scholarly journals Validation of Blood Collection Procedures for the Determination of Circulating Vascular Endothelial Growth Factor (VEGF) in Different Blood Compartments

2001 ◽  
Vol 16 (2) ◽  
pp. 87-96 ◽  
Author(s):  
R. Dittadi ◽  
S. Meo ◽  
F. Fabris ◽  
G. Gasparini ◽  
D. Contri ◽  
...  
Keyword(s):  
Platelet Count ◽  
Whole Blood ◽  
Relevant Information ◽  
Sampling Procedure ◽  
Systematic Evaluation ◽  
Blood Collection ◽  
Sample Collection ◽  
Analytical Phase

Aims of the study. Studies on circulating VEGF have reported mixed results, possibly due to a lack of standardization of the pre-analytical phase. The aim of our investigation was to standardize the sampling procedure for the determination of VEGF in different blood fractions. Basic procedures. We evaluated various clotting times for obtaining serum in 30 subjects, as well as different procedures for the preparation of plasma Edinburgh anticoagulant mixture (EDTA, PGE1, theophylline) and CTAD. VEGF was also assayed in lysed whole blood. In vitro platelet activation was monitored by measuring the levels of PF4. VEGF and PF4 were measured using commercially available enzyme-linked immunoassays. Main findings. Clotting time increased the release of VEGF, which reached a plateau between 2 and 4 hours. The percent increase of VEGF at 2 hours ranged from 118% to 4515% (median 327%) compared to samples centrifuged within 10 min from withdrawal. VEGF was not different and PF4 was very low or undetectable in Edinburgh plasma and CTAD plasma, while it was significantly higher in sodium citrate plasma. VEGF in CTAD plasma was not correlated with platelet count or leukocytes. Serum VEGF did not correlate with the leukocyte number, but it correlated significantly with the platelet count. Principal conclusions. The procedures for sample collection described above are highly standardized and easy to perform in a routine setting. We therefore suggest systematic evaluation of VEGF in CTAD plasma, in serum (clotting for 2 hours at room temperature) and in whole blood, until prospective controlled clinical studies will have clarified in which blood compartment(s) VEGF provides clinically relevant information.

scholarly journals Importance of Preanalytical Factors in Measuring Cr and Co Levels in Human Whole Blood: Contamination Control, Proper Sample Collection and Long-Term Storage Stability

2020 ◽  
Author(s):  
Yuliya L Sommer ◽  
Cynthia D Ward ◽  
Joaudimir Castro Georgi ◽  
Po-Yung Cheng ◽  
Robert L Jones
Keyword(s):  
Stainless Steel ◽  
Whole Blood ◽  
Consumer Products ◽  
Blood Collection ◽  
Sample Collection ◽  
Blood Samples ◽  
Contamination Control ◽  
Human Whole Blood

Abstract A number of errors with potentially significant consequences may be introduced at various points in the analytical process, which result in skewed, erroneous analytical results. Precautionary procedures such as contamination control, following established sample collection protocols, and having a complete understanding of the long-term stability of the elements of interest can minimize or eliminate these errors. Contamination control is critical in the quantification of Cr and Co in human whole blood. Cr and Co levels in most biological samples are low, but these elements occur naturally in the environment and are often found in commercial and consumer products, which increases the risk of contamination. In this paper, we demonstrated that lot screening process in which we pre-screen a sub-set of manufactured lots used in collecting, analyzing and storing blood samples is a critical step in controlling Cr and Co contamination. Stainless steel needles are often utilized in blood collection but are considered as a potential source of introducing metal contamination to the patient sample. We conducted two studies to determine if there is a possibility of Cr or Co leaching into the human whole blood from the needles during blood collection. We analyzed blood collected from 100 donors and blood collected in vitro in the laboratory from designated vessel containing spiked blood with higher levels of Cr and Co. Two blood tubes were consecutively collected through one needle. In both studies, Cr and Co concentration levels in the two consecutively collected tubes were compared. Based on the results from donor and in vitro blood collection studies, we concluded that there was no Cr and Co leaching from the limited sets of stainless steel needles used in these studies. Furthermore, we demonstrated that Cr and Co human whole blood samples are stable for 1 year stored at temperatures of −70, −20 and 4°C and 6 months at room temperature.

