Thiophilic adsorption ‐ a new method for protein fractionation

Porath, Jerker; Maisano, Federico; Belew, Makonnen

doi:10.1016/0014-5793(85)80928-5

Cited by 247 publications

(78 citation statements)

References 5 publications

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“…"Thiophilic-interaction chromatography" (TIC) technique was introduced originally by Porath and his colleagues for the isolation of immunoglobulins [12]. TIC has been used successfully for characterization of human transferring [13], alzheimer's beta-amyloid peptides [14] and for purification of immunoglobulins from chicken sera [15].…”

Section: Introductionmentioning

confidence: 99%

“…This chromatographic step is based upon salt-promoted adsorption of proteins to the resin, where the binding of proteins to a sulfone and thio-ether-containing hetero-aliphatic ligand takes place mainly through accessible tryptophan and/or phenylalanine residues. The desorption of bound proteins is achieved when the salt concentration is reduced [12]. In our earlier studies, we have shown that PSA and PSA complexes have strong affinity for different thiophilic gels (T-gels), and that T-gel affinity can be successfully applied for the purification of PSA and PSA-complexes from biological fluids, including serum and seminal plasma [16,17].…”

Section: Introductionmentioning

confidence: 99%

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Thiophilic-interaction chromatography of enzymatically active tissue prostate-specific antigen (T-PSA) and its modulation by zinc ions

Babu

Vijayalakshmi

Smith

et al. 2008

Journal of Chromatography B

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Prostate-specific antigen (PSA) is a serine protease secreted both by normal prostate glandular epithelia cells and prostate cancer cells. We explored "thiophilic-interaction chromatography" (TIC) to isolate tissue prostate-specific antigen (T-PSA) from fresh human prostate cancer tissue harvested by radical prostatectomy for the purpose to characterize T-PSA for its enzymatic activity and sensitivity to zinc ions. We have shown, for the first time, that T-PSA has strong affinity for the thiophilic gel (T-gel). The average recovery of T-PSA from T-gel is over 87%. The presence of PSA in the column eluate was confirmed by ELISA and SDS/PAGE. Western blot developed with monoclonal antibody to PSA revealed that T-PSA was predominantly in the "free" form having a molecular weight of 33kDA. Furthermore, T-PSA was found to be enzymatically active. T-PSA was found to be less enzymatically active as compared to seminal plasma PSA. The inhibition of enzymatic activity of both f-PSA and T PSA over a wide range of concentrations of Zn 2+ ions (10 nM to 50 μM) was comparable. In contrast, the enzymatic activity of chymotrypsin, another serineprotease, was affected differently. At higher concentrations of Zn 2+ (10 μM and higher) the enzymatic activity of chymotrypsin was inhibited, whereas, at lower concentrations of Zn 2+ (5 μM and lower), the enzymatic activity was enhanced.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Thiophilic-interaction chromatography of enzymatically active tissue prostate-specific antigen (T-PSA) and its modulation by zinc ions

Babu

Vijayalakshmi

Smith

et al. 2008

Journal of Chromatography B

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“…Serum was adjusted to pH 5.5 by adding one volume of 0.5 M sodium acetate (pH 5.5), and it was salted out by adding ammonium sulfate to 1.95 M. After 2 h at room temperature, the supernatant was recovered by centrifugation (4,000 ϫ g for 20 min), and the precipitate was washed with 1.95 M ammonium sulfate. The combined supernatants were dialysed extensively against 1.75 M ammonium sulfate at 4°C and applied to a thiophilic adsorbent (Affi-T agarose; Kem En Tec, Copenhagen) equilibrated in 1.75 M ammonium sulfate (25,26). By using a flow of Ϸ0.5 ml͞min, the adsorbent was washed to its baseline with the same buffer and eluted with 1.5 M ammonium sulfate, followed by 0.1 M NaCl.…”

Section: Methodsmentioning

confidence: 99%

Serpina1 is a potent inhibitor of IL-8-induced hematopoietic stem cell mobilization

