摘要:

AbstractA new electrophoretic transfer system for the recovery of proteins focused in Immobiline matrices is described. The gel strip containing the sample zone of interest is transferred to a horizontal tray and embedded in 1%, low‐gelling (37°C) agarose. For acidic to neutral proteins (up to pI7.7) the electrophoretic transfer is from the IPG strip into a layer of DEAE‐Sephadex, buffered at pH 8.5 in 100 mM Tris‐acetate. Recovery (better than 90% in all cases studied) is achieved by titrating the resin at pH 9.5, in 200 mM Tris‐Gly buffer, containing 200 mM salt. For basic proteins (pI>7.7) the electrophoretic retrieval is from the IPG strip into a zone of CM‐Sephadex, buffered at pH 6.0, in 50 mM citrate (cathodic migration). Recovery (again better than 90%) is accomplished by titrating the exchanger at pH 4.0, in 200 mM formate buffer, containing 200 mM NaCl. Exposure to pH 9.5 does not affect enzyme activity if the eluate is promptly titrated around neutrality; a pH 4 milieu might irreversibly alter enzymes, in which case elution is best performed by titrating the protein to its pI, rather than protonating the exchanger. It has been demonstrated that Immobiline gels, even when incorporating 5 times the standard amount of buffer (75 mM Immobiline at pH = pK,i. e.50 mM buffering ion and 25 mM titrant) exhibit, under the electric field, negligible ion‐exchange properties, thus ensuring ideal behavior as supports for isoelect

ISSN:0173-0835    

DOI:10.1002/elps.1150060202

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1985

数据来源: WILEY

摘要:

AbstractProtein separation in borate‐mannitol pH gradients using a free‐flow electrophoretic apparatus was studied. The pH gradients covering 0.6–1 pH unit were created by dilution of mannitol solution in borax with distilled water. Linear concentration gradients of mannitol and borax, an almost linear pH gradient, and a linear conductivity gradient, increasing from the alkaline toward the acidic end of the pH gradient, were obtained. By varying the initial mannitol concentration in borax, the gradients can be formed within the pH range 4.5–9.2. Under the experimental conditions the gradients were rapidly changed due to the influence of the electric field. The character of changes was dependent on the properties of the membranes between the electrode compartments and separation chamber, on residence time, voltage and presence of salts in the gradients. Because of gradient instability, isoelectric focusing was not achieved under the experimental conditions, and the protein separation resulted from electrophoresis in the pH gradient. The results of the separation were dependent on the point of injection of the sample into the pH gradient: the best separation was achieved when the proteins were injected above their isoelectric

ISSN:0173-0835    

DOI:10.1002/elps.1150060203

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1985

数据来源: WILEY

摘要:

AbstractProtein separation was investigated in the free‐flow apparatus using pH gradients generated by a concentration gradient of boric acid in a solution of borax. The linear pH gradient covering the range pH 8.2–9.2 was obtained in a solution of 2.5 mM borax due to the formation of the logarithmic concentration gradient of 0–45 mM boric acid. These pH gradients lacked a conductivity gradient in contrast to borate mannitol pH gradients, described in the joint report. By addition of mannitol into the gradient, at constant concentration, the pH was shifted toward more acidic values in proportion to the amount of mannitol added, without changing the range and shape of the pH gradient. According to this procedure of gradient formation the pH gradients can be obtained within the range from pH 3.5 to 9.2. The gradients were rapidly decaying under the experimental conditions. Thus, isoelectric focusing of proteins could not be achieved, and the separation resulted from electrophoresis in the pH gradient. The results of protein separation in the gradients practically did not depend on the point of injection of the samples into the pH gradient, at least in the cases when the proteins were injected near their isoelectric p

ISSN:0173-0835    

DOI:10.1002/elps.1150060204

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1985

数据来源: WILEY

摘要:

AbstractThe omega function, defined for strong and monovalent weak electrolytes, is a useful regulating principle for the description of electrophoretic processes. The omega function profile, which is established by the initial electrolyte distribution in an electrophoretic column, will not be changed by current flow. The function is based solely upon migration and is only applicable to those regions where diffusional dispersion can be neglected. Computer simulations are used to follow the evolution of the concentration gradients of various electrophoretic systems and to verify the proposed constancy of the omega function profile. The limitations of the theory are investigated and the value of these predictions to the experimentalist is discussed.

