scholarly journals EVALUATION OF CENTRIFUGING REGIMES FOR THE PURPOSE OF OPTIMIZING THE PLATELET RICH PLASMA HARVESTING PROTOCOL

2019 ◽  
Vol 2 ◽  
pp. 11-18
Author(s):  
Sergiy Chetverikov ◽  
Dmitro Atanasov
Keyword(s):  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Maximum Level ◽  
Calcium Chloride Solution ◽  
Optimal Conditions ◽  
Time Intervals ◽  
Basic Principles ◽  
Harvesting Efficiency ◽  
Centrifugation Protocol

Aim: Based on the classical principles, to determine the optimal conditions for centrifugation, PRP harvesing (platelet-rich plasma). To conduct a quantitative assessment of the substrate obtained under different conditions of centrifugation. Materials and methods. Based on the basic principles of obtaining platelet-rich plasma (PRP) by centrifuging in containers with an anticoagulant followed by phase separation to obtain the final substrate, the efficiency of the technique under the conditions of single and double centrifugation as well as under different conditions of acceleration and centrifugation was evaluated. Blood for follow-up was collected from 20 healthy volunteers (11 men, 9 women) average 25.3±4.1 in syringes of LuerLock design with ACD-A anticoagulant solution, and centrifuged. Centrifugation was carried out under controlled conditions using a centrifuge with rotating bowls of the rotor. Centrifugation was performed at an acceleration of 100-400g in time intervals up to 20 minutes. Activation of the substrate was performed with calcium chloride solution. Quantitative evaluation of platelets of whole blood and the final substrate of PRP was carried out with a semi-automatic analyzer. Results. The obtained results demonstrate the maximum level of harvesting efficiency when performing double centrifugation in the 150g×15 min+250g×10 min mode. Subject to this centrifugation protocol, it is possible to obtain a substrate that complies with the standardized requirements for PRP. The maximum level of an increase in the number of platelets in the substrate in comparison with whole blood is determined at the level of ×4.36 with concentration (volume reduction) x5 in comparison with the volume of whole blood. Conclusions. This study demonstrated the advantage of double centrifuging modes over single modes. According to the results of the study, it was possible to determine the conditions for an optimal double-centrifugation mode (acceleration and duration), which allows us to achieve the most efficient concentration of the substrate.

scholarly journals Study of a Two-Step Centrifugation Protocol for Concentrating Cells and Growth Factors in Bovine Platelet-Rich Plasma

2017 ◽  
Vol 2017 ◽  
pp. 1-8
Author(s):  
Claudia M. Gutiérrez ◽  
Catalina López ◽  
Carlos E. Giraldo ◽  
Jorge U. Carmona
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Whole Blood ◽  
Transforming Growth Factor ◽  
Platelet Rich Plasma ◽  
Distribution Width ◽  
Lack Of Information ◽  
Growth Factor Release ◽  
Pearson Correlations ◽  
Centrifugation Protocol

There is a lack of information about the methods used for bovine platelet-rich plasma (PRP)/platelet-rich gel (PRG) procurement, including information on platelet (PLT), white blood cell (WBC) in PRP, and growth factor release from PRG supernatants. The aims of this study were to compare and to correlate the PLT, WBC, transforming growth factor beta-1 (TGF-β1), and platelet-derived growth factor BB (PDGF-BB) concentrations in bovine whole blood, plasma, and four PRP layers and their respective PRG supernatants: A and B (obtained by a single centrifugation tube method at 720g/5 min) and C and D (obtained by a double centrifugation tube method, by using two centrifugation episodes at 720g/5 min). PLT and WBC counts were significantly higher in PRP-C, followed by whole blood, PRP-A, PRP-B, and PRP-D. TGF-β1 concentrations were significantly higher in PRG-B supernatants and its correspondent PRP-B lysate when compared to the other PRG supernatants and plasma. Supernatants from PRG-A, PRG-B, and PRG-D had equivalent TGF-β1 concentrations. PDGF-BB concentrations were not statistically different between the hemoderivatives. Significant Pearson correlations were noted between PLT counts and WBC counts (0.8) and between PLT counts and PLT distribution width (0.6). Further studies should be performed to assess the potential clinical applications of these PRPs.

