Non-coating method for non-adherent cell culture using high molecular weight dextran sulfate and bovine serum albumin

Blood trypomastigotes of Trypanosoma cruzi were isolated from infected athymic rnu/rnu rats and purified by an improved procedure of DEAE-Sephacel ion-exchange chromatography. Elution into a buffer supplemented with bovine serum albumin avoided column-induced changes on the surface of the parasites. Biotin-labelled bovine serum albumin, fluorescence microscopy, flow cytometry and Western blot analysis revealed a very intense binding of albumin to the parasite. Incubation and washing of cells without protein supplementation did not result in any damage or lysis of parasites but it did cause extensive shedding of cellular and surface proteins into the supernatant which could be prevented by using the protein-supplemented buffer. A decreasing yield of high molecular weight cellular proteins in relation to centrifugal force was a general phenomenon observed in scanning densitometry of SDS gels after isolation in either protein-supplemented buffer or protein-free buffer. The quantity of shed cellular components increased with increasing centrifugal force. In contrast, quantities of high molecular weight, biotin-labelled surface proteins increased with greater centrifugal force, indicating labelling of otherwise inaccessible residues. These data emphasize the importance of protein supplementation of buffers with proteins and of choosing low centrifugation forces (<400 g) during investigations of T. cruzi.

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Effect of surfaces on fluid-phase prekallikrein activation

Blood ◽

10.1182/blood.v57.3.553.bloodjournal573553 ◽

1981 ◽

Vol 57 (3) ◽

pp. 553-560

Author(s):

CF Scott ◽

EP Kirby ◽

PK Schick ◽

RW Colman

Keyword(s):

Molecular Weight ◽

Bovine Serum Albumin ◽

Serum Albumin ◽

High Molecular Weight ◽

Bovine Serum ◽

Plasma Proteins ◽

Fluid Phase ◽

Factor Xii ◽

Cold Insoluble Globulin ◽

Kallikrein Activity

The activation of prekallikrein by factor XII fragments (XIIf), during incubation in plastic tubes was previously noted to be increased by high molecular weight (HMW) kininogen as well as other plasma proteins. In this report, we investigated the mechanism responsible for this increase. Although we confirmed that HMW kininogen, bovine serum albumin, fibrinogen, cold insoluble globulin, and mixed phospholipids apparently increased prekallikrein activation, we found that the product of prekallikrein activation (kallikrein) lost substantial activity in less than 0.5 min after exposure to a variety of fresh surfaces. This loss was partially prevented by the presence of various proteins and phospholipids. Similar protection against inactivation of XIIf, the enzyme in this reaction, was also found. In contrast, no loss of the substrate, prekallikrein, was observed during incubation. The loss of kallikrein activity was found to be proportional to the surface area of the incubation vessel as well as the concentration of kallikrein. Further loss of kallikrein activity could also be prevented by pretreating the vessel with kallikrein. We therefore conclude that various substances apparently affect prekallikrein activation in a purified system by preventing the enzyme and product in the reaction mixture from losing activity due to adsorption to a surface.

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Natural killer cell-dependent immunoglobulin G2a anti-bovine serum albumin (BSA) response elicited by high molecular weight dextran-BSA conjugates associated with dextran-mediated macrophage-natural killer cell interaction

Immunology ◽

10.1046/j.1365-2567.2000.00135.x ◽

2000 ◽

Vol 101 (4) ◽

pp. 474-483

Author(s):

C. P. Ediriwickrema ◽

S. L. Tonkonogy ◽

B. Hammerberg

Keyword(s):

Molecular Weight ◽

Bovine Serum Albumin ◽

Natural Killer Cell ◽

Serum Albumin ◽

Natural Killer ◽

High Molecular Weight ◽

Bovine Serum ◽

Killer Cell ◽

Cell Interaction

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Effect of surfaces on fluid-phase prekallikrein activation

Blood ◽

10.1182/blood.v57.3.553.553 ◽

1981 ◽

Vol 57 (3) ◽

pp. 553-560 ◽

Cited By ~ 9

Author(s):

CF Scott ◽

EP Kirby ◽

PK Schick ◽

RW Colman

Keyword(s):

