A Protocol for the Preparation of Cryoprecipitate and Cryo-depleted Plasma for Proteomic Studies

Simpson, Richard J.; Greening, David W.

doi:10.1007/978-1-4939-7057-5_2

Cited by 14 publications

(17 citation statements)

References 19 publications

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“…The base for the formation of hydrogel scaffolds was a cryoprecipitate of blood plasma. It is known that such blood plasma cryoprecipitates contain fibrinogen, fibronectin, factor XIII, fibrinolysis inhibitors, and circulating molecules of cell adhesion in a considerably higher concentration than standard blood plasma [57,58]. Fibrinogen is the main biopolymer that takes part in the formation of a scaffold structure according to the method described by ourselves.…”

Section: Discussionmentioning

confidence: 99%

Hydrogel scaffolds based on blood plasma cryoprecipitate and collagen derived from various sources: Structural, mechanical and biological characteristics

Egorikhina

Aleynik

Rubtsova

et al. 2019

Bioactive Materials

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At present there is a growing need for tissue engineering products, including the products of scaffold-technologies. Biopolymer hydrogel scaffolds have a number of advantages and are increasingly being used to provide means of cell transfer for therapeutic treatments and for inducing tissue regeneration. This work presents original hydrogel biopolymer scaffolds based on a blood plasma cryoprecipitate and collagen and formed under conditions of enzymatic hydrolysis. Two differently originated collagens were used for the scaffold formation. During this work the structural and mechanical characteristics of the scaffold were studied. It was found that, depending on the origin of collagen, scaffolds possess differences in their structural and mechanical characteristics. Both types of hydrogel scaffolds have good biocompatibility and provide conditions that maintain the three-dimensional growth of adipose tissue stem cells. Hence, scaffolds based on such a blood plasma cryoprecipitate and collagen have good prospects as cell carriers and can be widely used in regenerative medicine.

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Section: Discussionmentioning

confidence: 99%

Hydrogel scaffolds based on blood plasma cryoprecipitate and collagen derived from various sources: Structural, mechanical and biological characteristics

Egorikhina

Aleynik

Rubtsova

et al. 2019

Bioactive Materials

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“…The workflow for the preparation of PFP prior to EV isolation is shown in Figure 3A. To ensure removal of platelet‐derived EVs, blood collection was performed in sRNA tubes and blood was initially centrifuged with cryoprecipitates removed [62]. For ExoQuick™ ULTRA, thrombin treatment was performed prior to psEV isolation as recommended by the manufacturer.…”

Section: Resultsmentioning

confidence: 99%

“…EVs were isolated from 1 mL of fresh or frozen PFP collected in EDTA or sRNA tubes. If frozen PFP was utilized, samples were quickly thawed in a water bath or dry heating system at 37°C, followed by centrifugation at 5000 x g for 15 min (4°C) to remove cryoprecipitates [62]. plEV were isolated as described in Section 2.3 above, while psEV were isolated using three different commercial methods following manufacturer's instructions.…”

Section: Methodsmentioning

confidence: 99%

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Human myeloma cell‐ and plasma‐derived extracellular vesicles contribute to functional regulation of stromal cells

Reale

Carmichael

et al. 2021

Proteomics

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Circulating small extracellular vesicles (sEV) represent promising non-invasive biomarkers that may aid in the diagnosis and risk-stratification of multiple myeloma (MM), an incurable blood cancer. Here, we comprehensively isolated and characterized sEV from human MM cell lines (HMCL) and patient-derived plasma (psEV) by specific EV-marker enrichment and morphology. Importantly, we demonstrate that HMCL-sEV are readily internalised by stromal cells to functionally modulate proliferation. psEV were isolated using various commercial approaches and pre-analytical conditions (collection tube types, storage conditions) assessed for sEV yield and marker enrichment. Functionally, MM-psEV was shown to regulate stromal cell proliferation and migration. In turn, pre-educated stromal cells favour HMCL adhesion. psEV isolated from patients with both pre-malignant plasma cell disorders (monoclonal gammopathy of undetermined significance [MGUS]; smouldering MM [SMM]) and MM have a similar ability to promote cell migration and adhesion, suggesting a role for both malignant and pre-malignant sEV in disease progression. Proteomic profiling of MM-psEV (305 proteins) revealed enrichment of oncogenic factors implicated in cell migration and adhesion, in comparison to non-disease psEV. This study describes a protocol to generate

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“…The first study was conducted on the preparation of thrombin using Method M 1.0 and M 2.0 to compare the presence and absence of RVV-X in the preparation process. Thrombin was prepared from CDP (or cryo-supernatant), which is rich in prothrombin complex and has a low level of fibrinogen [25,26], by the procedure described in 2.3. From starting volume 100 ml of CDP, we obtained about 25 ml of thrombin solution in the preparation batch.…”

Section: Rvv-xmentioning

confidence: 99%

Study of Optimum Condition for Rapid Preparation of Thrombin using Russell’s Viper Venom Factor X Activator

Kijkriengkraikul¹,

Nuchprayoon²

2018

TOBIOTJ

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Background: The purpose of this study is to investigate a simple method with the optimum condition for rapid thrombin preparation from Cryoprecipitate-depleted Plasma (CDP) using RVV-X in the process. Methods: Thrombin preparation from human CDP was studied with the presence of different factors in batch condition including: 1) RVV-X; 2) volume of calcium chloride solution; 3) volume of sodium chloride solution for final extraction; and 4) incubation time. The properties of the prepared sample were analyzed for fibrin clot formation, total protein by Kjeldahl method, thrombin time, molecular weight and protein patterns by SDS-PAGE, and thrombin concentration by coagulation analyzer. The method and process of preparing thrombin and the study of optimum condition for rapidly preparing the highest yield of thrombin from starting CDP 100 ml were introduced. Results: The best four conditions were concluded: 1) RVV-X 50 mcg should be present in the process; 2) volume of 0.25 M calcium chloride should be 3 ml; 3) volume of 0.85% sodium chloride for the final protein precipitate extraction should be 10 ml and; 4) no incubation time needed for prothrombin activation process. A solution prepared from the optimum condition showed an obvious band on SDS-PAGE at a molecular weight about 36,000 Da which is our target protein thrombin. The prepared solution had a total protein content of 0.065 g/dl and gave satisfactory results of thrombin time (9 seconds) and fibrin clot formation. The test results of thrombin concentration between the method with and without incubation time were 269.4 and 295.2 IU/ml, respectively. Conclusion: This result showed that the method with RVV-X but without incubation time for prothrombin activation (optimum condition) gave the highest yield of thrombin.

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A Protocol for the Preparation of Cryoprecipitate and Cryo-depleted Plasma for Proteomic Studies

Cited by 14 publications

References 19 publications

Hydrogel scaffolds based on blood plasma cryoprecipitate and collagen derived from various sources: Structural, mechanical and biological characteristics

Hydrogel scaffolds based on blood plasma cryoprecipitate and collagen derived from various sources: Structural, mechanical and biological characteristics

Human myeloma cell‐ and plasma‐derived extracellular vesicles contribute to functional regulation of stromal cells

Study of Optimum Condition for Rapid Preparation of Thrombin using Russell’s Viper Venom Factor X Activator

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