A Simple and Rapid Method to Determine GPIIb/IIIa-Receptor Inhibitor Activity in Whole Blood

1999 ◽  
Vol 19 (03) ◽  
pp. 134-138
Author(s):  
Gitta Kühnel ◽  
A. C. Matzdorff
Keyword(s):  
Flow Cytometry ◽  
Platelet Count ◽  
Blood Sample ◽  
Platelet Activation ◽  
Whole Blood ◽  
Receptor Occupancy ◽  
Aggregate Formation ◽  
Inhibitor Activity ◽  
Receptor Inhibitor

SummaryWe studied the effect of GPIIb/IIIa-inhibitors on platelet activation with flow cytometry in vitro. Citrated whole blood was incubated with increasing concentrations of three different GPIIb/IIIa-inhibitors (c7E3, DMP728, XJ757), then thrombin or ADP were added and after 1 min the sample was fixed. Samples without c7E3 but with 0.1 U/ml thrombin had a decrease in platelet count. Samples with increasing concentrations of c7E3 had a lesser or no decrease in platelet count. The two other inhibitors (DMP 725, XJ757) gave similar results. GPIIb/IIIa-inhibitors prevent aggregate formation and more single platelets remain in the blood sample. The agonist-induced decrease in platelet count correlates closely with the concentration of the GPIIb/IIIa inhibitor and receptor occupancy. This correlation may be used as a simple measure for inhibitor activity in whole blood.

Determinants of platelet conjugate formation with polymorphonuclear leukocytes or monocytes in whole blood

2007 ◽  
Vol 98 (12) ◽  
pp. 1276-1284 ◽  
Author(s):  
Simona Costanzo ◽  
Branislav Vohnout ◽  
Licia Iacoviello ◽  
Giovanni de Gaetano ◽  
Benedetta Izzi ◽  
...  
Keyword(s):  
Platelet Count ◽  
Whole Blood ◽  
Polymorphonuclear Leukocytes ◽  
Large Population ◽  
Mixed Cell ◽  
Cd11b Expression ◽  
Glucose Levels ◽  
Primary Platelet ◽  
Collagen Stimulation

SummaryFollowing preliminary in-vitro experiments, platelet-leukocyte conjugates and their determinants were evaluated in citrated whole blood from 349 subjects (209 women, age 16–92 years) randomly recruited from the general population. Platelet activation by ADP/collagen but not leukocyte stimulation by fMLP or LTB4 resulted in formation of platelet conjugates with PMN or monocytes. In the population study, mixed cell conjugates, platelet P-selectin and leukocyte CD11b were measured by flow cytometry both at baseline and after in-vitro stimulation with ADP/collagen. The latter significantly increased platelet conjugates with either PMN or monocytes, platelet P-selectin and leukocyte CD11b expression. Platelet count significantly correlated with platelet-PMN, platelet-monocyte conjugates and P-selectin both at baseline and upon stimulation. In all conditions, both conjugate levels correlated with each other, when adjusted for gender, age and platelet count. Age correlated with platelet-PMN conjugate numbers in basal and stimulated conditions and with basal P-selectin. ADP/collagen stimulation resulted in higher P-selectin and conjugates values in women. Among risk factors, a significant correlation was found between conjugate and glucose levels. In conclusion, the presence and formation in whole blood from a large population of plateletleukocyte conjugates reflects primary platelet – but not leukocyte – activation and varies with gender, age, platelet count and blood glucose.

Determination of the Effect of In-vitro Time, Temperature and Tourniquet Use on Whole Blood Venous Point-of-care Lactate Concentrations

2007 ◽  
Vol 14 (5 Supplement 1) ◽  
pp. S185-S185
Author(s):  
M. Leonard ◽  
A. Jones ◽  
J. Hernandez-Nino ◽  
J. Kline
Keyword(s):  
Whole Blood ◽  
Point Of Care

Abstract 571: A Novel High-Throughput Whole Blood Immuno-Counting Assay is Useful in the Assessment of Platelet Responses to Antiplatelet Therapy

2014 ◽  
Vol 34 (suppl_1) ◽  
Author(s):  
Marie Lordkipanidze ◽  
Natalia Dovlatova ◽  
Mohammad Algahtani ◽  
Martina H Lundberg ◽  
Timothy D Warner ◽  
...  
Keyword(s):  
Platelet Count ◽  
Antiplatelet Therapy ◽  
Platelet Function ◽  
Whole Blood ◽  
Dual Antiplatelet Therapy ◽  
Whole Blood Assay ◽  
Blood Assay ◽  
Treated Sample ◽  
Function Testing