ISSN:0173-0835    

DOI:10.1002/elps.1150060205

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1985

数据来源: WILEY

摘要:

AbstractAutomated cell electrophoretic measurements with Lazypher are based on the laser Doppler technique. Experiments are performed with high precision (of 1.2% S. D.) and in short time intervals of 1–3 min. Application to electrophoretic mobility tests with tanned sheep erythrocytes incubated in supernatants from sensitized murine thymus and spleen cells which had been prepared from cultures containing 5 μg purified protein derivative (PPD)/ml caused a reduction of the electrophoretic mobility (EM) of 5% and 13%, respectively. Control spleen cell supernatants caused a 4% decrease in mobility and no effect was observed with supernatants from control thymus cells. Peripheral blood lymphocytes from healthy adults (n = 56) and children with atopic diseases and congenital heart disease (n = 12) are separated into a high mobility peak (μ = 1.24 μmcm/Vs for adults, μ = 1.19 μmcm/Vs for the child group) corresponding to T‐cells and a low mobility peak, μ = 0.93 μcm/Vs, representing B‐cells. The mobility difference between the adult and child T‐cells is highly significant (99%). Electrophoretic measurements of myeloid cells from healthy donors and patients with chronic myeloid leukemia (CML) and acute myeloid leukemia (AML) show that progressing differentiation within this cell line is accompanied by a decreasing mobility. Blast cells from a patient with acute lymphoblastic leukemia (c‐ALL) had an EM, μ = 1.09 μmcm/Vs, distinct from that of myeloblasts in AML (μ = 1.16 and 1.25 μmcm/Vs) and CML (μ = 1.25 and 1.23 μmcm/Vs) and from norm

ISSN:0173-0835    

DOI:10.1002/elps.1150060206

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1985

数据来源: WILEY

摘要:

AbstractA three‐month study was conducted on blood stains to determine the reliability and viability of the transferrin (Tf) and plasminogen (PLG) genetic markers upon storage under varying conditions. Stains stored at room temperature (19–22 °C), 4 °C and −70 °C were tested at weekly intervals. Isoelectric focusing on agarose gels was followed by fixation and staining with Coomassie Brilliant Blue R‐250. Gels were subsequently treated with silver stain if band intensity was too faint for phenotyping. Stains stored at room temperature and 4 °C showed a gradual decrease in band intensity. By using silver stain, it was possible to phneotype blood stains stored for at least twice the length of time as compared to those stained with Coomassie Blue. This study demonstrates that isoelectric focusing and silver staining is reliable for determining Tf and PLG markers in forensic inv

ISSN:0173-0835    

DOI:10.1002/elps.1150060207

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1985

数据来源: WILEY

摘要:

AbstractThe conversion of a sulfydryl group of mercaptoacids to a thiuronium group by means of dimethylcynamide followed by electrophoresis on cellulose gel strips is suggested as a method for estimation of the number of carboxyl groups in the molecule and for separation of mono and polycarboxyl mercaptoacids. As a result of cyanamidation the molecule gains an additional positive charge reflected in a shift of pH‐mobility curves along the mobility axe

ISSN:0173-0835    

DOI:10.1002/elps.1150060208

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1985

数据来源: WILEY

摘要:

AbstractA heating‐cooling plate for open casting of agarose gels is described. Use of this plate provides a flawless and convenient means of producing 0.8 mm or thicker gels of any size up to 25 × 30 cm on glass plates (0.5 mm thick gels tend to pull slightly away from the edge of the glass plate during pouring). Gels of of any thickness (0.5 mm or greater) can be prepared if GelBond film is us

ISSN:0173-0835    

DOI:10.1002/elps.1150060209

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1985

数据来源: WILEY

摘要:

AbstractAn ultrathin‐layer polyacrylamide gel isoelectric focusing method is described for subtyping transferrin. After a one‐hour pretreatment at room temperature of transferrin in serum with ferrous ammonium sulfate, the samples were subjected to isoelectric focusing for 140 min with a final voltage of 3500 V. Thus, transferrin subtyping was achieved in one‐half of a working day. Population data on transferrin in Black and White samples from Baltimore, Maryland, were obtained using this method. The data were similar to that previously reported on these races from other geographical loca

ISSN:0173-0835    

DOI:10.1002/elps.1150060210

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1985

数据来源: WILEY

ISSN:0173-0835    

DOI:10.1002/elps.1150060212

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1985

数据来源: WILEY