摘要:

AbstractStudying the separation behavior of various native carbonic anhydrase isozymes from mammalian erythrocytes we found that the migration of these enzymes differs from that of the marker proteins commonly used in gradient gel electrophoresis. In alkaline buffer systems the enzymes from human, bovine, rabbit, and canine erythrocytes start to migrate with a size apparently 6 to 12 times larger than their monomeric size, then gradually lose in apparent size and finally end up in a size equivalent to their monomeric mol mass. We determined the monomeric mol mass of the various carbonic anhydrase forms to be 23 000 to 39 000 (g/mol). These values are in accordance with different data in the literature and the data which we obtained by sodium dodecyl sulfate‐gradient gel electrophoresis. The equations and the graphical solutions needed to calculate the apparent upper size as well as the monomeric size of mammalian carbonic anhydrases are presented. It is demonstrated by this example that only the time‐dependent version of non‐denaturing polyacrylamide gradient gel electrophoresis provides reliable mol mass estima

ISSN:0173-0835    

DOI:10.1002/elps.1150060402

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

年代:1985

数据来源: WILEY

摘要:

AbstractThe effect of protein complexing on quantitation by rocket immunoelectrophoresis was studied using group‐specific component (Gc), a serum protein which undergoes 1:1 interactions with both sterols (Vitamin D metabolites) and protein (G‐actin). Gc purified from serum was used as a reference. Complex formation with 25‐〈OH〉 D3 did not significantly affect the measurement of Gc. However, addition of excess G‐actin to purified Gc led to substantially higher rockets at every concentration of Gc tested. In dose‐response experiments performed with a constant quantity of Gc and increasing quantities of added G‐actin, the increment in rocket height was progressive up to equimolarity, at which point a plateau was reached. Further demonstration that the increased rocket heights were directly related to 1:1 complex formation between Gc and G‐actin (Gc:G‐actin) was obtained by [125I]G‐acting and isoelectric focusing (IEF) with autoradiography. Examination of the relative charge of Gc and of Gc:G‐actin by crossed immunoelectrophoresis showed that complexes exhibited faster mobility, whereas very little evidence of alteration in antigenicity of Gc:G‐actin complexes was apparent. These results suggest that Gc:G‐actin complexes exhibit increased electrophoretic mobility and that their presence in samples containing Gc causes an increase in the height of Gc rockets. The findings also indicate that interactions with specific ligands could cause artefacts in immunoelectrophorestic quantitation of other proteins p

ISSN:0173-0835    

DOI:10.1002/elps.1150060403

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

年代:1985

数据来源: WILEY

摘要:

AbstractThe stability with time of Immobiline solutions and of pre‐cast Immobiline gels has been investigated, for the former by high pressure liquid chromatography and for the latter by focusing a set of marker proteins. Temperature seems to affect these chemicals most: whatever degradation occurs at 20°C within a few weeks' time, is operative at 60°C within a few hours' time. These buffers are altered by pH in different ways: acidic Immobilines (pKs 3.6 and 4.6) are extensively degraded around neutrality (pH 6) while basic species (especially pK's 8.5 and 9.3) are massively destroyed in alkaline solutions (pH 9). Acidic pH values (pH 3–4) ensure maximum stability for all Immobiline chemicals. In terms of medium‐term storage of pre‐cast gels, wet matrices exhibit least degradation if titrated to pH 4. Formic acid is the best titrant since it can be efficiently removed during the focusing step with formation of the narrowest salt‐front at the anode. Dry matrices have much longer stability (>2 months) but should be stored with traces of humidity to prevent the cracking and peeling‐off from the plastic support which occurs at zero rela

ISSN:0173-0835    

DOI:10.1002/elps.1150060404

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

年代:1985

数据来源: WILEY

摘要:

AbstractSeminal acid phosphatase (SAP) and vaginal acid phosphatase (VAP) bands, visualised by conventional isoelectric focusing have a wide pH range extending from approximately pH 4.0–6.0. Although a large proportion of these bands had similar isoelectric point (pI) values, the extremities of their respective ranges do not overlap. For SAP it was found that coincidental focusing tended not to occur withion a pH range of approxiamtely 4.3–5.2. The introduction of zwitterionic buffers to a conventional pH 4.0–6.0 Ampholine gradient produces a narrow pH gradient (4.3–5.2) in which semen‐specific SAP bands can be distinguished from VAP. This system has been used in an attempt to qualitatively identify SAP in the presence of VAP from a number of semen‐contaminated swabs. The interconvertibility of SAP into VAP has been demonstrated by neuraminidase treatment of SAP. When SAP was visualised on 15 cm gels containing pH 4–6 Ampholines it was observed that anodal bands faded and were converted into cathodal forms. Detailed examination of cathodal VAP and neuraminidase‐treated SAP was carried out on narrow pH gradients which enabled direct comparisons of individual isoenzymes. This provided confirmation that interconversion between the two forms was possible. Finally, a population survey of SAP patterns was carried out which failed to reveal any genet