Platelet-Rich Plasma Compared With Other Common Injection Therapies in the Treatment of Chronic Lateral Epicondylitis

2016 ◽  
Vol 25 (1) ◽  
pp. 77-82 ◽  
Author(s):  
Tristan Rodik ◽  
Brendon McDermott
Keyword(s):  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Corticosteroid Injection ◽  
Lateral Epicondylitis ◽  
Clinical Scenario ◽  
Clinical Question ◽  
Bottom Line ◽  
And Function ◽  
Corticosteroid Injections

Clinical Scenario:Lateral epicondylitis (LE) is a relatively common pathology capable of producing chronic debilitation in a variety of patients. A newer treatment for orthopedic conditions is platelet-rich plasma (PrP) local injection.Focused Clinical Question:Is PrP a more appropriate injection therapy for LE than other common injections such as corticosteroid or whole blood?Summary of Key Findings:Four studies were included: 1 randomized controlled trial (RCT), 2 double-blind RCTs, and 1 cohort study. Two studies involved comparisons of PrP injection to corticosteroid injection. One of the studies involved a 2-y follow-up while another involved a 1-y follow-up. Another study involved the comparison of PrP injection with whole-blood injection with a 6-mo follow-up. The final study included a PrP-injection group and control group. The 2 studies involving PrP vs corticosteroid injections with 2-y and 1-y follow-ups both favored PrP over corticosteroid injection in terms of pain reduction and function increases. The third study favored PrP injections over whole-blood injections at 6 mo regarding pain reduction. All studies demonstrated significant improvements with PrP over comparison injections or no injection.Clinical Bottom Line:PrP injections provide more favorable pain and function outcomes than whole blood and corticosteroid injections for 1–2 y after injection.Strength of Recommendation:Consistent findings from RCTs suggest level 1b evidence in support of PrP injection as a treatment for LE.

scholarly journals Method to obtain platelet-rich plasma from rabbits (Oryctolagus cuniculus )

2016 ◽  
Vol 36 (1) ◽  
pp. 39-44 ◽  
Author(s):  
Josiane M. Pazzini ◽  
Andrigo B. De Nardi ◽  
Rafael R. Huppes ◽  
Ana P. Gering ◽  
Marília G.P.A. Ferreira ◽  
...  
Keyword(s):  
Calcium Gluconate ◽  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Preparation Time ◽  
Final Sample ◽  
Platelet Concentration ◽  
Initial Blood ◽  
Prp Gel ◽  
Blood Centrifugation ◽  
Centrifugation Protocol

Abstract: Platelet-rich plasma (PRP) is a product easy and inxpesnsive, and stands out to for its growth factors in tissue repair. To obtain PRP, centrifugation of whole blood is made with specific time and gravitational forces. Thus, the present work aimed to study a method of double centrifugation to obtain PRP in order to evaluate the effective increase of platelet concentration in the final product, the preparation of PRP gel, and to optimize preparation time of the final sample. Fifteen female White New Zealand rabbits underwent blood sampling for the preparation of PRP. Samples were separated in two sterile tubes containing sodium citrate. Tubes were submitted to the double centrifugation protocol, with lid closed and 1600 revolutions per minute (rpm) for 10 minutes, resulting in the separation of red blood cells, plasma with platelets and leucocytes. After were opened and plasma was pipetted and transferred into another sterile tube. Plasma was centrifuged again at 2000rpm for 10 minutes; as a result it was split into two parts: on the top, consisting of platelet-poor plasma (PPP) and at the bottom of the platelet button. Part of the PPP was discarded so that only 1ml remained in the tube along with the platelet button. This material was gently agitated to promote platelets resuspension and activated when added 0.3ml of calcium gluconate, resulting in PRP gel. Double centrifugation protocol was able to make platelet concentration 3 times higher in relation to the initial blood sample. The volume of calcium gluconate used for platelet activation was 0.3ml, and was sufficient to coagulate the sample. Coagulation time ranged from 8 to 20 minutes, with an average of 17.6 minutes. Therefore, time of blood centrifugation until to obtain PRP gel took only 40 minutes. It was concluded that PRP was successfully obtained by double centrifugation protocol, which is able to increase the platelet concentration in the sample compared with whole blood, allowing its use in surgical procedures. Furthermore, the preparation time is appropriate to obtain PRP in just 40 minutes, and calcium gluconate is able to promote the activation of platelets.

scholarly journals Effectiveness of Two Methods for Preparation of Autologous Platelet-Rich Plasma: An Experimental Study in Rabbits

