer simulation of this process is developed. The simulation depends on two separate stochastic processes. The first involves the probability that a division of a cytoplasmic particle of a specified size will occur. The second relates to the relative sizes of the two particles produced by the binary division. These two processes are controlled by three parameters only: a lower threshold L on the platelet volume, below which the probability of a binary division is zero; a parameter lambda which defines the probability of division for volumes larger than this lower threshold; and the standard deviation S of the Gaussian distribution of possible volumes created by the binary division. A minimization scheme is used to establish those values of L, lambda, and S which provide the best fit to the experimental data obtained from the different mammals.     

Measuring platelet aggregation with microplate reader. A new technical approach to platelet aggregation studies

Platelet aggregation measurements were done with the use of a commercially available microtiter plate reader with specific modification of the mode of agitation of the samples. Satisfactory aggregation curves were obtained with use of an external horizontal agitator, with an amplitude of 1.3 mm and minimum frequency of 1,360 cycles/minute. With the use of the 96 available wells in the microtiter plates, all test and control platelet samples, with replicates, were observed simultaneously and the output data obtained within 10-15 minutes. The technique was validated by demonstrating the similarity of dose-response curves, obtained with a standard aggregometer and with the microtiter technique, of platelets stimulated by adenosine diphosphate, thrombin, and arachidonic acid.     

A new flow cell for platelet adhesion studies

A flow cell has been designed and tested for the purpose of exposing platelets to various substrates. The design adopts the principle of flow relaminarization by acceleration to dissipate secondary fluid motions and turbulence so that platelet diffusion will be controlled by Brownian diffusion coefficients. To assess the effectiveness of this flow mechanism two critical tests were undertaken. First, hot film anemometry signals were obtained to observe visually the local fluid velocity in the flow cell. Second, in vitro platelet adhesion results were obtained by exposing glass and silane coated glass to platelet suspensions. Surface platelet concentrations for varying exposure times were compared to a theoretical model which is based on a laminar diffusion model. Both tests confirm that hydrodynamically the flow cell behaves in a manner which is consistent with that of a fully developed, laminar flow with a constant diffusion coefficient.     

A note on platelet adhesiveness in ischaemic heart disease

Platelet adhesiveness was measured in vitro in a group of patients with ischaemic heart disease and in a group of matched healthy subjects using either 1% EDTA or 3.8% sodium citrate as anticoagulant. When sodium citrate was used platelet adhesiveness was significantly greater in the group with ischaemic heart disease. However, no difference was found when EDTA was used.     

A case of pseudothrombocytopenia due to platelet cold agglutinins

We report a case of pseudothrombocytopenia due to platelet cold agglutinins. Platelet counts were decreased in blood drawn in a tube without anti-coagulant just after withdrawal as well as in blood drawn in a tube with anti-coagulant, such as EDTA-2K, MgSO4, citrate or heparin. In our case, platelet aggregates were noted on blood-smear made from blood samples obtained with and without anti-coagulant. RBC and WBC counts were within the normal range. Platelet aggregates mainly consisted of 2-5 platelets. Patient plasma agglutinated normal platelets at a temperature below 10 degrees C. Immunoglobulin class was determined as IgM by flow cytometry.