ISSN:0173-0835    

DOI:10.1002/elps.1150060405

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

年代:1985

数据来源: WILEY

摘要:

AbstractEsterase‐19 (ES‐19) of the house mouse, following specific visualization in electrophoretic and isoelectric focusing gels, is described. A list of diagnostic features was compiled including substrate specificity, inhibition and activation characteristics, temporal and spatial patterns of expression, distribution among laboratory and wild mouse strains. ES‐19 was classified as arylesterase (E.C. 3.1.1.2) and compared with ES‐16 and ES‐25, two other mouse arylesterases. Homology with brain esterases of rat, guinea pig, rabbit and man was found, and a uniform disegnation “arylesterase I

ISSN:0173-0835    

DOI:10.1002/elps.1150060406

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

年代:1985

数据来源: WILEY

摘要:

AbstractNon‐histone proteins (NHP) extracted from chromatin of two plasmocytoma cell lines, X63‐Ag8‐653 and Sp2/O‐Ag 14, with 0.35 M NaCl are analyzed by two‐dimensional polyacrylamide gel electrophoresis. These NHP have been fractionated into a high mobility group (HMG) and a low mobility group (LMG) using 2 % trichloroacetic acid (TCA). The HMG which is well‐characterized is soluble in 2 % TCA whereas the LMG precipitates. LMG resolubilization is difficult. Only a buffer containing glycerol, sodium dodecyl sulfate (SDS) and β‐mercaptoethanol enables them to be studied by polyacrylamide SDS electrophoresis. The LMG is clearly differentiated from the HMG by the predominance of high molecular weight proteins and by the high number of molecular species in a wide pH range. In the LMG there are also some diffrences between the two plasmocytoma cell lines. For example, a high molecular weight protein of 100 000 and pI7.2–8.2 is present in Sp2/O‐Ag 14 but absent in X63‐Ag8–653. Also noticeable in the LMG proteins is the presence of low molecular weight proteins with high el

ISSN:0173-0835    

DOI:10.1002/elps.1150060407

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

年代:1985

数据来源: WILEY

摘要:

AbstractPreviously reported anomalous results obtained on the separation of the factor XIIIB (F XIIIB*2) allele product have been reinvestigated by thinlayer polyacrylamide gel isoelectric focusing (IEF). The ordinary IEF fails to resolve the difference between the products of F XIIIB*1 and F XIIIB*2 alleles. However, IEF in polyacrylamide gel in the presence of 6 M urea has been found to be a useful method of differentiation between these two allele products.

ISSN:0173-0835    

DOI:10.1002/elps.1150060408

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

年代:1985

数据来源: WILEY

摘要:

AbstractAmino acid analysis shows that commercial grades of agarose contain peptides. The quantity varies with the purity of the agarose. These findings agree with our observations that the extent of the background after silver‐staining for agarose gels or where agarose was used in the system (as in two‐dimensional polyacrylamide gel electrophoresis) was determined mainly by the quality of agarose u

ISSN:0173-0835    

DOI:10.1002/elps.1150060409

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

年代:1985

数据来源: WILEY

摘要:

AbstractThe formation of anionic complexes with chelating ligands like ethylenediaminetetraacetic acid or nitrilotriacetic acid is useful for the anionic isotachophoretic determination of selected metals. A zone comprising such a metal complex may decompose gradually under the influence of the electric current. The kinetic behavior of various complex zones is determined in a capillary‐type apparatus with a linear array of 255 potential gradient sensors. The discussed examples comprise both systems where complex formation occurred in the sample solution prior to its injection into the isotachophoretic column and where anin situformation of the complex took place under current flo

ISSN:0173-0835    

DOI:10.1002/elps.1150060410

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

年代:1985

数据来源: WILEY

摘要:

AbstractWe have devised methodology which allows for the rapid mechanical fixation of proteins in polyacrylamide slab gels. Proteins so fixed remain in the gel even after four days of soaking. The potential utility of this approach is discussed.

ISSN:0173-0835    

DOI:10.1002/elps.1150060411

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

年代:1985

数据来源: WILEY