Background: The current gold standard in platelet function testing, light transmission aggregometry, is time- and labor-intensive, and uses platelet-rich plasma which makes it sub-optimal for high throughput testing. In order to reduce blood manipulation prior to platelet function testing and to study multiple platelet activation pathways simultaneously, we have developed a 96-well plate-based assay carried out in whole blood, where aggregation is measured as a decrease in the number of fluorescently-labeled single platelets by flow cytometry. Aim: To investigate whether a 96-well plate-based whole blood assay can be used to assess platelet function. Methods: Platelet function in response to 5 concentrations of lyophilized arachidonic acid (AA), ADP, collagen, epinephrine, TRAP, U46619, and ristocetin, was assessed in healthy volunteers (n=20) to establish normal ranges. The effect of antiplatelet drugs was assessed in vitro by incubation with aspirin (100 μM), cangrelor (1 μM) or both (n=20), and in patients on dual antiplatelet therapy (n=20). After addition of 40 μl of whole blood per well, the plate was shaken for 5 min at 1000 rpm at 37°C; a fixative solution (Platelet Solutions, Nottingham) was applied to stop platelet aggregation and allow analysis in a central laboratory. Fixed whole blood samples (stable for up to 9 days) were labeled with FITC-conjugated CD42a and assessed by flow cytometry. Aggregation was calculated as (Platelet count in vehicle-treated sample - Platelet count in agonist-stimulated sample) / Platelet count in vehicle-treated sample x 100. Results: Dose-response curves were readily assessable for all agonists and intra-individual variability was minimal in healthy volunteers (CV<10%). In vitro addition of aspirin alone resulted in inhibition of AA- and collagen-induced aggregation, whereas cangrelor induced a shift in dose-response to most agonists in addition to profound inhibition of ADP responses. In patients on dual antiplatelet therapy, the pattern of response was consistent with the results obtained with in vitro agents. Conclusions: A 96-well plate-based whole blood assay with a minimal blood volume requirement (<2 ml) could be used to provide a global portrait of platelet responses to antiplatelet agents.

Generation of Kinins during Preparation and Storage of Leukoreduced Platelet Concentrates.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 947-947
Author(s):  
Marie Eve Moreau ◽  
Louis Thibault ◽  
Anik Désormeaux ◽  
François Marceau ◽  
Marie-Joëlle de Grandmont ◽  
...  
Keyword(s):  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Storage System ◽  
Reference Population ◽  
Storage Conditions ◽  
Blood Collection ◽  
Converting Enzyme Inhibitors ◽  
Pharmacological Characterization ◽  
And Storage

Abstract BACKGROUND. Severe hypotensive reactions can occur following transfusion of blood components, particularly with platelets concentrates (PCs) leukoreduced with negatively- charged filters. Bradykinin (BK)-related peptides were proposed as a possible cause of this side-effect. This study evaluated the effects of leukoreduction and storage conditions on the levels of two kinins (BK and des-Arg9-BK) in PCs. METHODS. Whole blood donations (n=35) were processed using current PRP (platelet-rich plasma) procedure to prepare leukoreduced and unfiltered components by Leukotrap® RC-PL Whole Blood Collection, Filtration and Storage System. PCs and plasma were stored for 7 days at 20–24°C with agitation. Levels of BK and des-Arg9-BK were measured by specific enzyme immunoassays and HPLC at day 0, 2, 5 and 7 of storage. Mechanisms potentially responsible for accumulation of BK and des-Arg9-BK were studied. RESULTS. On day 0, kinins were measured in significantly higher concentrations in leukoreduced (BK: 101 ± 157 pg/mL; des-Arg9-BK: 194 ± 191 pg/mL) vs unfiltered PCs (BK: 71 ± 121 pg/mL; des-Arg9-BK: 98 ± 114 pg/mL). During storage, both kinins peaked on day 5, with concentrations higher than 1 ng/mL in 22% of leukoreduced as well as unfiltered PCs. Physicochemical and pharmacological characterization of immunoreactive kinins confirmed their nature. In vitro activation of the contact system of the corresponding platelet-poor plasma (PPP) showed that a high kinin concentration on day 5 of the storage corresponded to a low kinin-forming capacity of plasma. On day 7, BK was no longer elevated presumably due to its degradation and the depletion of kinin-forming capacity of the plasma in stored PCs. The activity of metallopeptidases that metabolize BK-related peptides in plasma from PCs were at levels similar to those recorded in the plasma of a normal reference population and were unaffected by storage. CONCLUSIONS. Our results indicate that filtration and storage conditions of PCs contribute to generation of pharmacologically relevant bradykinin levels that might pose a risk in susceptible patients. The clinical relevance of such high concentrations of kinins in PCs remains to be established but could potentially be significant especially in patients treated with angiotensin I-converting enzyme-inhibitors that affect the pathway of BK degradation.