2010 ◽  
Vol 04 (04) ◽  
pp. 395-402 ◽  
Author(s):  
Maria J. H. Nagata ◽  
Michel R. Messora ◽  
Flávia A. C. Furlaneto ◽  
Stephen E. Fucini ◽  
Alvaro F. Bosco ◽  
...  
Keyword(s):  
Platelet Count ◽  
Whole Blood ◽  
Repeated Measures ◽  
Platelet Rich Plasma ◽  
Blood Platelet ◽  
Protocol Group ◽  
Group I ◽  
Group Ii ◽  
Processing Errors ◽  
Centrifugation Protocol

Objectives: The purpose of this study was to compare the quantity and quality of platelets in platelet- rich plasma (PRP) samples prepared using either the single- or the double-centrifugation protocol.Methods: Ten adult white New Zealand rabbits were used. Ten ml of blood were drawn from each animal via cardiac puncture. Each blood sample was divided into two equal parts for PRP preparation: 5 ml of blood were centrifuged according to a single-centrifugation protocol (Group I), and 5 ml were centrifuged according to a double-centrifugation protocol (Group II). Manual platelet counts were performed on the whole blood and PRP samples of each group. Smears were also done on all samples in order to see the morphology of the platelets. The data obtained in the manual platelet count were submitted to statistical analysis (repeated measures ANOVA, Tukey, P<.05).Results: The average whole blood platelet count was 446,389/μl. The PRP samples in Group II presented an average platelet amount significantly higher than that of Group I (1,986,875 ± 685,020/μl and 781,875 ± 217,693/μl, respectively). The PRP smears from Group II were the only one to present platelets with altered morphology (75% of the smears). A few lymphocytes with increased cytoplasm were observed in the PRP smears of both Groups I (25% of the smears) and II (62.5% of the smears).Conclusions: Within the limits of this study, it can be concluded that the double-centrifugation protocol resulted in higher platelet concentrations than did the single-centrifugation protocol. However, the double-centrifugation protocol caused alterations in platelet morphology and was more sensitive to small processing errors. (Eur J Dent 2010;4:395-402)

scholarly journals The Effect of Platelet-Rich Plasma on Elbow Tendinopathies: A Systematic Review

2013 ◽  
Author(s):  
Sarah Schwetlik ◽  
Luke Strempel
Keyword(s):  
Systematic Review ◽  
Cohort Study ◽  
Physical Function ◽  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Autologous Blood ◽  
Sufficient Evidence ◽  
Quantitative Studies ◽  
And Function

Purpose: Elbow tendinopathies are common conditions that typically last 6 to 24 months. There is no clear consensus in the literature regarding the most effective management. Platelet-rich plasma (PRP) is an autologous blood product used for elbow tendinopathies with the aim of enhancing tissue regeneration. The aim of this systematic review was to evaluate the available evidence on the effectiveness and safety of PRP for reducing pain and physical function in elbow tendinopathies. Methods: Electronic databases were searched for relevant studies and data were extracted regarding the design, sample characteristics, interventions, and outcome measures. Each study was critically appraised for methodological quality using a modified tool for quantitative studies and presented in a narrative summary. Results: The search strategy identified 299 hits related to platelet-rich plasma and/or elbow tendinopathies. Five studies met the inclusion criteria; all were randomized controlled trials except one cohort study. All five studies showed improvements from baseline in pain and physical function with a PRP intervention. One study and its follow-up study showed significant improvements in pain and function with PRP compared to corticosteroid at 26, 52, and 104 weeks. Two studies compared PRP to whole blood, which did not find sufficient evidence to suggest one is more effective than the other. A cohort study found PRP was more effective than placebo at 4 and 8 weeks. Three studies reported on the safety of PRP and found no significant adverse effects. Conclusions: The current literature has some limitations and is insufficient to provide strong recommendations regarding the use of PRP in elbow tendinopathies over other modalities; however, these studies suggest that PRP may be more efficacious than corticosteroid injections, but that whole blood injections may be as effective as PRP.

Stability of antioxidant vitamins in whole human blood during overnight storage at 4°C and frozen storage up to 6 months

2018 ◽  
Vol 88 (3-4) ◽  
pp. 151-157 ◽  
Author(s):  
Scott W. Leonard ◽  
Gerd Bobe ◽  
Maret G. Traber
Keyword(s):  
Ascorbic Acid ◽  
Whole Blood ◽  
Healthy Subjects ◽  
Frozen Storage ◽  
Blood Collection ◽  
Plasma Samples ◽  
Optimal Conditions ◽  
Plasma Ascorbic Acid ◽  
Blood Samples ◽  
Sample Handling