Pel

Velders

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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Here, we report that cytokine-induced (granulocyte colony-stimulating factor and IL-8) hematopoietic stem cell (HSC) and hematopoietic progenitor cell (HPC) mobilization is completely inhibited after low-dose (0.5 Gy) total-body irradiation (TBI). Because neutrophil granular proteases are regulatory mediators in cytokineinduced HSC͞HPC mobilization, we considered a possible role for protease inhibitors in the induction of HSC͞HPC mobilization. Bone marrow (BM) extracellular extracts that were obtained from murine femurs after 0.5 Gy of TBI contained an inhibitor of elastase. Also, after low-dose TBI, both Serpina1 mRNA and protein concentrations were increased in BM extracts, compared with extracts that were obtained from controls. The inhibitory activity in BM extracts of irradiated mice was reversed by addition of an Ab directed against Serpina1. To further study a possible in vivo role of Serpina1 in HSC͞HPC mobilization, we administered Serpina1 before IL-8 injection. This administration resulted in an almost complete inhibition of HSC͞HPC mobilization, whereas heat-inactivated Serpina1 had no effect. These results indicate that low-dose TBI inhibits cytokine-induced HSC͞HPC mobilization and induces Serpina1 in the BM. Because exogenous administration of Serpina1 inhibits mobilization, we propose that radiation-induced Serpina1 is responsible for the inhibition of HSC͞HPC mobilization. Also, we hypothesize that cytokine-induced HSC͞HPC mobilization is determined by a critical balance between serine proteases and serine protease inhibitors.protease ͉ protease inhibitors ͉ bone marrow ͉ adhesion molecules

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“…Activity of factor II, V, VIII and X was determined with the help of coagulometric methods and Helena BioSciences reagents. Coagulation system ex vivo studies were performed in normal human plasma after addition of lyophilized monoclonal IgG, isolated with the help of thiophilic adsorption method from 20 ml of the patient's blood collected at the time admission [8][9][10]. Thereafter, the activity of factor II, factor V and aPTT, PT and TT in the prepared plasma samples (a series of dilutions of monoclonal IgG in standard human plasma at final concentration of 0, 0.468, 0.937, 1.875, 3.75, 7.5, 15 and 30 g/l, respectively) were measured.…”

Section: Casementioning

confidence: 99%

IgG Monoclonal Immunoglobulin (M-protein) as Factor V Inhibitor in Multiple Myeloma Patient: Case Report and Discussion

Krzysztof¹,

Szczepaniak²

2016

J Clin Cell Immunol

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We present a case of a 53-year-old male with a symptomatic relapse of IgGλ multiple myeloma who was readmitted to the hospital with clinical symptoms of retroperitoneal hematoma. Laboratory tests revealed high concentration of monoclonal immunoglobulin G (32 g/l, 91.6% of total IgG) in the serum, prolongation of activated partial thromboplastin time (aPTT), prothrombin time (PT), and thrombin time (TT), significantly decreased factor V procoagulant activity and the presence of factor V inhibitor. After successful retreatment with the use of six cycles of bortezomib, adriamycin and dexamethasone, the monoclonal IgG concentration in the patient's serum significantly decreased (up to 1.3 g/l). At that time, also the normalization of the results of aPTT, PT and TT was stated. Therefore, the presence of factor V inhibitor in monoclonal immunoglobulin fraction was suspected. To confirm this, the monoclonal immunoglobulin G from the patient's initial blood sample was isolated, a series of dilutions of it in standard human plasma was prepared and the activity of factor II, factor V and aPTT, PT and TT in the prepared plasma samples were measured. The results of ex vivo studies matched those obtained in vivo, proving that the monoclonal immunoglobulin G produced by multiple myeloma cells acted as a factor V inhibitor.

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Thiophilic adsorption ‐ a new method for protein fractionation

Cited by 247 publications

References 5 publications

Thiophilic-interaction chromatography of enzymatically active tissue prostate-specific antigen (T-PSA) and its modulation by zinc ions

Thiophilic-interaction chromatography of enzymatically active tissue prostate-specific antigen (T-PSA) and its modulation by zinc ions

Serpina1 is a potent inhibitor of IL-8-induced hematopoietic stem cell mobilization

IgG Monoclonal Immunoglobulin (M-protein) as Factor V Inhibitor in Multiple Myeloma Patient: Case Report and Discussion

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