Order of blood draw: Opinion Paper by the European Federation for Clinical Chemistry and Laboratory Medicine (EFLM) Working Group for the Preanalytical Phase (WG-PRE)

2017 ◽  
Vol 55 (1) ◽  
pp. 27-31 ◽  
Author(s):  
Michael Cornes ◽  
Edmée van Dongen-Lases ◽  
Kjell Grankvist ◽  
Mercedes Ibarz ◽  
Gunn Kristensen ◽  
...  
Keyword(s):  
Working Group ◽  
Clinical Chemistry ◽  
Venous Blood ◽  
Laboratory Medicine ◽  
European Federation ◽  
Blood Collection ◽  
Sample Collection ◽  
Blood Draw ◽  
Preanalytical Phase ◽  
Analytical Phase

AbstractIt has been well reported over recent years that most errors within the total testing process occur in the pre-analytical phase (46%–68.2%), an area that is usually outside of the direct control of the laboratory and which includes sample collection (phlebotomy). National and international (WHO, CLSI) guidelines recommend that the order of draw of blood during phlebotomy should be blood culture/sterile tubes, then plain tubes/gel tubes, then tubes containing additives. This prevents contamination of sample tubes with additives from previous tubes that could cause erroneous results. There have been a number of studies recently looking at whether order of draw remains a problem with modern phlebotomy techniques and materials, or it is an outdated practice followed simply because of historical reasons. In the following article, the European Federation of Clinical Chemistry and Laboratory Medicine Working Group for the Preanalytical Phase (EFLM WG-PRE) provides an overview and summary of the literature with regards to order of draw in venous blood collection. Given the evidence presented in this article, the EFLM WG-PRE herein concludes that a significant frequency of sample contamination does occur if order of draw is not followed during blood collection and when performing venipuncture under less than ideal circumstances, thus putting patient safety at risk. Moreover, given that order of draw is not difficult to follow and knowing that ideal phlebotomy conditions and protocols are not always followed or possible, EFLM WG-PRE supports the continued recommendation of ensuring a correct order of draw for venous blood collection.

Über den Nachweis der Plasminogenverarmung in retrahierten Vollblutgerinnseln und über deren Bedeutung für die Lysierbarkeit mit Streptokinase

1966 ◽  
Vol 15 (03/04) ◽  
pp. 570-590 ◽  
Author(s):  
R Gottlob ◽  
G Blümel
Keyword(s):  
Water Content ◽  
Calcium Ions ◽  
Whole Blood ◽  
Immunological Method ◽  
Blood Clots

Summary1. Blood clots are lysed by SK before the beginning of retraction. Retracted clots are only very little lysed.2. Recalcified citrated blood is easily lysed since the excess of added calcium ions inhibits the retraction.3. The determination of the water content of the clots gives information on their retraction state.4. The determination of water content made it possible to prove that thrombus retraction takes place in vivo. Already one day after thrombosis the water content is clearly lower than in fresh clots. Later the water content increases again slightly.5. Clots formed in vitro and in vivo are lysed by SK after retraction if they come into contact with fibrin or fibrinogen containing plasminogen.6. Heated fibrin plates or heated fibrin tubes both with SK, casein degradation and an immunological method allowed to prove that whole blood clots loose their activable plasminogen during retraction. One assumes that plasminogen is expressed during retraction.7. The possibility of a therapeutical fibrinolysis with SK is due to the fact that SK can penetrate into the clot and the SK containing thrombus continuously comes in contact with circulating blood rich in plasminogen. The importance of these results for thrombolysis with SK is discussed.

Sign in / Sign up

Export Citation Format

Share Document