Abstract. To determine optimal conditions for blood collection during clinical trials, where sample handling logistics might preclude prompt separation of erythrocytes from plasma, healthy subjects (n=8, 6 M/2F) were recruited and non-fasting blood samples were collected into tubes containing different anticoagulants (ethylenediaminetetra-acetic acid (EDTA), Li-heparin or Na-heparin). We hypothesized that heparin, but not EDTA, would effectively protect plasma tocopherols, ascorbic acid, and vitamin E catabolites (α- and γ-CEHC) from oxidative damage. To test this hypothesis, one set of tubes was processed immediately and plasma samples were stored at −80°C, while the other set was stored at 4°C and processed the following morning (~30 hours) and analyzed, or the samples were analyzed after 6 months of storage. Plasma ascorbic acid, as measured using HPLC with electrochemical detection (LC-ECD) decreased by 75% with overnight storage using EDTA as an anticoagulant, but was unchanged when heparin was used. Neither time prior to processing, nor anticoagulant, had any significant effects upon plasma α- or γ-tocopherols or α- or γ-CEHC concentrations. α- and γ-tocopherol concentrations remained unchanged after 6 months of storage at −80°C, when measured using either LC-ECD or LC/mass spectrometry. Thus, refrigeration of whole blood at 4°C overnight does not change plasma α- or γ-tocopherol concentrations or their catabolites. Ascorbic acid is unstable in whole blood when EDTA is used as an anticoagulant, but when whole blood is collected with heparin, it can be stored overnight and subsequently processed.

Time Dependence of Whole Blood Aggregation in Response to Platelet Activating Factor (PAF)

1988 ◽  
Vol 59 (02) ◽  
pp. 162-163 ◽  
Author(s):  
R R Taylor ◽  
J Strophair ◽  
M Sturm ◽  
R Vandongen ◽  
L J Beilin
Keyword(s):  
Time Dependence ◽  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Platelet Activating Factor ◽  
Sampling Time ◽  
Blood Sampling ◽  
Impedance Aggregometry

SummaryThe aggregation/adhesion response to platelet activating factor (PAF) was studied in diluted whole blood by impedance aggregometry. The extent of aggregation varied directly with the interval between blood sampling and aggregation measurement over the first 30 minutes from sampling, then remained stable for the next 60 minutes of observation. This is an effect opposite to that described for aggregation to PAF in platelet rich plasma which, however, cannot be studied soon after sampling. Time dependence of aggregation is important and comparative measurements should be made during the period of stable aggregability.

Macroaggregation of Platelets in Plasma, as Distinct from Microaggregation in Whole Blood (and Plasma), as Determined Using Optical Aggregometry and Platelet Counting respectively, is Specifically Impaired following Cardiopulmonary Bypass in Man

1994 ◽  
Vol 72 (04) ◽  
pp. 511-518 ◽  
Author(s):  
Valentine C Menys ◽  
Philip R Belcher ◽  
Mark I M Noble ◽  
Rhys D Evans ◽  
George E Drossos ◽  
...  
Keyword(s):  
Cardiac Surgery ◽  
Platelet Count ◽  
Cardiopulmonary Bypass ◽  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Interquartile Range ◽  
Contributory Factor ◽  
Platelet Aggregability ◽  
Aggregate Growth

SummaryWe determined changes in platelet aggregability following cardiopulmonary bypass, using optical aggregometry to assess macroaggregation in platelet-rich plasma (PRP), and platelet counting to assess microaggregation both in whole blood and PRP. Hirudin was used as the anticoagulant to maintain normocalcaemia.Microaggregation (%, median and interquartile range) in blood stirred with collagen (0.6 µg/ml) was only marginally impaired following bypass (91 [88, 93] at 10 min postbypass v 95 (92, 96] prebypass; n = 22), whereas macroaggregation (amplitude of response; cm) in PRP stirred with collagen (1.0µg/ml) was markedly impaired (9.5 [8.0, 10.8], n = 41 v 13.4 [12.7,14.3], n = 10; p <0.0001). However, in PRP, despite impairment of macroaggregation (9.1 [8.5, 10.1], n = 12), microaggregation was near-maximal (93 [91, 94]), as in whole blood stirred with collagen. In contrast, in aspirin-treated patients (n = 14), both collagen-induced microaggregation in whole blood (49 [47, 52]) and macroaggregation in PRP (5.1 [3.8, 6.6]) were more markedly impaired, compared with control (both p <0.001).Similarly, in PRP, macroaggregation with ristocetin (1.5 mg/ml) was also impaired following bypass (9.4 [7.2, 10.7], n = 38 v 12.4 [10.0, 13.4]; p <0.0002, n = 20), but as found with collagen, despite impairment of macroaggregation (7.2 [3.5,10.9], n = 12), microaggregation was again near-maximal (96 [93,97]). The response to ristocetin was more markedly impared after bypass in succinylated gelatin (Gelo-fusine) treated patients (5.6 [2.8, 8.6], n = 17; p <0.005 v control), whereas the response to collagen was little different (9.3 v 9.5). In contrast to findings with collagen in aspirin-treated patients, the response to ristocetin was little different to that in controls (8.0 v 8.3). Impairment of macroaggregation with collagen or ristocetin did not correlate with the duration of bypass or the platelet count, indicating that haemodilution is not a contributory factor.In conclusion: (1) Macroaggregation in PRP, as determined using optical aggregometry, is specifically impaired following bypass, and this probably reflects impairment of the build-up of small aggregates into larger aggregates. (2) Impairment of aggregate growth and consolidation could contribute to the haemostatic defect following cardiac surgery.