Molecular Weight ◽

Bovine Serum Albumin ◽

Serum Albumin ◽

High Molecular Weight ◽

Bovine Serum ◽

Plasma Proteins ◽

Fluid Phase ◽

Factor Xii ◽

Cold Insoluble Globulin ◽

Kallikrein Activity

Abstract The activation of prekallikrein by factor XII fragments (XIIf), during incubation in plastic tubes was previously noted to be increased by high molecular weight (HMW) kininogen as well as other plasma proteins. In this report, we investigated the mechanism responsible for this increase. Although we confirmed that HMW kininogen, bovine serum albumin, fibrinogen, cold insoluble globulin, and mixed phospholipids apparently increased prekallikrein activation, we found that the product of prekallikrein activation (kallikrein) lost substantial activity in less than 0.5 min after exposure to a variety of fresh surfaces. This loss was partially prevented by the presence of various proteins and phospholipids. Similar protection against inactivation of XIIf, the enzyme in this reaction, was also found. In contrast, no loss of the substrate, prekallikrein, was observed during incubation. The loss of kallikrein activity was found to be proportional to the surface area of the incubation vessel as well as the concentration of kallikrein. Further loss of kallikrein activity could also be prevented by pretreating the vessel with kallikrein. We therefore conclude that various substances apparently affect prekallikrein activation in a purified system by preventing the enzyme and product in the reaction mixture from losing activity due to adsorption to a surface.

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Improvement QCM Quality By Polystyrene Coating and Bovine Serum Albumin as Immobilization Agent

Jurnal Ilmiah Pendidikan Fisika Al-Biruni ◽

10.24042/jipfalbiruni.v8i1.3716 ◽

2019 ◽

Vol 8 (1) ◽

pp. 35-41 ◽

Cited By ~ 1

Author(s):

Lalu A. Didik ◽

Yahdi Yahdi ◽

Masruroh Masruroh

Keyword(s):

Thin Film ◽

Bovine Serum Albumin ◽

Serum Albumin ◽

Spin Coating ◽

Bovine Serum ◽

Quartz Crystal ◽

Measuring System ◽

Frequency Shifts ◽

Impedance Analyzer ◽

Coating Method

Polystyrene (PS) thin film with various concentration of mass (1%, 3%, and 5%) has been deposited onto Quartz Crystal Microbalance (QCM) surface by a spin coating method. The purpose of this research is the improvement of Bovine Serum Albumin (BSA) immobilization into the QCM surface. The QCM resonance frequency has been investigated by means of Impedance Analyzer. The film thickness was calculated by the Sauerbrey equation. The surface roughness of the PS layer was observed by noncontact Topography Measuring System (TMS-1200). Calculation result using a Sauerbrey equation shows the thickness of the PS layer increases with the increasing PS concentration. The QCM/PS with 1% and 5% concentration of mass has rougher morphology and more frequency shifts caused by BSA injection compared to 3% concentration of PS

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Elucidation of the Interaction between Flavan-3-ols and Bovine Serum Albumin and Its Effect on Their In-Vitro Cytotoxicity

Molecules ◽

10.3390/molecules24203667 ◽

2019 ◽

Vol 24 (20) ◽

pp. 3667

Author(s):

Yasuyuki Fujii ◽

Yoshitomo Suhara ◽

Yusuke Sukikara ◽

Tomohiro Teshima ◽

Yoshihisa Hirota ◽

...

Keyword(s):

Cell Culture ◽

Bovine Serum Albumin ◽

Serum Albumin ◽

Bovine Serum ◽

Fetal Bovine Serum ◽

In Vitro Cytotoxicity ◽

Binding Energies ◽

Docking Simulations ◽

Fetal Bovine

Flavan-3-ols (FLs), specifically catechin and its oligomer B-type procyanidins, are suggested to potently bind to bovine serum albumin (BSA). We examined the interaction between BSA and FLs by fluorescence quenching and found the following order of binding activities to BSA: cinnamtannin A2 (A2; tetramer) > procyanidin C1 (C1; trimer) ≈ procyanidin B2 (B2, dimer) > (−)epicatechin (EC, monomer). Docking simulations between BSA and each compound at the binding site showed that the calculated binding energies were consistent with the results of our experimental assay. FLs exerted cytotoxicity at 1000 μg/mL in F11 cell culture with fetal bovine serum containing BSA. In culture containing serum-free medium, FLs exhibited significant cell proliferation at 10−4 μg/mL and cytotoxicity was observed at concentrations greater than 10 μg/mL. Results of this study suggest that interactions between polyphenols and BSA should be taken into account when evaluating procyanidin in an in vitro cell culture system.

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