Haemostatic Platelet Plug Formation in the Isolated Rabbit Mesenteric Preparation - An Analysis of Red Blood Cell Participation

1980 ◽  
Vol 44 (01) ◽  
pp. 006-008 ◽  
Author(s):  
D Bergqvist ◽  
K-E Arfors
Keyword(s):  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Adenosine Diphosphate ◽  
In Vitro Test ◽  
Pharmacological Agents ◽  
Vitro Test ◽  
Releasing Effect ◽  
Plug Formation

SummaryIn a model using an isolated rabbit mesenteric preparation microvessels were transected and the time until haemostatic plugs formed was registered. Perfusion of platelet rich plasma gave no haemostasis whereas whole blood did. Addition of chlorpromazine or adenosine to the whole blood significantly prolonged the time for haemostasis, and addition of ADP to the platelet rich plasma significantly shortened it. It is concluded that red cells are necessary for a normal haemostasis in this model, probably by a combination of a haemodynamic and ADP releasing effect.The fundamental role of platelets in haemostatic plug formation is unquestionable but there are still problems concerning the stimulus for this process to start. Three platelet aggregating substances have been discussed – thrombin, adenosine diphosphate (ADP) and collagen. Evidence speaking in favour of thrombin is, however, very minimal, and the discussion has to be focused on collagen and ADP. In an in vitro system using polyethylene tubings we have shown that "haemostasis" can be obtained without the presence of collagen but against these results can be argued that it is only another in vitro test for platelet aggregation (1).To be able to induce haemostasis in this model, however, the presence of red blood cells is necessary. To further study this problem we have developed a model where haemostatic plug formation can be studied in the isolated rabbit mesentery and we have briefly reported on this (2).Thus, it is possible to perfuse the vessels with whole blood as well as with platelet rich plasma (PRP) and different pharmacological agents of importance.

Antiheparin Activity of Human Serum and Platelet Factor 4

1973 ◽  
Vol 30 (01) ◽  
pp. 093-105 ◽  
Author(s):  
C.H.J Sear ◽  
L Poller ◽  
F.R.C Path
Keyword(s):  
Molecular Weight ◽  
Ionic Strength ◽  
Blood Serum ◽  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Gel Filtration ◽  
Normal Serum ◽  
Platelet Factor ◽  
Mean Values ◽  
Antiheparin Activity

SummaryThe antiheparin activity of normal serum has been studied by comparing the antiheparin activities of sera obtained from normal whole blood, platelet-rich plasma and platelet-’free’ plasma with a purified platelet extract during differential isoelectric precipitation and by gel filtration chromatography.The mean values for the activity of PRP-serum and PFP-serum were 106% (S.D. 11) and 10% (S.D. 3) of untreated whole blood respectively. The activity of whole blood serum, PRP serum and whole blood serum plus platelet extract precipitated under identical physical conditions, i.e. pH 7.0, I =0.008, indicating that the activities of the three samples are probably associated with PF4. PF4 precipitated from human platelet extract at pH 4.0, but this is probably due to the difference in the two biochemical environments investigated, i.e. serum and platelet extract.The gel filtration experiments revealed striking similarities between the major antiheparin activities of serum and platelet extract. At physiological pH and ionic strength both activities were associated with high molecular weight material, but at physiological pH and elevated ionic strength both activities behaved as much smaller entities of molecular weight between 25,000 and 30,000 daltons and it seems very likely that both activities are associated with the same molecule, i.e. PF4.

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